Background: Infections caused by E. coli cause considerable disease burden and range from frequently occurring and relatively innocent urinary tract infection (UTI) to severe bloodstream infection (BSI). The incidence of infections caused by ESBL-producing E. coli (ESBL-PEc) is increasing, justifying surveillance and development of preventive strategies in several domains. Faecal carriage is universal and believed to be the most important reservoir for E. coli from which infections can originate. It is currently unknown to what extent Dutch E. coli carriage strains in the community reflect isolates causing disease. In this study, we will perform comparative genomics to infer the population structures of human-derived ESBL-PEc from community- and hospital-acquired infections and from community-based faecal carriage samples in the Netherlands. Furthermore, we will describe the molecular epidemiology of E. coli isolates causing invasive disease (BSI). Methods: This study uses four different microbiological data sources: 1) ESBL-PEc from patients with community-acquired UTI tested in primary care between May and November 2017, 2) ESBL-PEc from urine cultures obtained from patients hospitalized between January 2014 and December 2016, 3) E. coli from blood cultures obtained from patients hospitalized between January 2014 and December 2016, and 4) ESBL-PEc from faecal samples collected in a national population- prevalence study performed between January 2014 and January 2017. Clinical epidemiological data was collected from all patients and all isolates were subjected to whole genome sequencing. Discussion: The EPIGENEC study (EPIdemiology and GENetics of E. coli) will describe the molecular epidemiology of E. coli BSI and assess the genomic population structure of ESBL-PEc strains from community-acquired and nosocomial infections, and of ESBL-PEc reflecting community-based faecal carriage. Information from these studies may assist in optimizing surveillance strategies and determining targets and potential impact of future new preventive measures.

Legionnaires’ disease is normally acquired by inhalation of legionellae from a contaminated environmental source. Water systems of large buildings, such as hospitals, are often contaminated with legionellae and therefore represent a potential risk for the hospital population. In this study, we demonstrated the constant presence of Legionella in water samples from the water system of a large university hospital in Messina (Sicily, Italy) consisting of 11 separate pavilions during a period of 15 years (2004-2018). In total, 1346 hot water samples were collected between January 2004 and December 2018. During this period, to recover Legionella spp. from water samples the standard procedures reported by the Italian Guidelines emanated in 2000 were adopted; from May 2015 to 2018 Italian Guidelines revised in 2015 (ISS, 2015), were used. The most water samples (72%) were positive to L. pneumophila serogroups 2-14 whereas L. pneumophila serogroup 1 accounted for the 18% and Legionella spp. for the 15%. Most of the positive samples were found in the buildings where are situated critical wards as ICU, Neurosurgery, Surgeries, Pneumology and Neonatal Intensive Unit Care. We highlighted the importance of a continuous monitoring of hospital water samples to prevent the potential risk of nosocomial legionellosis.

Mycobacteria are a large family of over 100 species, most of which do not cause diseases in humans. The majority of the mycobacterial species are referred to as nontuberculous mycobacteria (NTM), meaning they are not the causative agent of tuberculous (TB) or leprosy, i.e. Mycobacterium tuberculous complex and Mycobacterium leprae, respectively. The latter group is undoubtedly the most infamous, with TB infecting an estimated 10 million people and causing over 1.2 million deaths in 2017 alone (1). TB and leprosy also differ from NTM in that they are only transmitted from person to person and have no environmental reservoir, whereas NTM infections are commonly acquired from the environment (2) (3). It took until the 1950's for NTM to be recognised as a potential lung pathogen in people with underlying pulmonary disease and another 3 decades for NTM to be widely recognised by the medical community when NTM, particularly Mycobacterium avium complex (MAC) was recognised as the most common group of opportunistic pathogens in AIDS patients (4). This review focusses on an emerging NTM called Mycobacterium abscessus (M. abs). M. abs is a rapidly growing (RGM) NTM that is responsible for opportunistic pulmonary infections in patients with structural lung disorders such as cystic fibrosis (CF) and bronchiectasis (5), as well as a wide range of skin and soft tissue infections (SSTIs) in humans (6) (7). M. abs is a weakly staining Gram-positive mycobacterium that is neverand is, like other NTM, most often seen in soil and aquatic environments (8). The bacillus-shaped bacterium is 1-6µm long and 0.2-0.5µm in diameter, with curved ends and the presence of cord factor, or trehalose 6-6'-dimycolate, a glycolipid found in the cell wall of virulent species of mycobacteria that results in "serpentine cord" cell morphology is sometimes observed (8) (9). On solid growth medium, M. abs can display either a rough (M. abs-R) or smooth (M. abs-S) morphotype, with the rough morphotype displaying a more virulent phenotype than its smooth variant (10). The rough morphotype is characterised by irregular parallel filaments that form ridges across the colony, whereas a smooth morphology is displays a wet, smooth colony with no filaments or ridges (79). This morphology is driven by cell wall glycopeptidolipid (GPL); a loss of GPL results in the reversion from rough to smooth morphotype (80) (81). Moreover, it has been shown using human tissue culture models of infection that M. abs-R is able to persist and multiply within the host macrophage whereas M. abs-S lacks this capacity, hence its role in virulence (82). Like all other mycobacteria, M. abs are aerobic, non-motile and acid-fast organisms with a characteristically thick, lipid-rich cell wall that is hydrophobic. Due to their unusually impermeable, thick cell wall, mycobacteria are notoriously resistant to many antibiotics, disinfectants and heavy metals (11). When the genome of M. abs became available in 2009, elucidation of the resistance mechanisms of M. abs became an area of focus for scientific research, as the considerable threat it poses to public health became more apparent (12) (13) (14). In this review we will discuss how we came to understand the pathogen, how it is currently treated, as well as a discussion of drug resistance mechanisms and novel treatments currently in development.

The USDA-FSIS considers mechanically-tenderized beef as ‘non-intact’ and a food safety concern because of the potential for translocation of surface E. coli O157:H7 into the interior of the meat that may be cooked ‘rare or medium-rare’ and consumed. We evaluated 14 potential spray interventions on E. coli O157:H7-inoculated lean beef wafers (~10⁶ CFU/cm² , n=80) passing through a spray system (18 sec dwell time, ~40 PSI) integrated into the front end of a Ross TC-700MC tenderizer. Inoculated and processed beef wafers were stomached with D/E neutralizing broth and plated immediately, or were held in refrigerated storage for 1-, 7-, or 14 days prior to microbial plating. Seven antimicrobials that showed better performance in preliminary screening on beef wafers were selected for further testing on beef subprimals in conjunction with blade tenderization. Boneless top sirloin beef subprimals were inoculated at ~2 x 10⁴ CFU/cm² with a four-strain cocktail of Escherichia coli O157:H7 and passed once, lean side up, through an integrated spray system and blade tenderizer. Core samples obtained from each subprimal were examined for the presence/absence of E. coli O157:H7. Absence of E coli O157:H7 translocated into core samples correlated with the ability of the antimicrobials to reduce bacterial levels on the surface of beef prior to blade tenderization.

In many regions where drinking water supply is intermittent and unreliable, households adapt by storing water in cisterns or rooftop tanks. Both intermittent supply and stored water can be vulnerable to contamination by microorganisms with deleterious health effects. The Guadalajara Metropolitan Area is a rapidly growing urban center with over five million residents where household storage is nearly ubiquitous. This pilot study was conducted in July 2018 to examine the microbiological quality of drinking water in Guadalajara. Samples were tested for free available chlorine residual, total coliform bacteria, and E. coli. A survey on access to water and public perspectives was also conducted. Water exiting rooftop tanks exceeded regulatory limits for total coliform levels in half of the homes studied. Piped water arriving at two homes had total coliform levels that far exceeded regulatory limits. No E. coli were detected in any of the samples. Only 35% of homes had a chlorine residual between the recommended 0.2 and 1.5 mg/L. Many homes reported unpleasant odors and colors. Only 7% of residents drank the piped water. Future studies are needed, especially during April and May when many homes reported a higher disruption to water service.

As described in previous work, the use of synthetic chemical ingredients in modern cosmetics is postulated to be a cause of damage to the skin microbiome. The discovery that biodiversity on the human skin is currently the only reliable indicator of skin health, meant that for the first time, a mechanism to test for healthy skin was possible. Using this mechanism and in collaboration with The Medical University of Graz, who carried out the independent study, this work aimed to help answer whether modern day synthetic cosmetics are a main cause of long term damage to the skin microbiome. Thirty-two human participants tested three different face washes for their effect on the skin’s microbial diversity, along with skin pH, moisture and TEWL (trans-epidermal water loss), washing twice a day for four weeks. The upper volar forearm of the volunteers was swabbed at the beginning, two weeks in and end (four weeks). 16S rRNA sequencing was used. One leading ‘natural’ brand full of synthetic ingredients, a leading synthetic brand and a 100% natural face wash were used. Results give the first indications of a link between synthetic ingredients in a cosmetics product, and its effect on skin microbiome biodiversity. It paves the way for future studies on the topic with a larger sample group, longer test period and standardised methodology to create a universal standard for testing the health of skin using benchmark diversity values. This can be used in the future to test the effectiveness of cosmetics or ingredients on skin health, leading to the banning of products proven to harm the skin’s natural environment.

In this study we announce the draft genome sequence of a newly identified Acinetobacter species cross-reacting with E. coli serotype 0157:H7. The advent of Next-Generation technology has paved to way to discover new species which could otherwise be misidentified using conventional cultural and serotyping methods. The whole genome sequence of this isolate will help to identify potential marker/s of intervention and further genomic analysis might also shed light onto the virulence properties of this newly identified Acinetobacter species which has been provided the new name of Acinetobacter maqsudiensis.

The present study evaluates the bioremediation potential of indigenous bacterial species isolated from dye-contaminated soil samples from small dyeing outlet located in Ilorin. The water pollution index was estimated based on the physicochemical characteristics and heavy metal concentrations of the raw (Day 0) and treated textile wastewater such as pH, biochemical oxygen demand-5, chemical oxygen demand, total suspended solids and total dissolved solid with mean values of 8.85±0.45 mg/L, 1200±21.3 mg/L, 2440±31.3 mg/L, 1660±17.2 mg/L and 2650±28.1 mg/L respectively, similarly, Lead was the most abundant heavy metal detected in the sample while Cadmium concentration was the lowest with the mean values of 3.52±0.00 mg/L and 2.18±0.00 mg/L respectively. The bacterial strain with highest dye decolorization capacity was screened and identified as Bacillus licheniformis ZUL012.The isolate was consequently used for the bioremediation of the wastewater over a period of 10 days. The results showed an incredible reduction in the physiochemical characteristics and heavy metal concentrations of the textile wastewater in the following ranges (8.85-6.55), (1200-300) mg/L, (2440-518) mg/L, (1660-666) mg/L and (2650-920) mg/L with the highest removal efficiency of 75 %, 78 %, 60%, 65%, recorded for biochemical oxygen demand, chemical oxygen demand, total suspended solid, total dissolved solid, respectively while that of heavy metals such as lead, cadmium, chromium and nickel were 80 %, 60 %, 67 %, 72 % reduction, respectively. Laccase and Azoreductase activities tend to decrease as the pH gradually moved towards acidic condition during the bioremediation process. Toxicity of the treated effluent was assessed using Maize and Bean seed germination test. Conclusively, these research findings can serve as a framework for the outlet design of wastewater treatment plant for local textile outlets.

Drug resistance has become an extremely serious problem worldwide. Antibiotic resistance genes (ARGs) entering the environment with wastewaters promote replenishment of the resistome of natural microbioms. Distribution of several clinically significant ARGs in wastewaters of Rostov-on-Don (Southern Russia), lower reaches of the Don River and natural waters of the neighboring region was investigated. Metagenomic DNA samples isolated from 250 ml of wastewaters or natural waters and 200 mg of surface sediments were used for the study. Identification of the ARGs was carried out with end-point detection PCR. Presence of NDM, OXA-48, CTX-M, VanA, VanB, ErmB, and TetM/TetO genes was detected in urban wastewaters. Samples of wastewater treatment plant (WWTP) sewage were enriched with ARGs in contrast to non-treated wastewaters from the sewage collector. NDM, VanA, ErmB, TetM/TetO genes were found only in wastewaters and were absent in samples of natural waters and surface sediments. Only OXA-48, VanB and CTXM genes were found in natural waters and surface sediments. The described ARGs are quite typical for urban and hospital wastewaters. The target ARGs were detected in the samples connected to the anthropogenous sources of pollution such as Rostov municipal WWTP or livestock enterprise effluents.

This study evaluated the microbial safety of vended boiled, pasteurized and UHT milk sold in Nakawa, Kampala-Uganda. 15 milk samples were analyzed; 2 samples had Salmonella, 5 had S. aureus with a count of 1.66 0.02 log₁₀CFU/ml. E. coli was detected in 8 samples with 1.0 0.02 to 3.0 0.01log₁₀CFU/ml count. A high load of 3.0 CFU/ml was obtained in 3 samples with E. coli. Four E. coli positive samples had a contamination load of 2.0 0.015log₁₀ CFU/ml of which one was pasteurized milk. Only a pasteurized milk showed a low E. coli load at 1.0 0.02log₁₀ CFU/ml. All UHT milk had no microbial contamination. Both boiled and pasteurized milk had Salmonella, S. aureus and E. coli in levels above the set threshold limits. Milk consumers in Nakawa stand a potential public health risk of food poisoning reflected by presence of Salmonella, S. aureus and E. coli in some milk sold in the area.

The present study was conducted on the Ertec SU-1 sterilizer, a device designed to test sterilization processes of food samples using pulsed electric fields (PEF) of high intensity and for rapid sterilization of biological samples at low temperatures. The operation of the sterilizer is based on electro-perforation of the cell membrane. The aim of the paper is to show that the PEF method increases the concentration of polyphenolic compounds and decreases the antioxidant activity. Orange juice was used as a material to eliminate growth of microorganisms contained in it. The microorganisms were exposed to pulsed electric fields. The study results proved that a certain number of PEF pulses definitively eliminated microorganisms in the examined material.

Tuberculosis (TB) is the primary cause of death by a single infectious agent; responsible for around two million deaths in 2016. A major virulence factor of TB is the ability to enter a latent or Non-Replicating Persistent (NRP) state which is presumed untreatable. Approximately, 1.7 billion people are latently infected with TB and on reactivation many of these infections are drug resistant. As the current treatment is ineffective and diagnosis remains poor, millions of people have the potential to reactivate into active TB disease. The immune system seeks to control the TB infection by containing the bacteria in a granuloma, where it is exposed to stressful anaerobic and nutrient deprived conditions. It is thought to be these environmental conditions that trigger the NRP state. A number of in vitro models have been developed that mimic conditions within the granuloma to a lesser or greater extent. These different models have all been utilised for the research of different characteristics of NRP Mycobacterium tuberculosis, however their disparity in approach and physiological relevance often results in inconsistencies and a lack of consensus between studies. This review provides a summation of the different NRP models and a critical analysis of their respective advantages and disadvantages relating to their physiological relevance.

Natural spa springs are diffused all over the world and their use in pools is known since ancient times. This review underlines the cultural and social spa context focusing on hygiene issues, public health guidelines and emerging concerns regarding water management in wellness or recreational settings. The question of the "untouchability" of therapeutic natural waters and their incompatibility with traditional disinfection processes is addressed considering the demand for effective treatments that would respect the natural properties. Available strategies and innovative treatments are reviewed, highlighting potentials and limits for a sustainable management. Alternative approaches comprise nanotechnologies, photocatalysis systems, advanced filtration. State of the art and promising perspectives are reported considering the chemical-physical component and the biological natural complexity of the spa water microbiota.

Heavy metals are compounds that can be hazardous and impair growth of living organisms. Bacteria have evolved the capability not only to cope with heavy metals but also to detoxify polluted environments. Three heavy metal-resistant strains of Mucilaginibacer rubeus and one of Mucilaginibacter kameinonensis were isolated from the gold/copper Zijin mining site, Longyan, Fujian, China. These strains were shown to exhibit high resistance to heavy metals with minimal inhibitory concentration reaching up to 3.5 mM Cu(II), 21 mM Zn(II), 1.2 mM Cd(II), and 10.0 mM As(III). Genomes of the four strains were sequenced by Illumina. Sequence analyses revealed the presence of a high abundance of heavy metal resistance (HMR) determinants. One of the strain, M. rubeus P2, carried genes encoding 6 putative P1B-1-ATPase, 5 putative P1B-3-ATPase and 4 putative Zn(II)/Cd(II) P1B-4 type ATPase, and 16 putative RND-type metal transporter systems. Moreover, the four genomes carry a high abundance of genes coding for putative metal binding chaperones. Analysis of the close vicinity of these HMR determinants uncovered the presence of clusters of genes potentially associated with mobile genetic elements. These loci include genes coding for tyrosine recombinases (integrases) and subunits of mating pore (type 4 secretion system) respectively allowing integration/excision and conjugative transfer of numerous genomic islands. Further in silico analyses revealed that their genetic organization and gene products resemble the Bacteroides integrative and conjugative element CTnDOT. These results highlight the pivotal role of genomic islands in the acquisition and dissemination of adaptive traits, allowing for rapid adaption of bacteria and colonization of hostile environments.

Globally, the gastroenteritis or diarrhoea has become a more significant problem today due to infection caused by foodborne/ waterborne pathogen Vibrio cholera. In this concern, an investigation was carried out to evaluate the vibriocidal potential of the different solvent extracts of leaf and rhizome of Maranta arundinacea under in vitro condition. For this, aqueous, methanolic, ethanolic and hexane extracts of both leaf and rhizome of M. arundinacea were tested against the pre-isolated strains of Vibrio cholerae such as SPAB1, SPAB4 and SPAB5  by agar well diffusion and minimum inhibitory concentration (MIC) method. All the solvent extracts of both leaf and rhizome were found to be active against the tried strains of V. cholera however, ethanolic extract showed maximum inhibitory effect against SPAB1 strain with an inhibition zone of 26.23 ± 0.53 mm (MIC of 80.00 ± 10.06 µg/ ml) and 24.27 ± 0.12 mm (MIC of 100.00 ± 12.82 µg/ ml) in rhizome and leaf samples, respectively. Then, the effectiveness was followed in SPAB4 and SPAB5 however, it was not much more significant to that of SPAB1. Therefore, it was suggested that the rhizome and leaf extracts which proved to be potentially effective can be used as the natural alternative for the treatment of diarrhoea caused by Vibrio infection.

The human intestinal microbiota, establishing a symbiotic relationship with the host, plays a significant role for the human health. It is also well known that a disease status is frequently characterized by a dysbiotic condition of the gut. A probiotic treatment can represent an alternative therapy for enteric disorders and human pathologies not apparently linked to the gut environment. Among bifidobacteria, strains of the species Bifidobacterium breve are widely used in pediatrics. B. breve is the dominant species in the gut of breast-fed infants and it has also been isolated from human milk. It has antimicrobial activity against human pathogens, it does not possess transmissible antibiotic resistance traits, it is not cytotoxic and it has immuno-stimulating abilities. This review describes the applications of B. breve strains mainly for the prevention/treatment of pediatric pathologies. The target pathologies range from widespread gut diseases, including diarrhea and infant colics, to celiac disease, obesity, allergic and neurological disorders. Moreover, B. breve strains are used for the prevention of side infections in pre-term newborns and during antibiotic treatments or chemotherapy. With this documentation, we hope to increase knowledge on this species to boost the interest in the emerging discipline known as “therapeutic microbiology”.

Catechol dioxygenases in microorganisms cleave catechol into cis-cis-muconic acid or 2-hydroxymuconic semialdehyde via the ortho- or meta-pathway, respectively. The aim of the study was to purify, characterize and predict template-based three-dimensional structure of catechol 1,2-dioxygenase (C12O) from indigenous Pseudomonas chlororaphis strain UFB2 (PcUFB2). Preliminary studies showed that PcUFB2 could degrade 40 ppm of 2,4-dichlorophenol (2,4-DCP). The crude cell extract showed 10.34 U/mL of C12O activity with a specific activity of 2.23 U/mg of protein. A 35 kDa protein was purified to 1.5-fold with total yield of 13.02 % by applying anion exchange and gel filtration chromatography. The enzyme was optimally active at pH 7.5 and temperature 30 °C. The Lineweaver-Burk plot showed the v_max and K_m values of 16.67 µM/min and 35.76 µM, respectively. ES-MS spectra of tryptic digested SDS-PAGE band and bioinformatics studies revealed that C12O share 81% homology to homogentisate 1,2-dioxygenase reported in other Pseudomonas chlororaphis strains. Characterization and optimization of C12O activity can assist in understanding the 2,4-DCP metabolic pathway in PcUFB2 and its possible application in bioremediation strategies.

The application of 16S rRNA gene pyrosequencing has expanded our knowledge about the respiratory tract microbiome originally obtained using conventional, culture-based methods. In this study, we employed DNA-based molecular techniques for examining the sputum microbiome in bronchiectasis patients in relation to disease severity. Of 63 study subjects, 42 had mild and 21 had moderate or severe bronchiectasis, which was classified by calculating the FACED score based on FEV₁ (forced expiratory volume in 1 s, %) (F, 0–2 points), age (A, 0–2 points), chronic colonization by Pseudomonas aeruginosa (C, 0-1 point), radiographic extension (E, 0–1 point), and dyspnoea (D, 0–1 point). Bronchiectasis was defined as mild at 0–2 points, moderate at 3–4 points, and severe at 5–7 points. The mean age was 68.0 ± 9.3 years; 33 patients were women. Haemophilus (p=0.005) and Rothia (p=0.043) were significantly more abundant in the mild bronchiectasis group, whereas Pseudomonas (p=0.031) was significantly more abundant in the moderate or severe group. However, the alpha or beta diversity did not significantly differ among sputum microbiota, i.e. the same dominating genera were found in all samples. Further large-scale studies are needed to investigate the sputum microbiome in bronchiectasis.

Malassezia species are part of human commensal microbiota and is also related to diseases. Little is known about the interaction of these microorganisms with their host. Here we established two standard culture conditions for Malassezia spp. to perform infection assays using C. elegans and T. molitor. Invertebrate hosts infected by Malassezia spp. cultured in M9M resulted in higher death rate on survival assays when compared to yeasts cultured in standard Dixon medium indicating that M9M cultured Malassezia species have increased virulence. The culture and infection conditions established in this work, using invertebrate alternative models, are valuable tools to understand Malassezia-host interaction.

Abstract: Supplementation of infant formula with ingredients potentially able to manage, in a way positive for the host, the gut microbiota of infants, is nowadays widely used by infant food producers. The impact of this class of compounds, named prebiotics, on gut microbiota composition, was initially measured on the enrichment of Bifidobacterium and Lactobacillus spp., but the use of culture-independent analytical techniques has allowed to establish that a larger number of intestinal microorganisms are affected by the ingestion of prebiotics. However, despite a relevant number of scientific publications, actually there is no consensus on the amount of prebiotics to be administered daily, the potentially different actions exerted by prebiotics which differ in biochemical structures, as well as the impact of different percentages of the same prebiotics when used in mixtures and/or combination. This paper is aimed to critically review the available data on the use of prebiotics in infant formula, with special attention paid to identify a link between, the dosage used, the mixture composition and the observed outcomes, keeping in mind the influences in nutrition quality and growth influence.

Actinobaculum massiliense, a Gram-positive anaerobic coccoid rod colonizing the human urinary tract, belongs to the taxonomic class of Actinobacteria. We identified A. massiliense as a cohabitant of urethral catheter biofilms (CB). The CBs also harbored common uropathogens such as Proteus mirabilis and Aerococcus urinae, supporting the notion that A. massiliense is adapted to a life style in polymicrobial biofilms. We isolated a strain from an agar colony derived from a clinical sample. Using 16S rRNA gene sequencing and shotgun proteomics, we identified and characterized A. massiliense, comparing the isolate grown in vitro and four clinical ‘in vivo’ samples. Based on abundances of proteins in the in vivo milieu, we assessed their functions related to nutrient import and responses to hostile host conditions characterized by neutrophil infiltration. Two putative subtilisin-like proteases and a heme/oligopeptide transporter were highly expressed in vivo and are perhaps important for survival in the host milieu. The uptake of xylose/glucuronate and oligopeptides apparently enables feeding metabolites into mixed acid fermentation and peptidolysis pathways, respectively, to generate energy. A putative polyketide synthase which may generate a secondary metabolite interacting with either the host or co-colonizing microbes was identified. The enzyme may contribute to A. massiliense persistence in CBs.   

The application of quantitative PCR (qPCR) as a routine method to detect and enumerate Paenibacillus larvae in honey and hive debris could greatly speed up the estimation of prevalence and outbreak risk of the American foulbrood (AFB) disease of Apis mellifera. However, none of the qPCR tests described so far has been officially proposed as a standard procedure for P. larvae detection and enumeration for surveillance purposes. Therefore, in this study inclusivity, exclusivity and sensitivity in detection of P. larvae spores directly in samples of honey and hive debris were re-evaluated for the previously published qPCR methods. To this aim recently acquired P. larvae sequence data were considered to assess inclusivity in silico and more appropriate non-target species were used to verify exclusivity experimentally. This led to the modification of one of the previously described methods resulting in a new test capable to allow the detection of P. larvae spores in honey and hive debris down to 1 CFU/g. The application of the qPCR test optimized in this study can allow to reliably detect and quantify P. larvae in honey and hive debris, thus circumventing the disadvantages of late AFB diagnosis based on clinical symptoms and possible underestimation of spore numbers that is the main drawback of culture-dependent procedures.

This mini-review provides an overview of traditional, emerging, and future applications of lactic acid bacteria (LAB) and discusses how genome editing tools can be used to overcome current challenges in all these applications. It also describes currently available tools and how these can be further developed, and takes current legislation into account. Genome editing tools are necessary for the construction of strains for new applications and products, but can also play a crucial role in traditional ones, such as food and probiotics, as a research tool for understanding mechanistic insights and discovering new properties. Traditionally, recombinant DNA techniques for LAB have strongly focused on being food-grade, but they lack throughput and the number of genetically tractable strains is still rather limited. Further tool development in this direction will enable rapid construction of multiple mutants or mutant libraries on a genomic level in a wide variety of LAB strains. We also propose an iterative Design-Build-Test-Learn workflow cycle for LAB cell factory development based on systems biology, with “cell factory” expanding beyond its traditional meaning of production strains and making use of high-throughput genome editing tools to advance LAB understanding, applications and strain development.

Background Legionella pneumophila is one of the causes of legionellosis. Water environments serve as the natural habitat and the main sources of Legionella pneumophila. Objectives The aims of this study was to understand the differences of Legionella pneumophila serogroups distribution in well water, tap water, ice cubes, hospital and hotel water in East Java-Indonesia. Methods a total of 60 water samples in east java-Indonesia; from well water (n=25), tap water (n=5), ice cubes (n=5), water from the hospital (n=16), and hotel water (n=9) were detected using polymerase chain reaction with mip gene spesific primers and then it was analyzed by phylogenetic tree. Results For the 60 water samples collected in East Java, 12% of the samples (7/60) were positively contaminated by L. pneumophila. In details, there was 8% of the well water samples (2/25), 2% of the tap water samples (1/5), 2% of the ice cubes samples (1/5), 0% of the hospital water samples (0/16) and 33.33% of the hotel water samples (3/9). The phylogenetic tree showed that Legionella pneumophila contaminating well water isolate 1 from Surabaya and tap water isolate from Sidoarjo was closer to L.pneumophila serogroup 2, 4, 6, 8, 10, 12, isolates from Brazil, China, Spain and Australia. L.pneumophila contaminating the ice cubes from Sidoarjo was closer to serogroup 1, 2, 4, 7, 8, 11, 13, 14, while the bacteria contaminating well water isolate 2 from Sidoarjo as well as water in hotel of Surabaya (hotel water isolate 1, 2 and 3) classified into their own group. Conclusion There is a difference in the distribution of L. pneumophila serogroups between well water, tap water, ice cubes, and hotels.

Despite a considerable body of published research on Hepatitis B in Bangladesh, researchers continue to lament the lack of reliable information about Hepatitis B epidemiology. The present review aims to provide a comprehensive survey of the literature with particular focus on a number of epidemiological questions, as well as a commentary on the trends of Hepatitis B research as it has taken place in Bangladesh. The key themes to emerge from this review are: first, beyond noting a declining trend, it is difficult to provide conclusive estimates about Hepatitis B prevalence in the general population of Bangladesh. The majority of the studies, even the ones conducted on apparently healthy populations, fail to be adequately representative for the reasons explored in the article. Secondly, Hepatitis B in Bangladesh is sharply stratified across sociodemographic lines, which speaks to the role of awareness and risk exposure in Hepatitis B prevalence. Third, more research on occult infection rates is required to estimate the extent of risk posed by the current blood donation screening program, which relies exclusively on Hepatitis B surface antigen as a biomarker. The same considerations apply for the comparative importance of vertical vs. horizontal transmission, and prevalence among particular risk groups like healthcare workers with high occupational exposure. Finally, while recent studies do allow us, albeit with some ambiguity, to draw conclusions about distribution of Hepatitis B genotypes in Bangladesh, there needs to be an added emphasis on molecular epidemiology. It is hoped that the present review, the first of its kind in Bangladesh, will serve as an up-to-date summary of the course Hepatitis B epidemiology research in Bangladesh has taken thus far, as well as crucial gaps to address going forward.

The urinary tract infection (UTI) is the most common bacterial infection, especially in women. The increased incidence of UTIs, at the last decades have paralleled with the growing emergence of antibiotic resistance. The aim is to evaluate aqueous garlic extract (AGE) susceptibility against multidrug-resistant (MDR) bacteria isolated in urine of women. The investigation of antibacterial propriety and time kill effect of AGE was performed by the well method, microdilution method and spectrophotometer assay. Antibiotics susceptibility assay revealed that the nine MDR bacteria had high resistance against Amoxicillin/ clavulanic acid (100%) and Erythromycin (100%), Cefotaxime (83.33%) and Ceftazidime (83.33%). AGE exhibited potent antibacterial activity against the nine MDR bacteria tested. In Gram-negative bacteria, the inhibition diameters ranged from 20 ± 3 to 32 ± 4 mm, with Minimum Inhibitory Concentrations (MICs) ranging from 10% to 12.5% (w/v) and Minimum Bactericidal Concentration (MBCs) was 12.5 % (w/v). Gram-positive bacteria exhibited diameters ranging from 38 ± 2 to 45 ± 1 mm; MIC and MBC values ranged from 05 to 10 % (w/v) and were found more susceptible than Gram-negative bacteria. To conclude, this investigation shown that AGE have high potential antibacterial to use as an alternative to treat women UTIs.

Infectious diarrhoea such as shigellosis causes considerable morbidity and mortality, especially in infants, immune-compromised individuals and those living with HIV/AIDS. It is endemic in developing countries and in Sub-Saharan Africa, including South Africa, where diarrhoeal disease remains a leading cause of morbidity and mortality. This study was undertaken to establish incidences of Shigella, its serotype and resistant pattern of isolates from human faeces from residence of Johannesburg, South Africa. All stools received between January to April from the private healthcare system were cultured on Xylose Lysine Deoxycholate and MacConkey Agar and Shigella was confirmed by standard biochemical reactions and a serological method. An antimicrobial sensitivity test was used. A total of 11 009 samples from patients between 22 days to 94 years old yielded 110 Shigella isolates, of which 47 (43%) were S. flexneri, 61 (55%) were S. sonnei, 1 (1%) was S. dysenteriae and 1 (1 %) was S. boydii. The majority of patients were children between < 1 to 5 years, 76 (69%), followed by those between 6 to 10 years 13 (12%). In children up to 10 years, S. sonnei was confirmed in 52 cases (59%) and S. flexneri in 36 cases (41%). Overall, 53 (48%) males and 57 (52%) females were infected.

Prasinoviruses are large dsDNA viruses commonly found in aquatic systems worldwide, where they can infect and lyse unicellular prasinophyte algae such as Ostreococcus. Host susceptibility is virus strain-specific, but resistance of susceptible Ostreococcus tauri strains to a virulent virus arises frequently. In clonal resistant lines that re-grow, viruses are usually present for many generations, and genes clustered on chromosome 19 show physical rearrangements and differential expression. Here, we investigated changes occurring during the first two weeks after inoculation of viruses. By serial dilutions of cultures at the time of inoculation, we estimated the frequency of resistant cells arising in virus-challenged O. tauri cultures to be 10⁻³–10⁻⁴ of the inoculated population. Re-growing resistant cells were detectable by flow cytometry 3 days post-inoculation (dpi), visible re-greening of cultures occurred by 6 dpi, and karyotypic changes were visually detectable at 8 dpi. Resistant cell lines showed a modified spectrum of host-virus specificities and much lower levels of OtV5 adsorption.

Background: Staphylococcus aureus (S. aureus) occasionally threatens the life of the host as a persistent pathogen even though it is normal flora of humans and animals. We characterized drug resistance in S. aureus isolated from animal carcasses and milk samples from the abattoirs and dairy farms in the Eastern Cape Province. Methods: A 1000 meat swab samples and 200 raw milk samples were collected from selected abattoirs and dairy farms in the Eastern Cape Province, South Africa. S. aureus was isolated and positively identified using biochemical tests and confirmed by molecular methods. Antibiotic susceptibility test against 14 different antibiotics was performed against all isolates. Antibiotic resistance genes were also detected. Results: Of the 1200 samples collected, 134 (11.2%) samples were positive for S. aureus. Resistance ranged from 71.6% for penicillin G to 39.2% for tetracycline. Resistance gene (blaZ) was detected in 13 (14.9%), while msrA was found in 31 (52.5%) of S. aureus isolates. Conclusions: The present result shows the potential dissemination of multidrug-resistant S. aureus strains in the dairy farms and abattoirs in the Eastern Cape. Therefore, this implies that the organism may rapidly spread through food and pose serious public health risk

Fungal endophytes are the most ubiquitous plant symbionts on earth and are phylogenetically diverse. Studies on the fungal endophytes in tobacco have shown that they are widely distributed in the leaves, stems, and roots, and play important roles in the composition of the microbial ecosystem of tobacco. Herein, we analyzed and quantified the endophytic fungi of healthy tobacco leaves at the seedling (SS), resettling growth (RGS), fast-growing (FGS), and maturing (MS) stages at three altitudes [600 (L), 1000 (M), and 1300 m (H)]. We sequenced the ITS region of fungal samples to delimit operational taxonomic units (OTUs) and phylogenetically characterize the communities. The result showed the number of clustering OTUs at SS, RGS, FGS and MS were greater than 170, 245, 140, and 164, respectively. At the phylum level, species in Ascomycota and Basidiomycota had absolute predominance, representing 97.8% and 2.0 % of the total number of species. We also found the number of unique fungi at the RGS and FGS stages were higher than those in the other two stages. Additionally, OTU richness was determined by calculating the Observed Species, Shannon, Simpson, Chao1, ACE, Goods coverage and PD_whole_tree indices based on the total number of species. Our results showed RGS samples had the highest diversity indices. Furthermore, we found that the diversity of fungal communities tended to decrease with increasing tobacco growth altitude. The results from this study indicated that the tobacco harbors an abundant and diverse endophytic fungal population, which provides new opportunities for exploration their potential utilization.

The oral cavity harbors hundreds of microbial species that are present either as planktonic cells, or incorporated into biofilms. The majority of the oral microbes are commensal organisms. Those that are pathogenic microbes can result in oral infections, and at times initiate systemic diseases. Biofilms that contain pathogens have been challenging to control. Many conventional antimicrobials have proven to be ineffective. Recent advances in science and technology are providing new approaches for pathogen control and containment and methods to characterize biofilms. This perspective provides: 1) A general understanding of biofilm development; 2) A description of emerging chemical and biological methods to control oral biofilms; 3) An overview of high-throughput analytical approaches to analyze biofilms.

The antibiotic or antimicrobial resistance is rapidly spreading in microbes relevant to human health. Two visible major contributory factors have been the indiscriminate overuse of antimicrobials for preventing diseases in human and to enhance the productivity in agriculture sector. To mitigate the potential threat posed by post-antibiotic era, the global health stakeholders have been making extra efforts at a war footing to formulate and implement global and national plans of action. In the current article, an endeavour is made to provide a perspective to look beyond the current focus on just use of the antimicrobials. Attention has been drawn towards various obvious and not-so-obvious self-preservation infection-prevention practices in vogue from the pre-antibiotic era whose usage has been on decline in the antibiotic era for various reasons. Particularly, the practices with a clear potential to effectively decrease the spread of pathogens through contact, curtail the evolution and dissemination of the antimicrobial resistance in local environment and its introduction into the global community, should be Identified and strengthened to make them part of comprehensive hygiene and quarantine practices. Broadly, the suggestions pertaining to the personal and community hygiene including bereavement practices, isolation and quarantine of suspected pathogen carriers, and water and environment security have been made to invoke a constructive debate and discussion among various stakeholders for their evaluation and implementation to effectively delay the development of antimicrobial resistance wherever possible and disrupt its spread to pathogens.

The challenge posed by multi-drug resistance (MDR) of pathogenic organisms, spectacularly manifested in the 6 “ESKAPE” bacterium (two Gram-positive, four Gram-negative) species, should invoke new comprehensive strategies, and needs cooperation of scientists with medical, veterinary and natural science background. This review is aimed at informing newcomers, coming from the field of biology and genetics, about problems related to rapidly emerging, new multi-drug resistant, pathogenic, bacteria. Unlike persistence, the antibiotic resistance is inherited. A functioning “resistance gene” makes a susceptible organism resistant to a given antibiotic, encoding for polypeptides capable of acting either as decomposing enzymes, or acting as trans-membrane pumps, or membrane structure components capable of modifying the permeability implementing a «by pass» mechanism enabling the antibiotic molecule to reach its cellular target(s). A functioning “sensitivity gene” encode for a polypeptide, capable (directly or indirectly) of transferring toxic molecules into target cells, or of metabolizing non-transferable to transferable, or non-toxic molecules to toxic derivatives. A gene of a normal function could act as a “sensitivity” gene in the presence of antibiotics of chemical structures similar to the natural substrate of the gene product, (enzyme or binding/
trans-membrane protein). The Agrocin 84 story is a good example. Multi-drug resistance is a phenotypic consequence of the sequential accumulation of mutations, and/or up-take of plasmids or genomic islands carrying resistance genes from the environment via horizontal gene transfer, mediated by conjugative plasmid or bacteriophage carrying mobile genetic elements. Both multi-drug resistance and collateral sensitivity are evolutionary products. Some revealed evolutionary process and their Lamarckian and Darwinian interpretations are discussed. Toolkits of comparative full-genome sequencing, genomics, experimental evolution and population genetics may provide perspectives for overcoming the invincibility of multi-drug panresistance. The status of some recently emerging pathogenic bacterium species with zoonic features and of veterinary background is also discussed.

Cadmium is a highly toxic heavy metal for biological systems. Cupriavidus metallidurans CH34 is a model strain for heavy metal resistance and bioremediation. The aim of this study was to determine the role of the c-di-GMP pathway in the C. metallidurans CH34 response to cadmium in both planktonic and biofilm cells. Increasing cadmium concentrations correlates with an inhibition of biofilm formation and EPS production in C. metallidurans cells. Planktonic and biofilm cells showed similar tolerance to cadmium. During exposure to cadmium an acute decrease of c-di-GMP levels in planktonic and biofilm cells was observed. Transcription analysis by RT-qPCR showed that cadmium induced in planktonic cells and strongly induced in biofilm cells the expression of the urf2 gene and the mercuric reductase encoding merA gene, which belong to the Tn501/Tn21 mer operon. After exposure to cadmium the cadA gene involved in cadmium resistance was equally upregulated in both lifestyles. Bioinformatic analysis and null mutant complementation assays indicated that the protein encoded by the urf2 gene is a functional phosphodiesterase involved in the c-di-GMP metabolism. We propose to rename the urf2 gene as mrp gene for metal regulated phosphodiesterase. An increase of the second messenger c-di-GMP content by the heterologous expression of the constitutively active diguanylate cyclase PleD* correlated with an increase in biofilm formation and cadmium susceptibility. These results indicate that the response to cadmium in C. metallidurans CH34 involves a decrease in c-di-GMP content that inhibits the biofilm lifestyle.

New approaches to deal with drug-resistant pathogenic bacteria are urgent. We studied the antibacterial effect of chitosans against an E. coli quorum sensing biosensor reporter strain, and selected a non-toxic chitosan to evaluate its QS inhibition activity and its effect on bacterial aggregation. To this end, chitosans of varying DA (12 to 69%) and M_w (29 to 288 KDa) were studied. Only chitosans of low DA (~12%) inhibited the bacterial growth, regardless of the M_w. Chitosan MDP DA30 (DA 42% and M_w 115 kDa) was selected for further QS inhibition and SEM imaging studies. MDP DA30 chitosan exhibited QS inhibition activity in an inverse dose-dependent manner (≤12.5 µg/mL). SEM images revealed that this chitosan, when added at low concentration (≤30.6 µg/mL), induced substantial bacterial aggregation, whereas at high concentration (234.3 µg/mL), it did not. Aggregation explains the QS inhibition activity as the consequence of retardation of the diffusion of AHL.

Neisseria gonorrhoeae is a strict human pathogen responsible for more than 100 million new sexually transmitted infections worldwide each year. Due to the global emergence of antibiotic resistance, the CDC recently listed N. gonorrhoeae as an urgent threat to public health. No vaccine is available in spite of the huge disease burden and the possibility of untreatable gonorrhea. The aim of this study is to investigate the immunogenicity of a novel whole-cell based inactivated gonococcal microparticle vaccine formulation loaded in dissolvable microneedles for transdermal administration. The nanotechnology-based vaccine formulation consists of inactivated whole-cell gonococci strain CDC-F62, spray dried and encapsulated into biodegradable cross-linked albumin matrix with sustained slow antigen release. The dry vaccine nanoparticles were then loaded in a dissolvable microneedle skin patch for transdermal delivery. The efficacy of the whole-cell microparticles vaccine formulation loaded in microneedles was assessed in vitro using dendritic ,cells and macrophages as well as in vivo mouse model. Antibody titers were measured using an ELISA and antigen-specific T lymphocytes were assessed in spleens and lymph nodes. Here we report that whole-cell based gonococcal microparticle vaccine loaded in dissolvable microneedles for transdermal administration induced significant increase in antigen-specific IgG antibody titers and antigen-specific CD4 and CD8 T lymphocytes in mice compared to gonococcal antigens in solution or empty microneedles. Significant increase in antigen-specific IgG antibody levels was observed at end of week 2 in groups that received the vaccine compared to the group receiving empty nanoparticles. The advantages of using formalin-fixed whole-cell gonococci that all immunogenic epitopes are covered and preserved from degradation. The spherical shaped micro and nanoparticles are biological mimics of gonococci, therefore present to the immune system as invaders but without the ability to suppress adaptive immunity. In conclusion, the transdermal delivery of microparticles vaccine via a microneedle patch was shown to be an effective system for vaccine delivery. The novel gonorrhea nanovaccine is cheap to produce in a stable dry powder and can be delivered in microneedle skin patch obviating the need for needle use or the cold chain.

Escherichia coli phage Eco_BIFF was isolated from several laboratory stocks of E. coli K-12 MG1655 derivatives. The source of the contamination is unknown. Eco_BIFF is a lytic phage that shows effective growth inhibition of E. coli K-12. Here, we announce the complete genome sequence of Eco_BIFF, and major findings from its genome annotation.

Streptococcus mutans (S. mutans) is the main etiological bacteria present in the oral cavity that leads to dental caries. All of the S. mutans in the oral cavity form biofilms that adheres to the surfaces of teeth. Dental caries are infections facilitated by the development of biofilm. An esterified derivative of epigallocatechin-3-gallate (EGCG), epigallocatechin-3-gallate-sterate (EGCG-S) was used in this study to assess its ability to inhibit the growth and biofilm formation of S. mutans. The effect of EGCG-S on bacterial growth was evaluated with colony forming units (CFU) and log reduction; biofilm formation was qualitatively determined by Congo red assay, and quantitatively determined by crystal violet assay, fluorescence-based LIVE/DEAD assays to study the cell viability, and scanning electron microscopy (SEM) was used to evaluate the morphological changes. The results indicated that EGCG-S was able to completely inhibit growth and biofilm formation at concentrations of 250 µg/ml. Its effectiveness was also compared with a commonly prescribed mouthwash in the United States, chlorhexidine gluconate. EGCG-S was shown to be equally effective in reducing S. mutans growth as chlorhexidine gluconate. In conclusion, EGCG-S is potentially a natural anticariogenic agent by reducing bacterial presence in the oral cavity.

The human virome is an area of increasing interest with relation to human health and disease. It has been demonstrated to alter in concert with the bacterial microbiome in early life and was also found to be different in patients with certain diseases such as inflammatory bowel disease. However, all virome analyses are hampered by a lack of annotated representative database sequences, often referred to as the ‘viral dark matter’. Here we provide the first description of the gut DNA virome in elderly individuals (>65 years old) as well as the description of novel bacteriophages not present in current reference databases. Diversity analysis comparing elderly persons from different residence locations (community living vs long term care facilities) did not reveal any difference in their virome diversity profiles despite the reported differences at the bacteriome level. An abundance of Microviridae of the subfamily Gokushovirinae were present in the faeces of elderly individuals. Several novel members of the order Caudovirales were also characterized and annotated. Assignment of host bacteria to detected viral genomes was attempted using a combination of CRISPR spacers, tRNA genes and a probabilistic approach. Further characterization of the viral dark matter is necessary for developing tools and expanding databases to study the human virome. This study focused on the virome of an aging human cohort with the goal of illuminating part of the viral dark matter.

Proper species identification from ancient DNA samples is a difficult task that sheds light on the evolutionary history of pathogenic microorganisms. The field of palaeomicrobiology has undoubtedly benefited from the advent of untargeted metagenomic approaches that use next-generation sequencing methodologies. Nevertheless, assigning ancient DNA at the species level is a challenging process. Recently, the gut microbiome analysis of three pre-Columbian Andean mummies [1](Santiago-Rodriguez et al. 2016) has called into question the identification of Leishmania in South America. Here, the metagenomic data filed in MG-RAST (Metagenomics RAST server) were used for a further attempt to identify members of the Trypanosomatidae family infecting these ancient remains. For this purpose, we used two metagenomic analysis tools. In the first step, data were analysed using the ultrafast metagenomic sequence classifier, based on exact alignment of k-mers (Kraken). In the second step, we used Bowtie2, an ultrafast and memory-efficient tool for aligning sequencing reads to long reference sequences. We then compared the output results. These approaches highlight some interesting findings on potential infections by human pathogenic trypanosomatids in these three pre-Columbian mummies.

The human virome is an area of increasing interest with relation to human health and disease. It has been demonstrated to alter in concert with the bacterial microbiome in early life and was also found to be different in patients with certain diseases such as inflammatory bowel disease. However, all virome analyses are hampered by a lack of annotated representative database sequences, often referred to as the ‘viral dark matter’. Here we provide the first description of the gut DNA virome in elderly individuals (>65 years old) as well as the description of novel bacteriophages not present in current reference databases. Diversity analysis comparing elderly persons from different residence locations (community living vs long term care facilities) did not reveal any difference in their virome diversity profiles despite the reported differences at the bacteriome level. An abundance of Microviridae of the subfamily Gokushovirinae were present in the faeces of elderly individuals. Several novel members of the order Caudovirales were also characterized and annotated. Assignment of host bacteria to detected viral genomes was attempted using a combination of CRISPR spacers, tRNA genes and a probabilistic approach. Further characterization of the viral dark matter is necessary for developing tools and expanding databases to study the human virome. This study focused on the virome of an aging human cohort with the goal of illuminating part of the viral dark matter.

Whey proteins have excellent nutritional characteristics due to their levels of essential amino acids with high bioavailability. However, it has a high biochemical oxygen demand (BOD) and a considerable polluting potential, thus food manufacturers have opted to add whey to food formulations. The demand for beverages containing vitamins, probiotics, prebiotics, minerals, and bioactive compounds (antioxidants) with health benefits has increased and driven market growth. Therefore, this study aimed to develop a probiotic functional carbonated beverage from cheese whey and evaluate its microbiological, and physicochemical characteristics soon after the production and during storage. The viability and stability of probiotic, the microbiological characteristics, titratable acidity and sedimentation of the beverage were monitored during refrigerated storage for a month. The probiotic to be added to the formulation was established in a preliminary step. The production of this beverage proved to be a simple technology and the product was suitable for incorporation of the probiotic Bifidobacterium animalis subsp. lactis. The probiotic showed good viability and stability during storage. The microbiological quality of the beverage met the Brazilian legal standards. The pH and titratable acidity of the probiotic carbonated beverage remained stable during storage, and slight sedimentation was observed after one week of refrigerated storage.

Markets from Maputo City have been reported to have poorly hygienic premises and sellers unaware of the safest food handling practices. This study aimed to analyze associations between some lettuce (Lactuca sativa) handling features and the levels of total, fecal coliforms and Escherichia coli. Cabbage was purchased in the 3 markets. Just before the acquisition, the researchers recorded the source of water used to wash it, if they were sold in booths or on the floor, if they were sold mixed with other products, and if the sellers were wearing gloves. The cabbage heads (42) were analyzed through the technique of multiple tubes to estimate the most probable number. Average levels were 14.43 MPN/g for total coliforms, 13.9 MPN/g for fecal coliforms and 12.02 MPN/g for E. coli. The highest levels of contamination seemed associated with the use of well water, the food sold on the floor, the lettuce sold together with other products and the lack of gloves. The mixture of products only showed a major impact on the level of E. coli (p < 0.001). Yet, all levels were within the satisfactory range, according to the guidelines from The International Commission on Microbiological Specifications for Foods (ICMSF).

African trypanosomes are an early branch in eukaryotic evolution and developed a unique endomembrane system as an adaptation to their parasitic life style. The key virulence mechanism of many pathogens is successful immune evasion to enable survival within the host, which is a feature requiring both genetic events and membrane transport in African trypanosomes. Intracellular trafficking not only plays a role in immune evasion, but also in homeostasis of intracellular and extracellular compartments and interactions with the environment. Significantly, historical and recent work has unravelled some of the connections between these processes and highlighted how immune evasion mechanisms associated with adaptations in membrane trafficking may have, paradoxically, provided specific sensitivity to drugs. Here we explore these advances in understanding the membrane composition of the trypanosome plasma membrane and organelles and provide a perspective for how transport could be exploited for therapeutic purpose.

Urinary tract infections are a major cause of morbidity and mortality in Mozambique. They are sometimes treated empirically with nitrofurantoin. However, little is known about this antibiotic’s performance and bacterial resistance in the country. This study analyzed the results of nitrofurantoin sensitivity tests requested in the Central Hospital of Maputo during 2012 and 2013. As result, 181 samples were tested and most cases (66.9%) showed absolute sensitivity but there were considerable cases of resistance (29.8%). Morganella morganii was the only bacteria presenting no absolute or intermediate resistance. The sensitivity was also high in the case of Escherichia coli (90%) and Gram-negative bacteria (66.7%). Serratia marcescens was mostly resistant (64.3%). The remaining bacteria showed inconclusive results. Thus they shall be subjected to a sensitivity test before prescription. Factors such as seasonality, patients’ sex and urine transparency did not seem to be reliable indicators of microbial resistance in the urine. Yet, a longer time span (over 5 years) might be sufficient for the sensitivity profile to change.

Improperly prepared fresh fruit and vegetable juices are recognized as an emerging cause of food borne illnesses. Therefore, this study was aimed at evaluating the microbiological safety of fresh fruit juices marketed in Debre-Markos town and their hygienic conditions of preparations. Thirty six fruit juices samples were collected from 6 cafés and restaurants of Debre-Markos town and analyzed for total aerobic viable bacterial count (TAVBC), total staphylococcal count (TSC), aerobic spore forming bacterial count (ASFBC), total coliform count (TCC), fecal coliform count (FCC), yeast and mould count (YMC). The spread plate method was used for the isolation of microorganisms on appropriate selective media. All isolates were characterized following standard methods. Bacterial and fungal species were isolated following standard methods. Questionnaires were distributed for 30 juice makers to obtain preliminary information on hygienic and safety practices of fruit juice makers. Results show that the mean TAVBC, ASFBC, TSC, yeast and mold, TCC and FCC of mango were 2.2±0.48x10⁶,0.13±0.04x10⁵ ,0.004x10⁵ ,1.1±0.2x10⁶ ,0.15±0.05x10⁵,5.7±3.73x10⁴ and 0.06±0.04x10⁴ cfu/ml respectively. The mean of TAVBC, ASFBC, TSC, YMC, TCC, and FCC of avocado juice were 3.6±0.6x10⁶, 0.08±0.02x10⁵, 0.27±0.07x10⁵, 1.2±0.4x10⁶, 0.02±0.01x10⁵, 6.46±3.7x10^4, and 0.2±0.1x10⁴ cfu/ml respectively. The bacterial isolates were identified as Staphylococcus aureus, Escherichia coli, Klebsiella spp. Bacillus cereus, Enterobacter spp., Enterococcous spp., Streptococcus spp., and Serratia spp. while the identities of the fungal isolates were Fusarium spp., Mucor spp. and Saccharomyces cerevisiae. The results also showed that the microbial loads of most of the fruit juices were higher than the specifications set for fruit juices sold in the Gulf region and other parts of the world. Most venders obtained fruit from the open market and all juice makers lacked special training in food hygiene and safety. Therefore, regular training and health education on food hygiene and safety is recommended for juice handlers to improve the quality of fresh fruit juices in the study area.

This study investigated whether biogas reactor performance, including microbial community development, in response to a change in substrate composition is influenced by initial inoculum source. Test reactors were first started with two different inocula and operated with the same grass-manure mixture for more than 120 days. These reactors initially showed great differences depending on inoculum source, but eventually showed similar performance and overall microbial community structure. At the start of the present experiment, the substrate was complemented with milled feed wheat, added all at once or divided into two portions. The starting hypothesis was that process performance depends on initial inoculum source and microbial diversity, and thus that reactor performance is influenced by the feeding regime. In response to the substrate change, all reactors showed increases and decreases in volumetric and specific methane production, respectively. However, specific methane yield and development of the microbial community showed differences related to initial inoculum source, confirming the hypothesis. The different feeding strategies had however only minor effects on process performance and overall community structure, but still induced differences in the cellulose-degrading community and in cellulose degradation.

Current research in the field of antimicrobials is focused on the development of novel antibiotics and antimicrobial agents to counteract the huge dilemma that the human population is mainly facing in regards to the rise of bacterial resistance and biofilm infections. Host Defense peptides (HDPs) are a promising group of molecules for antimicrobial development as they share unique characteristics suitable for antimicrobial activity including their broad spectrum of activity and potency against bacteria. AamAP1 is a novel HDP that was identified through molecular cloning from the venom of the North African scorpion Androctonus amoeruxi. In vitro antimicrobial assays revealed that the peptide displays moderate activity against different strains of Gram-positive and Gram-negative bacteria. Additionally, the peptide proved to be highly hemolytic and displaying significantly high toxicity against mammalian cells. In our study, a novel synthetic peptide analogue named A3 was designed from the naturally occurring scorpion venom host defense peptide. The design strategy depended on modifying the amino acid sequence of the parent peptide in order to increase its net positive charge, percentage helicity and optimize other physico-chemical parameters involved theoretically in HDPs activity. Accordingly, A3 was evaluated for its in vitro antimicrobial and anti-biofilm activity individually and in combination with four different types of conventional antibiotics against clinical isolates of multi-drug resistant (MDR) Gram-positive bacteria. A3 was also evaluated for its cytotoxicity against mammalian cells. A3 displayed potent and selective in vitro antimicrobial activities against a wide range of MDR Gram-positive bacteria. Our results also showed that combining A3 with conventional antibiotics displayed a synergistic mode of action which resulted in decreasing the MIC value for A3 peptide as low as 0.125 µM. These effective concentrations were associated with negligible toxicities on mammalian cells. In conclusion, A3 exhibits enhanced activity and selectivity when compared with the parent natural scorpion venom peptide. The combination of A3 with conventional antibiotics may be pursued as a potential novel treatment strategy against MDR and biofilm forming bacteria.

Butanol extracts from leaves, twigs and fruits of Elaeagnus rhamnoides (L.) A. Nelson (sea buckthorn, SBT) were fractionated into phenolic and non-polar lipid components. Their chemical composition was analyzed using the Thermo Ultimate 3000RS chromatographic system, equipped with a diode array detector, a corona-charged aerosol detector, and coupled with a (Q-TOF) mass spectrometer. Assuming that an effect on natural microbiotaand host epithelial cells needs to be assessed, regardless of the purpose of using SBT formulations in vivo, the MIC/MBC/MFC of the fractions and reference phytocompounds were screened involving 17 species of Gram-positive and Gram-negative bacteria and Candida species. The impact of the fractions (subMIC) on the important in vivo persistence properties of S. aureus and C. albicans strains was evaluated. Tests for adhesion and biofilm formation on an abiotic surface and the surfaces conditioned with fibrinogen, collagen, plasma or artificial saliva showed the inhibitory activity of the fractions. The effects on FITC-labeled staphylococci adhesion to fibroblasts (HFF-1) and epithelial cells (Caco-2), and on fungal morphogenesis, indicated that SBT extracts have high anti-virulence potential. Cytotoxicity tests (MTT-reduction) on the standard fibroblast cell line showed variable biological safety of the fractions depending on their composition and concentration.

Polyketides are large group of secondary metabolites that have notable variety in their structure and function. Polyketides exhibit a wide range of bioactivities such as antibacterial, antifungal, anticancer, antiviral, immune-suppressing, anti-cholesterol and anti-inflammatory activity. Naturally, they are found in bacteria, fungi, plants, protists, insects, mollusks and sponges. Streptomyces is a genus of Gram-positive bacteria that has a filamentous form like fungi. This genus is best known as one of polyketides producers. Some examples of polyketides produced by Streptomyces are rapamycin, oleandomycin, actinorhodin, daunorubicin and caprazamycin. Biosynthesis of polyketides involves a group of enzyme activities called polyketide synthases (PKSs). There are three types of PKSs (type I, type II, and type III) in Streptomyces that responsible for producing polyketides. This paper focuses on biosynthesis of polyketides in Streptomyces with three structurally different types of PKSs.

The aim of our work was to check if one of the products of natural origin, namely honey bee propolis may be an alternative or supplement to currently used antifungal agents. The activity of 50 ethanolic extracts of propolis (EEPs), harvested in Polish apiaries, was tested on a group of 69 clinical isolates of C. albicans. Most of the EEPs showed satisfactory activity, with minimum fungicidal concentrations (MFC) mainly in the range of 0.08-1.25% (v/v). Eradication of biofilm from polystyrene microtitration plates in 50% (MBEC50) required concentrations in the range of 0.04% (v/v) to more than 1.25% (v/v). High activity was also observed in eradication of biofilm formed by C. glabrata and C. krusei on the surfaces of PVC and silicone catheters. EEPs at subinhibitory concentrations inhibited yeast-to-mycelia morphological transformation of C. albicans in liquid medium and mycelial growth on solid medium. A synergistic effect was observed for the action of EEPs in combination with fluconazole (FLU) and voriconazole (VOR) against C. albicans. In the presence of EEP at concentration as low as 0.02%, the MICs of FLU and VOR were 256 to 32 times lower in comparison to those of the drug alone. Evidences for the fungal cell membrane as the most probable target of EEPs are presented.

The order Mucorales, the largest in number of species within the Mucoromycotina, comprises typically fast-growing saprotrophic fungi. During a study of the fungal diversity of undiscovered taxa in Korea, two novel mucoralean strains, CNUFC-GWD3-9 and CNUFC-EGF1-4, were isolated from specific habitats including freshwater and fecal samples, respectively, in Korea. On the basis of their morphological characteristics and sequence analyses of internal transcribed spacer (ITS) and large subunit (LSU) of 28S ribosomal DNA regions, the CNUFC-GWD3-9 and CNUFC-EGF1-4 isolates were confirmed to be Gilbertella persicaria and Pilobolus crystallinus, respectively. It is ecologically, pathologically and mycologically significant to find such rare zygomycetous fungi in such specific habitats. 

Under abiotic stress conditions, arbuscular mycorrhizal (AM) fungi help plants by improving nutrient and water uptake. Finger millet is an arid crop having soils with poor water holding capacity. Therefore, it is difficult for the plants to obtain water and mineral nutrients from the soil to sustain life. To understand the role of mycorrhizal symbiosis in water and mineral up-take from the soil, we studied the role of Rhizophagus intraradices colonization and its beneficial role for drought stress tolerance in finger millet seedling. Under severe drought stress condition, AM inoculation led to the significant increase in plant growth (7%), phosphorus, and chlorophyll content (29%). Also, the level of osmolytes including proline and soluble sugars were found in higher quantities in AM inoculated seedlings under drought stress. Under water stress, the lipid peroxidation in leaves of mycorrhized seedlings was reduced by 29%. The flavonoid content of roots in AM colonized seedlings was found 16% higher compared to the control, whereas the leaves were accumulated more phenol. Compared to the control, ascorbate level was found to be 25% higher in leaf tissue of AM inoculated seedlings. Moreover, glutathione (GSH) level was increased in mycorrhiza inoculated seedlings with a maximum increment of 182% under severe stress. The results demonstrated that AM provided drought tolerance to the finger millet seedlings through a stronger root system, greater photosynthetic efficiency, a more efficient antioxidant system and improved osmoregulation.

Filamentous fungi such as Aspergillus spp. are opportunistic pathogens, which cause highly invasive infections, especially in immunocompromised individuals. Control of such fungal pathogens is increasingly problematic due to the small number of effective drugs available for treatment. Moreover, the increased incidence of fungal resistance to antifungal agents makes this problem a global human health issue. The cell wall integrity system of fungi is the target of antimycotic drugs echinocandins, such as caspofungin (CAS). However, echinocandins cannot completely inhibit the growth of filamentous fungal pathogens, which results in survival/escape of fungi during treatment. Chemosensitization was developed as an alternative intervention strategy, where co-application of CAS with the intervention catalyst octyl gallate (OG; chemosensitizer) greatly enhanced CAS efficacy, thus achieved ≥ 99.9% elimination of filamentous fungi in vitro. Based on hypersensitive responses of Aspergillus antioxidant mutants to OG, it is hypothesized that, besides destabilizing cell wall integrity, the redox-active characteristic of OG may further debilitate fungal antioxidant system.

In the last years evidence has emerged that neurodegenerative diseases (NDs) are strongly associated with the microbiome composition in the gut. Parkinson’s disease (PD) is the most intensively studied neurodegenerative disease in this context. In this review, we performed a systematic evaluation of published literature comparing changes in colonic microbiome in PD to the ones observed in other NDs including Alzheimer’s Disease (AD), Multiple system atrophy (MSA), Multiple sclerosis (MS), Neuromyelitis optica (NMO) and Amyotrophic lateral sclerosis (ALS). To warrant comparability of different studies, only human case-control studies were included. Several studies showed an increase of Lactobacillus, Bifidobacterium, Verrucomicrobiaceae and Akkermansia in PD. A decrease in PD was observed of Faecalibacterium spp., Coprococcus spp., Blautia spp., Prevotella spp. and Prevotellaceae. On low taxonomic resolution, like phylum level, the changes are not disease specific and inconsistent. However, on higher taxonomic resolution like genus or species level, a minor overlap was observed between PD and MSA, both alpha synucleinopathies. We show that a methodical standardization of sample collection and analysis is necessary for ensuring the reproducibility and comparability of data. We also provide the evidence, that assessing the microbiota composition at high taxonomic resolution, reveals changes in relative abundance, that may be specific or characteristic for one disease, or a disease-group and might evolve discriminative power. The interactions between bacterial species and strains and moreover the co-abundances must be more deeply investigated before assumptions of the effects of specific bacteria on the host can be made with certainty.

Scrub typhus threatens one billion people in the Asia-Pacific area and cases have emerged outside this region. It is caused by infection with any of the multitude of strains of the bacterium, Orientia tsutsugamushi. A vaccine that affords heterologous protection and a commercially available molecular diagnostic assay are lacking. Herein, we determined that the nucleotide and translated amino acid sequences of outer membrane protein A (OmpA) are highly conserved among 51 O. tsutsugamushi isolates. Molecular modeling revealed the predicted tertiary structure of O. tsutsugamushi OmpA to be very similar to that of the phylogenetically related pathogen, Anaplasma phagocytophilum, including the location of a helix that contains residues functionally essential for A. phagocytophilum infection. PCR primers were developed that amplified ompA DNA from all O. tsutsugamushi strains, but not from negative control bacteria. Using these primers in quantitative real-time PCR enabled sensitive detection and quantitation of O. tsutsugamushi ompA DNA from organs of mice that had been experimentally infected with the Karp or Gilliam strains. The high degree of OmpA conservation among O. tsutsugamushi strains evidences its potential to serve as a molecular diagnostic target and justifies its consideration as a candidate for developing a broadly protective scrub typhus vaccine.

Fatty acids play a major role in determining membrane biophysical properties. Staphylococcus aureus produces branched-chain fatty acids (BCFAs) and straight-chain fatty acids (SCFAs), and can incorporate exogenous SCFAs and straight-chain unsaturated fatty acids (SCUFAs). Many S. aureus strains produce the triterpenoid pigment staphyloxanthin, and the balance of BCFAs, SCFAs and staphyloxanthin determines membrane fluidity. Here, we investigated the relationship of fatty acid and carotenoid production in S. aureus using a pigmented strain (Pig1), its carotenoid-deficient mutant (Pig1ΔcrtM) and the naturally non-pigmented Staphylococcus argenteus that lacks carotenoid biosynthesis genes and is closely related to S. aureus. Fatty acid compositions in all strains were similar under a given condition indicating that staphyloxanthin does not influence fatty acid composition. Strain Pig1 had decreased membrane fluidity as measured by fluorescence anisotropy than the other strains under all conditions indicating that staphyloxanthin helps maintain membrane rigidity. We could find no evidence for correlation of expression of crtM and fatty acid biosynthesis genes. Supplementation of medium with glucose increased SCFA production and decreased BCFA and staphyloxanthin production, whereas acetate-supplementation also decreased BCFAs but increased staphyloxanthin production. We believe that staphyloxanthin levels are influenced more through metabolic regulation than responding to fatty acids incorporated into the membrane.

In Belgium, silages are often infected by Penicillium roqueforti sensu lato (s.l.). These toxigenic fungi are well adapted to silage conditions, and their prevention during feed-out is difficult. Bacillus velezensis strain NRRL B-23189 has been reported to inhibit P. roqueforti s.s. conidiospore germination in vitro by the production of lipopeptides. In the present study, the antagonistic effect of this B. velezensis strain towards P. roqueforti s.l. was evaluated in vitro and in vivo. In vitro, corn silage conditions were simulated, and the impact of B. velezensis culture supernatant or cell suspension on P. roqueforti s.l. growth, conidiospore germination and survival and roquefortine C production was evaluated. The antagonism was promising, but growth of B. velezensis in corn silage infusion was poor. An in vivo experiment with microsilos containing a mixture of perennial ryegrass and white clover artificially contaminated with P. roqueforti s.l. was carried out to determine if B. velezensis cell suspension could be used as an antagonistic silage inoculant. The B. velezensis cell suspension applied was unsuccessful in reducing P. roqueforti s.l. numbers at desiling after 56 days compared to no additive application. However, feed-out of the silage was not simulated, so it remains elusive whether or not B. velezensis exerts antagonistic activity during this phase.

Moonmilk are cave carbonate deposits that host a rich microbiome including antibiotic-producing Actinobacteria making these speleothems appealing for bioprospecting. Here we investigated the taxonomic profile of the actinobacterial community of three moonmilk deposits of the cave “Grotte des Collemboles” via high-throughput sequencing of 16S rRNA amplicons. Actinobacteria was the most common phylum after Proteobacteria, ranging from 9 to 23% of the total bacterial population. Next to actinobacterial OTUs attributed to uncultured organisms at the genus level (~44%), we identified 47 actinobacterial genera with Rhodoccocus (4 OTUs, 17%) and Pseudonocardia (9 OTUs, ~16%) as the most abundant in terms of absolute number of sequences. Streptomycetes presented the highest diversity (19 OTUs, 3%), with most of OTUs unlinked to the culturable Streptomyces strains previously isolated from the same deposits. 43% of OTUs were shared between the three studied collection points while 34% were exclusive to one deposit indicating that distinct speleothems host their own population despite their nearby localization. This important spatial diversity suggests that prospecting within different moonmilk deposits should result in the isolation of unique and novel Actinobacteria. These speleothems also host a wide range of non-streptomycetes antibiotic-producing genera, and should therefore be subjected to methodologies for isolating rare Actinobacteria.

Soil microbial fuel cells (sMFC) are a novel technique that use organic matters in soils as an alternative energy source. External resistance (ER) is a key factor influencing sMFC performance and, furthermore, alters the soil’s biological and chemical reactions. However, little information is available on how the microbial community and soil component changes in sMFC with different ER. Therefore, the effects of anodes of sMFC at different ER (2000 Ω, 1000 Ω, 200 Ω, 80 Ω and 50 Ω) were examined by measuring organic matter (OM) removal efficiency, trace elements in porewater and bacterial community structure in contaminated paddy soil. The results indicated that ER has significant effects on sMFC power production, OM removal efficiency and bacterial beta diversity. Moreover ER influences iron, arsenic and nickel concentration as well in soil porewater. In particular, greater current densities were observed at lower ER (2.4mA, 50Ω) compared to a higher ER (0.3mA, 2000Ω). The removal efficiency of OM increased with decreasing ER whereas it decreased with soil distance away from the anode. Furthermore, principal coordinate analysis (PCoA) revealed that ER may shape the bacterial communities that develop in the anode vicinity but have minimal effect on that of the bulk soil. The current study illustrates that lower ER can be used to selectively enhance the relative abundance of electrogenic bacteria and lead to high OM removal.

Neisseria meningitidis causes most cases of bacterial meningitis. Meningococcal meningitis is a public health burden to both developed and developing countries throughout the world. There are a number of vaccines (polysaccharide-based, glycoconjugate, protein-based and combined conjugate vaccines) that are approved to target five of the six disease-causing serogroups of the pathogen. Immunization strategies have been effective at helping to decrease the global incidence of meningococcal meningitis. Researchers continue to enhance these efforts through discovery of new antigen targets that may lead to a broadly protective vaccine and development of new methods of homogenous vaccine production. This review describes current meningococcal vaccines and discusses some recent research discoveries that may transform vaccine development against N. meningitidis in the future.

Native strains of Trichoderma, isolated from mangrove sediments of PE, Brazil were determining their morphological and molecular characterization, and were investigated to assess of their biocontrol potential over the phytopathogenic Fusarium strains isolated from Caatinga soil, PE, Brazil. The Trichoderma strains were characterized by polyphasic approach, which combined their morphological characteristics, macro- and microculture results, growth evaluation by Tukey test, with significance of 5%. The DNA was extracted and the product was amplified with primers ITS 1 and 2, and sequenced. Trichoderma strains were compatible morphologically with the description of the genus. The molecular identification of Trichoderma, sequences of 500 bp were amplified, deposited in GenBank and used for phylogenetic analysis. The growth rate analysis showed rate of 0.1207 cm^h-1 to Trichoderma strains and Fusarium spp. lower growth rate (0.031 cm^h-1) was observed. The antibiosis tests showed the best antagonistic level of effectiveness to T. asperellum UCP 0149 against F. solani UCP 1395(82.2%) and F. solani UCP 1075(70.0%), followed of T. asperellum UCP 0319 against F. solani UCP1083 (73.4%), and T. asperellum UCP 0168 against F. solani UCP1098 (71.5%), respectively. The data obtained could serve as the basis for developing several biotechnological processes of safe use.

Ticks are key vectors of some important diseases of humans and animals. Although they are carriers of disease agents, the viability and development of ticks are not harmed by the infectious agents due to their innate immunity. Antimicrobial peptides directly protect hosts against pathogenic agents such as viruses, bacteria, and parasites. Among the identified and characterized antimicrobial peptides, defensins have been considerably well studied. Defensins, which contain intramolecular disulfide bridges between cysteine residues, are commonly found among fungi, plants, invertebrates, and vertebrates. The sequence of the tick hemolymph defensin (HEdefensin) gene from the hard tick Haemaphysalis longicornis was analyzed after identification and cloning from a cDNA library. HEdefensin has a predicted molecular mass of 8.15 kDa and a theoretical isoelectric point of 9.48. Six cysteine residues were also identified in the amino acids. The synthetic HEdefensin peptide only showed antibacterial activity against Gram-positive bacteria such as Micrococcus luteus. A fluorescence propidium iodide exclusion assay also showed that HEdefensin increased the membrane permeability of M. luteus. Additionally, an indirect fluorescent antibody test showed that HEdefensin binds to M. luteus. These results suggested that HEdefensin strongly affects the innate immunity of ticks against Gram-positive bacteria.

Many marine animals use chemicals to defend themselves and their eggs from predators. Beyond their ecologically relevant functions, these chemicals may also have properties that make them beneficial for humans, including with biomedical and industrial applications. For example, some chemical defenses are also powerful antimicrobial or anti-tumor agents with relevance to human health and disease. One such chemical defense, Escapin, an L–amino acid oxidase in the defensive ink of the sea hare Aplysia californica, and related proteins have been investigated for their biomedical properties. This review details our current understanding of Escapin’s antimicrobial activity, including the array of chemicals generated by Escapin’s oxidation of its major substrates, L–lysine and L–arginine, and mechanisms underlying these molecules’ bactericidal and bacteriostatic effects on planktonic cells and the prevention of formation and removal of bacterial biofilms. Models of Escapin’s effects are presented, and future directions are proposed.

In most industrialized countries, screening programs for cervical cancer have shifted from cytology (Pap smear or ThinPrep) alone on clinician-obtained samples to the addition of screening for human papillomavirus (HPV), its main causative agent. For HPV testing, self-sampling instead of clinician-sampling has proven to be equally accurate, in particular for assays that use nucleic acid amplification techniques. In addition, HPV testing of self-collected samples in combination with a follow-up Pap smear in case of a positive result is more effective in detecting precancerous lesions than a Pap smear alone. Self-sampling for HPV testing has already been adopted by some countries, while others have started trials to evaluate its incorporation into national cervical cancer screening programs. Self-sampling may result in more individuals willing to participate in cervical cancer screening, because it removes many of the barriers that prevent women, especially those in low socioeconomic and minority populations, from participating in regular screening programs. Several studies have shown that the majority of women who have been underscreened but who tested HPV-positive in a self-obtained sample, will visit a clinic for follow-up diagnosis and management. Additionally, a self-collected sample can also be used for vaginal microbiome analysis, which can provide additional information about HPV infection persistence as well as vaginal health in general.

Rice is currently the most important food crop in the world and we are only just beginning to study the bacterial associated microbiome. It is of importance to perform screenings of the core rice microbiota and also to develop new plant-microbe models and simplified communities for increasing our understanding about the formation and function of its microbiome. In order to begin to address this aspect, we have performed the isolation of bacterial strains from the endorhizosphere of two rice cultivars from Venezuela. The validation of plant-growth promoting bacterial activities in vitro has led us to select and characterize 15 isolates for in planta studies such as germination test, endophytism ability and plant growth promotion. Consequently, a set of 10 isolates was selected for the set-up of an endophytic consortium as a simplified model of the natural rice bacterial endomicrobiota. Upon inoculation, the colonization and abundance of each strain within the rice roots was tracked by a culture-independent technique in gnotobiotic conditions in a 30 days period. Four strains belonging to Pseudomonas, Agrobacterium and Delftia genera have shown a promising capacity for colonizing and coexistence in root tissues. On the other hand, a bacterial community taxonomic profiling of the rhizosphere and the endorhizosphere of both cultivars were obtained and are discussed. This study is part of a growing body of research on core crops microbiome and simplified microbiomes, which strengthens the formation process of the endophytic community leading to a better understanding of the rice microbiome.

Although the European reports highlight an increase in community-acquired Legionnaires’ disease cases, the risk of Legionella spp. in private houses is underestimated. In Pisa (Italy) we performed a three-year survey on Legionella presence in 121 buildings with an independent hot water production (IB); 64 buildings with a central hot water production (CB); and 35 buildings with a solar thermal system for hot water production (TB). From all the 220 buildings Legionella spp. was researched in two hot water samples collected either at the recirculation point or at on the first floor and on the last floor, while the potable water quality was analyzed in three cold water samples collected at the inlet from the aqueduct network, at the exit from the autoclave, and at the most remove remote? tap. Legionella pneumophila sg1, Legionella pneumophila sg2-16 and not-pneumophila Legionella species were detected in 26% of the hot water networks, mostly in CB and TB. In these buildings we detected correlations between the presence of Legionella and the total chlorine concentration decrease or/and the increase of the temperature. Cold water resulted free from microbiological hazards, with the exception of Serratia liquefaciens and Enterobacter cloacae isolated at the exit from two different autoclaves. We observed an increase in total microbial counts at 22 and 37°C between the samples collected at the most remote taps compared to the ones collected at the inlet from the aqueduct. The study highlights a condition of potential risk for susceptible categories of population and supports the need for measures of risk assessment and control.

By providing useful tools to study host-pathogen interactions, next-generation omics has recently enabled the study of gene expression changes in both pathogen and infected host simultaneously. However, since great discriminative power is required to study pathogen and host simultaneously throughout the infection process, the depth of quantitative gene expression profiling has proven to be unsatisfactory when focusing on bacterial pathogens, thus preferentially requiring specific strategies or the development of novel methodologies based on complementary omics approaches. In this review, we focus on the difficulties encountered when making use of omics approaches to study bacterial pathogenesis. Besides, we review different omics strategies (i.e. transcriptomics, proteomics and secretomics) and their applications for studying interactions of pathogens with their host.

Italian Directives recommends the good quality of natural mineral waters but literature data assert a potential risk from several microorganisms colonizing wellsprings and mineral water bottling plants. Aim of study is the identification of microorganisms from spring waters (SW) and bottled mineral waters (BMW) samples. Methods: Routine microbiological indicators, Legionella spp., Nontuberculous mycobacteria (NTM), protozoa (FLA) and physical-chemical parameters were assessed in 24 SW and 10 BMW samples performing culture methods and molecular tests as PCR and qPCR. Results: In 33 out of 34 samples no cultivable bacteria were isolated with the exception of 83 CFU/L of Mycobacterium gilvum, detected in one warm rich-mineralized SW. qPCR showed the presence of Legionella genomic units in 24% of samples (mean 2,9x10²±1,7x10² GU/L) and NTM genomic units in 18% of samples (mean 5,7x10^3±4,1x10³ GU/L). Vermamoeba vermiformis and Acanthamoeba polyphaga were recovered respectively in 70% of BMW samples (counts from 1,3x10³ to 1,2x10⁵) and 42% of SW samples (counts from 1,1x10³ to 1,3x10⁴). Vahlkampfia spp. was detected in 42% of SW and 70% of BMW samples (mean 1,3x10⁴ ±2,9x10³ GU/L). Conclusion: The study highlights a low rate of microbial risk and the importance of risk assessment in natural mineral waters.

Agavins consumption has lead to accelerate body weight loss in mice. We investigated the changes on cecal microbiota and short chain fatty acids (SCFA) associated to body weight loss in overweight mice. Firstly, mice were fed with standard (ST5) or high fat (HF5) diet for 5 weeks. Secondly, overweight mice were shifted to standard diet alone (HF-ST10) or supplemented with agavins (HF-ST+A10) or oligofructose (HF-ST+O10), five more weeks. Cecal contents were collected before and after supplementation to determine microbiota and SCFA concentrations. At the end of first phase, HF5 mice showed a significant increase of body weight, which was associated with reduction of cecal microbiota diversity (PD whole tree; non-parametric t-test, P < 0.05), increased Firmicutes/Bacteroidetes ratio and reduced SCFA concentrations (t-test, P < 0.05). After diet shifted, HF-ST10 normalized its microbiota, increase its diversity and SCFA levels, whereas agavins (HF-ST+A10) or oligofructose (HF-ST+O10) led to partial microbiota restoration, with normalization of the Firmicutes/Bacteroides ratio as well as higher SCFA levels (P < 0.1). Moreover, agavins noticeably enriched Klebsiella and Citrobacter (LDA > 3.0); this enrichment has not been reported previously under a prebiotic treatment. In conclusion, agavins or oligofructose modulated cecal microbiota composition, reduced extent of diversity and increased SCFA. Furthermore, identification of bacteria enriched by agavins, opens opportunities to explore new probiotics.

A bioelectrochemical study of charge transfer in the biofilm/chalcopyrite interface was performed to investigate the effect of surficial sulfur reduced species (SRS), as non-stochiometric compounds or polysulfides (S_n^2-), and elemental sulfur (S⁰) on a biofilm structure during the earliest stages (1, 12 and 24 h) of chalcopyrite biooxidation by A. thiooxidans alone and adding Leptospirillum sp. The surface of massive chalcopyrite electrodes was exposed to the bacteria, which were analyzed electrochemically, spectroscopically, and microscopically. At the studied earlier times, charge transfer and significant differences in the biofilm structure were detected, depending on the presence of Leptospirillum sp. acting on A. thiooxidans biofilms. Such differences were a consequence of a continuous chalcopyrite pitting and promoting changes in biofilm hydropathy. A. thiooxidans modifies the reactive properties of SRS and favors an acidic dissolution, which shifts into ferric when Leptospirillum sp. is present. A. thiooxidans allows H⁺ and Fe³⁺ diffusion, and Leptospirillum sp. allows surpassing the charge transfer (reactivity) barrier between the mineral interface and the ions. The observed changes of hydropathy on the interface are associated to ions and electrons activity and transfer. Finally, a model of S⁰ biooxidation by A. thiooxidans alone or with Leptospirillum sp., is proposed.

Staphylococcus aureus bacteria are one of the leading microorganisms responsible for nosocomial infections as well as being the primary causative pathogen of skin and wound infections. Currently, the therapy of staphylococcal diseases faces many difficulties, due to a variety of mechanisms of resistance and virulence factors. Moreover, a number of infections caused by S. aureus is connected with biofilm formation that impairs effectiveness of the therapy. Short cationic lipopeptides that are designed on the basis of the structure of antimicrobial peptides structure are likely to provide a promising alternative to conventional antibiotics. Despite the fact that many research groups have proved a high antistaphylococcal potential of lipopeptides, the lack of unified protocols in determination of antimicrobial activity may be the reason of inconsistency of the results. The aim of this study was to learn how the use of various bacteriological media as well as solvents may affect activity of lipopeptides and their cyclic analogs.

A novel binary toxin-positive non-027, non-078 Clostridium difficile strain designated LC693 whose sequence type was ST201 was isolated from the fecal sample of a patient with severe diarrhea in China. To understand the pathogenesis basis of C. difficile ST201, this recently recovered isolate LC693 was then chosen for whole genome sequencing. The project finally generated an estimated genome size of approximately 4.07 Mbp. The genome sequence was then analyzed together with the other two ST201 strains VL-0104 and VL-0391 and compared to the epidemic 027/ST1 and 078/ST11 strains. Phylogenetic analysis demonstrated that the ST201 strains belonged to clade 3. Genome size of the three ST201 strains ranged from 4.07 Mb~4.16 Mb, with an average GC content between 28.5%~28.9%. The ST201 genomes contained more than 40 antibiotic resistance genes and 15 of them were predicted to be associated with vancomycin-resistance, suggesting that they may have a strong antibiotic resistance. The ST201 strains contained a typical clade 3 specific PaLoc with a Tn6218 element inserted, and those genes harbored on their PaLoc that participated in the toxin expression and regulation were highly homologues to the epidemic 027 and 078 strains, with the exception of tcdC. A truncated TcdC was found in the ST201 strains, which is suggestive to have a contribution to the toxin production of the ST201 strains. In addition, the ST201 strains contained intact binary toxin coding and regulation genes, which is also proposed to contribute to the virulence. Genome comparison of the ST201 strains with the epidemic 027 and 078 strain identified 641 genes specific for the C. difficile ST201, and a number of them were predicted as fitness and virulence associated genes. The identification of those genes also contributes to the pathogenesis of the ST201 strain. To our knowledge, this is the first study that the genome sequence of C. difficile ST201 was discussed in detail, and the present study would have a contribution to understanding the pathogenesis basis of C. difficile ST201.

Water borne pathogens present a threat to human health and their disinfection from water poses a challenge, prompting search for newer methods and newer materials. Disinfection of Gram-negative bacterium Escherichia coli and Gram-positive coccal bacterium Staphylococcus aureus in aqueous matrix was achieved within 60 and 90 minutes respectively at 35⁰C using solar-photocatalysis mediated by sonochemically synthesized [email protected] core-shell nanoparticles. The efficiency of the process increased with increase in temperature and at 55⁰C the disinfection could be achieved in 45 and 60 min respectively for the two bacteria. A new ultrasound assisted chemical precipitation technique was used for the synthesis of [email protected] core-shell nanoparticles. The characteristics of the synthesized material were established using physical techniques. The material remained stable even at 400o C. Disinfection efficiency of the [email protected] core-shell nanoparticles was confirmed in case of real world water samples from pond, river, municipal tap and was found to be better than that of pure ZnO and TiO2 (Degussa P25). When the nanoparticle based catalyst was recycled and reused for subsequent disinfection experiments, its efficiency did not change remarkably even after three cycles. The sonochemically synthesized [email protected] core-shell nanoparticles have a good potential for application in solar photocatalytic disinfection of water borne pathogens.

Flavonoids, compounds present in many dietary supplements, affect growth of different bacterial species when tested as purified or synthetic substances. Here, we asked if soy isoflavone extracts, commonly used in many products sold as anti-menopausal dietary supplements, influence bacterial growth similarly to synthetic isoflavone, genistein. Four commercially available products were tested in amounts corresponding to genistein concentrations causing inhibition of growth of Vibrio harveyi (a model bacterium sensitive to this isoflavone) and Escherichia coli (a model bacterium resistant to genistein). Differential effects of various extracts on V. harveyi and E. coli growth, from stimulation, through no changes, to inhibition, were observed. Moreover, contrary to genistein, tested extracts caused a decrease (to different extends) in viability of human dermal fibroblasts. These results indicate that effects of various soy isoflavone extracts on bacterial growth and viability of human cells are different, despite similar declared composition of the commercially available products.

In this study, we evaluated the physicochemical and microbial qualities of source and stored household waters in some communities in Southwestern Nigeria using standard methods. Compared parameters include physicochemical constituents; Temperature (T), pH, Total Dissolved Solids (TDS), Total Hardness (TH), Biological Oxygen Demand (BOD), Magnesium ion (Mg²⁺) and Calcium ion (Ca²⁺) and microbiological parameters included Total Coliform Counts (TC), Faecal Coliform Counts (FC), Fungal Counts (Fung C), Heterotrophic Plate Counts (HPC). Comparing Stored and Source samples, the mean values of some physicochemical parameters of most of the stored water samples significantly (P<0.05) exceeded that of Sources and ranged in the following order: T (15.3±0.3^oC - 28.3±0.5^oC), pH (6.4±0.1 - 7.6±0.1), TDS (192.1±11.1 ppm - 473.7±27.9 ppm), TH (10.6±1.7 mg/L - 248.6±18.6 mg/L), BOD (0.5±0.0 mg/L - 3.2±0.3 mg/L), Mg²⁺ (6.5±2.4 mg/L - 29.1±3.2 mg/L) and Ca²⁺ (6.5±2.4 mg/L - 51.6±4.4 mg/L). The mean microbial counts obtained from microbial comparison of different points (Stored and Source) of collection showed that most of the stored water had counts significantly exceeding (P<0.05) those of the source water samples (cfu/100 mL) which ranged as follows: TC (3.1±1.5 - 156.8±42.9), FC (0.0±0.0 - 64.3±14.2) and HPC (47.8±12.1 - 266.1±12.2) across all sampled communities. Also, the predominant isolates recovered from the samples were identified as Escherichia coli, Klebsiella pneumonia, Pseudomonas aeruginosa, Enterobacter aerogenes, Aspergillus spp, Mucor spp, Rhizopus spp and Candida spp. The presence of these pathogenic and potentially pathogenic organisms in the waters and the high counts of the indicator organisms suggest the waters to be a threat to public health.

The development of low-cost, open-source Remotely Operated Vehicle (ROV) systems has provided almost unrestricted access for researchers looking to monitor the marine environment in ever greater resolution. Sampling microbial communities from the marine environment, however, still usually relies on Niskin-bottle sampling (ROV or CTD based), a method which introduces an inaccuracy and variability that is incompatible with metatranscriptomic analysis. Here, we describe a versatile, easily-replicated platform which achieves in situ mRNA preservation, via the addition of RNAlater to filtered microbial cells, to enhance ROV or CTD functionality.

Objectives: To evaluate the effectiveness of hydrogen peroxide (HP) use in the hospital water network disinfection to control Legionella spp. colonization.
Methods: Following the detection of high levels of Legionella contamination in a 136-bed general hospital water network, an HP treatment of the hot water (25 mg/L) was adopted. During a period of 34 months, the effectiveness of HP on Legionella colonization was assessed. Legionella was isolated in accordance with ISO-11731 and identification was carried out by sequencing of the mip gene.
Results: Before HP treatment L.pneumophila sg 2-15 was isolated in all sites with a mean count of 9950±8279 CFU/L. After one month of HP-treatment, we observed the disappearance of L. pneumophila 2-15, however other Legionella species never cultured before appeared; Legionella pneumophila 1 was isolated in 1 out of 4 sampling sites (2,000 CFU/L) and other non-pneumophila species in all sites (mean load 3,000 ±2887 CFU/L). Starting from September 2013, HP-treatment was modified adding food-grade polyphosphates and in the following months we observed a progressive reduction of the mean load of all species (p<0.05), until to a substantial disappearing of Legionella colonization.
Conclusion: Hydrogen peroxide demonstrated a good efficacy in controlling Legionella. Although in the initial phases of treatment it seemed unable to eliminate all the species, by keeping HP levels to 25 mg/L and adding food-grade polyphosphates, a progressive and complete control of colonisation was obtained.

Pathogenic Salmonella strains have a set of virulence factors allowing them to generate systemic infections and damage in a variety of hosts. Among these factors, bacterial proteins secreted by specialized systems are used to penetrate the host’s intestinal mucosa, through the invasion and destruction of specialized epithelial M cells in the intestine. On the other hand, numerous studies have demonstrated that humans, as well as experimental animal hosts, respond to Salmonella infection by activating both innate and adaptive immune responses. Here, through live cell imaging of S. Typhimurium infection of zebrafish larvae, we showed that besides the intestinal colonization, a deformed cloacae region and a concomitant accumulation of S. Typhimurium cells was observed upon bacterial infection. The swelling led to a persistent inflammation of infected larvae, although the infection was non-lethal. The in vivo inflammation process was confirmed by the co-localization of GFP-tagged S. Typhimurium with mCherry-tagged neutrophils at 72 hours post exposition. Our live-cell analyses suggest that Salmonella Typhimurium induce cloacitis-like symptoms in zebrafish larvae.  Typhimurium induce cloacitis-like symptoms in zebrafish larvae.

Background: Exposure to chronic low levels of aflatoxin B1 (AFB1) contamination can lead to immune suppression and nutritional consequences that might greatly contributed in the increase of hepatocellular carcinoma (HCC). The toxicity of AFB1 is greatly vary between different population, affected by age, gender, and environmental factors. Material and subjects: Aflatoxin B1 (AFB1) was measured in 50 blood samples collected from non B, C hepatitis viruses and non CMV-Ab liver disease patients from different general hospitals and polyclinic in KSA during period 01-2013 to 06-2014. All Patients demonstrate elevation of ALT and AST with unknown etiology. Serum samples were obtained and kept at −20 °C for AFB1detection. Results: Out of the 50 blood samples, 38 demonstrate a detectable serum level of AFB1 while the remaining 12 patients were AFB1 negative and used as control participants. While AST was non-significantly different in AFB1 exposed patients, ALT was significantly higher in AFB1 positive samples compared to control AFB1-negative. AFB1 was positively correlated with AST and ALT as liver function enzymes and with age as a risk factor of long duration of AFB1 chronic exposure. Multiple linear regression analysis ascertained the association between AFB1 chronic exposure and ALT increase in liver dysfunction Saudi patients. Conclusion: Measurement of elevated ALT as marker of liver injury in AFB1 chronically exposed Saudi patients can help to avoid the future development of HCC. Moreover, early detection of AFB1 exposure, together with early vaccination against HBV and HCV can remove the synergistic effects of these two etiological factors and thus decrease the risk of developing liver cancer.

Application of toxic antibacterial agents is considered necessary to control prevalent fresh water microorganisms in evaporative cooling water systems, but these agents can adversely affect the environment and human health. Alternatively, natural antibacterial water chemistry has been applied in industrial cooling water systems for over 10 years with excellent results. The tower water chemistry method concentrates natural salts in highly-softened water to produce elevated pH and dissolved solids, with low calcium and magnesium. This practice conserves water while generating only a small volume of non-toxic natural salt concentrate for cost efficient separation and disposal if required. This review presents a novel perspective of natural antimicrobial chemistry for inhibiting parasitic microbiome functional relationships within the bio-triad of Legionella outbreaks, "Trojan Protozoans" and biofilms. The review further examines practical application and function of polyvalent metal ions in the inhibition of biofilms. Reducing global dependence on toxic antibacterial agents discharged to the environment is an emerging concern due to their impact on the natural microbiome, plants, animals and humans. Discharge of antibacterial agents also contributes to development of pathogen resistance. Use of natural antibacterial chemistry can play a key role in managing the cooling water environment in a more ecologically sustainable manner.

In 2016, two special issues have been launched and attended in this journal, "Challenges in New Technologies for Security" (http://www.mdpi.com/journal/challenges/special_issues/tech_security) and “Food Microbiology: Technologies and processes, microbiology analysis methods, and antimicrobials” (http://www.mdpi.com/journal/challenges/special_issues/food-microbiology). In this editorial, I will review one of the topics of biological hazards detection, i.e. the rapid analysis of biological agents (DNA and antigens of fungi, bacteria, and viruses) and their toxins.

Consumption of refined foods and beverages high in sugar make the teeth susceptible to the formation of biofilm, and lead to dental caries and diseases of the oral cavity such as periodontitis and periodontal disease. The aim of the present study was to determine the ability of selected probiotics to inhibit growth and biofilm formation by the cariogenic bacterium Streptococcus mutans in vitro. We screened strains of latic acid bacteria (LAB) (n=120) from the Bioresources Collection and Research Center (BCRC), saliva of healthy adults and infant stool. The antimicrobial activity of LAB in vitro was evaluated by agar spot culture and co-culture of the S. mutans strains. We determined the effect of heating and dilution factors (2- and 4-fold) on the antagonistic activity of LAB. Antagonistic substances in the spent culture suspensions (SCS) of LAB were precipitated by extraction with ammonium sulphate and chloroform to characterise the protein and lipophilic fractions. Results of co-culturing show that the SCS of the three LAB strains (Lactobacillus pentosus 13-1, 13-4 and Lactobacillus crispatus BCRC 14618) subjected to heat treatment showed significantly high antimicrobial activity. We found that substances produced by L. pentosus 13-4 which have the potential to exhibit antimicrobial properties might be lipophilic proteins. Additionally, we infer that the mechanism of reducing biofilm formation by Lactobacillus strains is associated with sucrose-dependent cell–cell adhesion and the gtfC level of glucosyltransferases (Gtfs) in the biofilm. Native LAB strains screened in our study may be used in chewing gums and other processed foods for preventing tooth decay.

Current knowledge of the RNA world indicates two different genetic codes being present throughout the living world. In contrast to non-coding RNAs that are built of repetitive nucleotide syntax, the sequences that serve as templates for proteins share – as main characteristics – a non-repetitive syntax. The differences in their syntax structure is coherent with the difference of the functions they represent. Whereas non-coding RNAs build groups that serve as regulatory tools in nearly all genetic processes, the coding sections represent the evolutionarily successful function of the genetic information storage medium. The DNA genomes themselves are rather inactive, whereas the non-coding RNA domain is highly active, even as non-random genetic innovation operators. This indicates that repetitive syntax is the essential pre-requisite for RNA interactions to install variable RNA-group-identities, whereas the non-repetitive syntax serves as a stable conservation tool for successful selection processes out of RNA-groups cooperation and competition. The interaction opportunities of RNA loops with repetitive syntax are higher than with non-repetitive ones. Interestingly, these two genetic codes resemble the function of all natural languages, i.e., (a) everyday language use for organization and coordination of biotic group behavior, and (b) artificial (instrumental) language use for conservation of blueprints for complex protein-body constructions.

Acellular bacterial ghosts (BGs) are empty non-living bacterial cell envelopes, commonly generated by controlled expression of the cloned lysis gene E of bacteriophage PhiX174. In this study, Vibrio parahaemolyticus ghosts (VPGs) were generated by chemically induced lysis and the method is based on minimum inhibitory concentration (MIC) of sodium hydroxide (NaOH), acetic acid, boric acid, citric acid, maleic acid, hydrochloric acid and sulfuric acid. The MIC values of the respective chemicals were 3.125, 6.25, < 50.0, 25.0, 6.25, 1.56 and 0.781 mg/ml. Except boric acid, the lysis efficiency was reached more than 99.99% at 5 min after treatment of all chemicals. Among those chemicals, NaOH-induced VPGs showed completely DNA-free that was confirmed by quantitative real-time PCR. Besides, lipopolysaccharides (LPS) extracted from the NaOH-induced VPGs showed no distinctive band on SDS-PAGE gel after silver staining. On the other hand, LPS extracted from wild-type bacterial cells as well as the organic acids-induced VPGs showed triple major bands and LPS extracted from the inorganic acids-induced VPGs showed double bands. It suggests that some surface structures in LPS of the NaOH-induced VPGs may be lost, weakened or modified by the MIC of NaOH. Nevertheless, Limulus amoebocyte lysate assay revealed that there is no significant difference in endotoxic activity between the NaOH-induced VPGs and wild-type bacterial cells. Macrophages exposed to the NaOH-induced VPGs at 0.5 × 106 CFU/mL showed cell viability of 97.9%, however the MIC of NaOH did not reduce the cytotoxic effect of wild-type bacterial cells. Like Escherichia coli LPS, the NaOH-induced VPGs are an excellent activator of pro-inflammatory cytokines (IL-1β and iNOS), anti-inflammatory cytokine (IL-10) and dual activities (IL-6) in the stimulated macrophage cells. On the other hand, the induction of TNF-α mRNA was remarkable in the macrophages exposed with wild-type cells. Scanning electron microscopy showed the formation of trans-membrane lysis tunnel structures in the NaOH-induced VPGs. SDS-PAGE and agarose gel electrophoresis also confirmed that cytoplasmic proteins and genomic DNA released from the VPGs to culture medium through the lysis tunnel structures. Taken together, all these results indicated that the NaOH-induced VPGs show the potency of safe, economical and effective inactivated bacterial vaccine candidate.

Adlay seed samples were collected from 3 adlay growing regions (Yeoncheon, Jeonnam and Eumseong regions) in Korea during 2012. Among all the samples collected, 400 seeds were tested for fungal occurrence by standard blotter and test tube agar methods and different taxonomic groups of fungal genera were detected. The most predominant fungal genera encountered were Fusarium, Phoma, Alternaria, Cladosporium, Curvularia, Cochliobolus and Leptosphaerulina. The occurrence of Fusarium species were 45.6% and based on the combined sequences of two protein coding genes, EF-1a, Beta-tubulin and phylogenetic analysis, 10 species were characterized as F. incarnatum (11.67%), F. kyushense (10.33%), F. fujikuroi (8.67%), F. concentricum (6.00%), F. asiaticum (5.67%), F. graminearum (1.67%), F. miscanthi (0.67%), F. polyphialidiom (0.33%), F. armeniacum (0.33%) and F. thapsinum (0.33%). The ability of these isolates to produce mycotoxins fumonisin (FUM) and zeralenone (ZEN) were tested by ELISA quantitative analysis method. The result revealed that fumonisin (FUM) was produced only by F. fujikuroi and zeralenone (ZEN) by F. asiaticum & F. graminearum. Mycotoxigenic species were then examined for their morphological characteristics to confirm their identity. Morphological observations of the species correlated well with their molecular identification and confirmed as F. asiaticum, F. fujikuroi and F. graminearum.

The Leguminosae (legume family) is divided into three sub-families, the Caesalpiniodeae, Mimosoideae and Papilionoideae. Here, the literature on legume-rhizobia symbioses was reviewed, and genotypically characterised rhizobia related to the taxonomy of the legumes they were isolated from. Only data from field soils were considered. The objective of the work was to assess to what extent legume specificity for rhizobial symbiont is related to legume taxonomy. Bradyrhizobium spp. were the exclusive rhizobial symbionts of species in the Caesalpinioideae but data are limited. Where tested, species within the two Mimosoideae tribes, Ingeae and Mimoseae were nodulated by different rhizobial genera. Generally, Papilionoideae species with indeterminate nodules were promiscuous in relation to rhizobial symbionts but high specificity for rhizobial partners appears to hold at tribe level for the Fabeae (Rhizobium spp.), genus level for Medicago (Ensifer spp.), Cytisus (Bradyrhizobium spp.) and Lupinus (Bradyrhizobium spp.), and species level for Galega spp. (Neorhizobium galegeae), Hedysarum coronarium (Rhizobium sullae), Cicer arietinum (Mesorhizobium spp.) and New Zealand native Sophora spp. (Mesorhizobium spp.). High legume specificity for rhizobial symbionts was linked to specific rhizobial symbiosis genes. For Papilionoideae species with determinate nodules, the Dalbergieae were primarily nodulated by Bradyrhizobium but were promiscuous with respect to Bradyrhizobium spp. while those in the Desmodieae, Phaseoleae, Psoraleae and Loteae were promiscuous across different rhizobial genera. Possible advantages and disadvantages of high specificity or promiscuity are discussed.