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Preprint ARTICLE | doi:10.20944/preprints201909.0142.v1

Chronic Kidney Disease–Associated Pruritus is Caused by Dysregulated Ion Channels in Sensory Neurons

Akishi Momose, Michihiko Yabe, Shigetoshi Chiba, Kenjirou Kumakawa, Yasuo Shiraiwa, Hiroki Mizukami

Subject: Medicine And Pharmacology, Urology And Nephrology Keywords: uremic pruritus; ion channels; cell signaling; Cav3.2 calcium channel; RT-PCR; skin

Online: 14 September 2019 (19:00:34 CEST)

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Preprint ARTICLE | doi:10.20944/preprints202311.1192.v1

Disposition Kinetics of Cathinone and Its Metabolites after Oral Administration in Rats

Fahad Y. Sabei, Ibrahim Khardali, Mohamed A. Al-Kasim, Emad S. Shaheen, Magbool Oraiby, Ahmad Alamir, David Banji, Saeed Alshahrani, Abdulmajeed Jali, Mohammad Attafi, Mohammed Y. Albeishy, Ibraheem Attafi

Subject: Medicine And Pharmacology, Pharmacology And Toxicology Keywords: Cathinone; Cathine; Kinetics; HPLC-IT/MS; Disposition; Organ-specific response; UPLC-QTOF/MS

Online: 20 November 2023 (07:36:02 CET)

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Cathinone is a natural stimulant found in Catha edulis and commonly used in new psychoactive substances. The objective of this study was to examine the disposition kinetics and metabolic profile of cathinone and its metabolite cathine through a single oral dose of cathinone administration in rats. Cathine and cathinone concentrations were identified and quantified using ion trap liquid chromatography (LC-IT/MS), while the metabolic profile was determined using the quadrupole time of flight UPLC-QTOF/MS method in the serum, brain, lung, liver, kidney, and heart over a period of 0, 0.5, 2.5, 6, 12, 24, 48, and 72 hours. The findings revealed a dynamic interplay between the pharmacokinetics of cathinone and its impact on organ-specific biomarkers. The distribution of cathinone and cathine was determined in serum, brain, heart, lung, kidney, and liver. The highest concentration of cathinone was found in the kidney at 1438.6 ug/L. It gradually decreased to 1.97 within 48 hours and disappeared after 72 hours. Within half an hour, cathinone levels in the lungs, liver, and heart were found to be 859, 798.9, and 385.8 ug/L, respectively. However, within 2.5 hours, these levels decreased to 608.1, 429.3, and 309.1 ug/L and kept decreasing until they became undetectable after 24 hours. The concentration of cathinone in the rat brain reduces quickly, dropping to undetectable levels within six hours, with levels decreasing from 712.7 ug/L after just 30 minutes. In the brain and serum, cathine reached its highest levels at 2.5 hours, while in other organs, it peaked at 0.5 hours. This indicates that the conversion of cathinone to cathine is slower in the brain and serum. The study found that cathinone's pharmacokinetics have a significant impact on organ-specific biomarkers. After administering the cathinone, there was a sharp change in biomarker levels, particularly in the liver and brain, during the initial hours. It is essential to investigate the correlation between the changes in biomarkers found in the brain and the levels of cathinone and cathine. The results have significant implications for drug development, pharmacovigilance, and clinical practices involving cathinone. In conclusion, this comprehensive analysis of cathinone's impact on organ-specific biomarkers provides a basis for informed decision-making in medical practices and further research into the cathinone's pharmacological properties.

Preprint ARTICLE | doi:10.20944/preprints202305.1201.v1

A New HPLC-UV Method Using Hydrolyzation with Sodium Hydroxide for Quantitation of Trans-p-hydroxycinnamic Acid and Its Esters in the Leaves of Ligustrum Robustum

Shi-Hui Lu, Xiao-Na Liang, Xiao-Jin Nong, Ran Chen, Xiu-Xia Li

Subject: Chemistry And Materials Science, Analytical Chemistry Keywords: trans-p-hydroxycinnamic acid; esters; Ligustrum robustum; HPLC-UV; hydrolyzation; sodium hydroxide; quantification

Online: 17 May 2023 (05:54:10 CEST)

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The leaves of Ligustrum robustum have been applied as Ku-Ding-Cha, a functional tea to clear heat and remove toxins; and served as a folk medicine to deal with diabetes, obesity, and hypertension. Trans-p-hydroxycinnamic acid and its esters, which were isolated from the leaves of L. robustum, might be a new resource to prevent diabetes and its complications because of their strong -glucosidase and fatty acid synthase (FAS) inhibitory activities and antioxidant effect. However, the quantitative analytical method for trans-p-hydroxycinnamic acid and its esters in the leaves of L. robustum hasn’t been reported so far. In addition, it was difficult and troublesome to analyze no less than 34 trans-p-hydroxycinnamic acid esters by usual HPLC. Therefore, a new HPLC-UV method using hydrolyzation with sodium hydroxide for quantitation of trans-p-hydroxycinnamic acid and its esters in the leaves of L. robustum was developed in the present study. The optimal hydrolyzation conditions were as follows: 1 mL sodium hydroxide aqueous solution (1 M) was added into 1 mL the extract of L. robustum and incubated at 80 °C for 2 h , and then 1 mL hydrochloride (1 M) solution was added. HPLC analysis was performed in reverse phase mode using a C-18 column, eluting with methanol-0.1% acetic acid aqueous solution (40:60, v/v) in isocratic mode at a flow rate of 1.0 mL/min, and detecting at 310 nm. The analytical method was simple and rapid, which simplified greatly the analytical process. The method was validated in terms of linearity, limit of detection (LOD), limit of quantification (LOQ), precision, and accuracy. High linearity was demonstrated with correlation coefficient of 1.000 for trans-p-hydroxycinnamic acid. The LOD and LOQ values were 1.55 and 4.69 μg/mL, respectively. The relative standard deviations (RSD) of intra-day and inter-day variabilities for trans-p-hydroxycinnamic acid were less than 2%. The percentage recovery of trans-p-hydroxycinnamic acid was 101.2% ± 0.8%. Finally, the proposed method was applied successfully to the determination and comparison of the contents of trans-p-hydroxycinnamic acid and its esters in various extracts of the leaves of L. robustum. The 60%-70% ethanol extracts of L. robustum showed the highest contents of free trans-p-hydroxycinnamic acid (3.96-3.99 mg/g), and the 50%-80% ethanol extracts of L. robustum displayed the highest contents of trans-p-hydroxycinnamic acid esters (202.6-210.6 mg/g). The method can be applied also to the quality control of the products of L. robustum.

Preprint ARTICLE | doi:10.20944/preprints202010.0475.v1

Antioxidant Properties of Lyophilized Rosemary and Sage Extract and its Effect to Prevent Lipid Oxidation In Poultry Pátê

Mirelli Bianchin, Daiane Pereira, Jacqueline Florio Almeida, Cristiane Moura, Rafaelly Simionatto Pinheiro, Leila Fernanda Serafini Heldt, Charles Windson Isidoro Haminiuk, Solange Teresinha Carpes

Subject: Biology And Life Sciences, Biochemistry And Molecular Biology Keywords: Rosmarinus officinalis; Salvia officinalis; natural antioxidant; antibacterial activity; total phenolic compounds; HPLC; TBARS.

Online: 23 October 2020 (09:48:36 CEST)

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Preprint ARTICLE | doi:10.20944/preprints201901.0041.v1

Metabolomics Characterizes the Effects and Mechanisms of Quercetin in the Nonalcoholic Fatty Liver Disease Development

Yan Xu, Jichun Han, Jinjin Dong, Xiangcheng Fan, Yuanyuan Cai, Jing Li, Tao Wang, Jia Zhou, Jing Shang

Subject: Biology And Life Sciences, Anatomy And Physiology Keywords: nonalcoholic fatty liver disease; high fat-sucrose diet; metabolomics; HPLC-QTOF-MS; quercetin

Online: 4 January 2019 (14:04:13 CET)

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Preprint ARTICLE | doi:10.20944/preprints202404.1845.v1

Determination Mitochondria's Quality Control of Parkinsonism Using Real-Time Reverse-Transcription Polymerase Chain Reaction

Al-Baraa Akram

Subject: Neuroscience And Neurology, Biology And Life Sciences Keywords: RT: real time, PCR: polymerase chain reaction, WT: wild type, dNTP: deoxynucleotide triphosphate

Online: 28 April 2024 (10:36:25 CEST)

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Nuclear DNA mutations, mitochondrial DNA mutations, combined nuclear and mitochondrial DNA defects, and random occurrences are the main causes of mitochondrial disease. Black pigmentation loss is a major sign of Parkinsonism. Because of the loss of dopaminergic neurons in the substantia nigra, it drops to less than 80%. The degree of motor impairment was correlated with the amount of black pigmentation loss. After RNA extraction, reverse transcription is carried out because reverse transcriptase was used to create cDNA. One microliter of total RNA supplemented with IORT Buffer, dNTPs, random hexamers, Multi-Scribe Reverse Transcriptase, and nuclease-free was used to create cDNA. The following was how the reaction was carried out in a thermal cycler: 10 minutes at 25°C, 120 minutes at 37°C, 5 seconds at 85°C, and lastly 4°C. The data was expressed as a v-value, which shows how much the target gene is enriched in comparison to the reference gene. Earlier research using mice injected with functional HtrA2/Omi protease activity revealed similar outcomes, including induced up-regulation of CHOP, decreased mitochondrial function, and early brain cell aging because of an increase in mtDNA deletions. The cell viability of the genotypes exhibits a dmg concentration-dependent effect in these results, suggesting that high content ratios of drug treatment with 6-OHDA may be the cause of cellular content depletion. After 6-OHDA neurotoxin treatment, CHOP is up-regulated. Prior in vitro research has shown that treatment with a model based on 6-OHDA neurotoxin causes the activation of mitochondrial stress pathways, which in turn promotes the death of neurons. 6-OHDA reversibly inhibited complexes I and IV's activities in rat brain cells, which led to dopaminergic neurodegeneration, a defining feature of PD pathogens. As a result, the number of living cells will directly correlate with the dmg concentration, supporting the results of the current study. Real-time PCR can generally demonstrate that the removal of HtrA 2 results in a significant up-regulation of Hsp60 expression when compared to WT cells. This could be linked to a decreased threshold for mitochondrial stress in HtrA2 KO cells, which would explain why, following ADEP4 and ACP5 treatment, Hsp60 is significantly up-regulated in HtrA2 KO cells compared to WT cells.

Preprint ARTICLE | doi:10.20944/preprints202404.0301.v1

Evaluation of Safety and Efficacy of Repeated Mesenchymal Stem Cell Transplantation in Patients with ALS by Investigating Patient's Specific MicroRNAs as Novel Biomarkers: A Retrospective Study

Zahraa Alkhazaali, Sajad Sahab-Negah, Amir Reza Boroumand, Najmeh Kaffash Farkhad, Mohammad Ali Khodadoust, Rashin Ganjali, Jalil Tavakol-Afshari

Subject: Medicine And Pharmacology, Neuroscience And Neurology Keywords: bone marrow-derived mesenchymal stem cells; ALS; microRNA; Mir206; Mir338-3p; RT -PCR.

Online: 3 April 2024 (14:25:24 CEST)

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Preprint ARTICLE | doi:10.20944/preprints202403.0205.v1

The Probiotic Paradox: Thriving in High-Hopped Sour Beer Bitter

Marcos Edgar Herkenhoff

Subject: Biology And Life Sciences, Agricultural Science And Agronomy Keywords: craft beer; probiotic; gene expression; PMA-qPCR; RT-qPCR; microbiological resistance to hops.

Online: 5 March 2024 (12:25:02 CET)

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Preprint ARTICLE | doi:10.20944/preprints202307.1362.v1

The Rise and Fall of Omicron BA.1 Variant as Seen in Wastewater Supports Epidemiological Model Predictions

Michal Liddor Naim, Yu Fu, Marilou Shagan, Itay Bar-Or, Robert Marks, Qun Sun, Rony Granek, Ariel Kushmaro

Subject: Public Health And Healthcare, Health Policy And Services Keywords: SARS-CoV-2; Omicron; Wastewater-based epidemiology; RT-qPCR; SIR model; SIRS model

Online: 20 July 2023 (11:44:24 CEST)

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Preprint REVIEW | doi:10.20944/preprints202104.0481.v1

Minimizing Errors in RT-PCR Detection and Quantification of SARS-CoV-2 RNA for Wastewater Surveillance

Warish Ahmed, Stuart Simpson, Paul Bertsch, Kyle Bibby, Aaron Bivins, Linda Blackall, Silvia Bofill-Mas, Albert Bosch, Joao Brandao, Phil Choi, Mark Ciesielski, Erica Donner, Nishita D'Souza, Andreas Farnleitner, Daniel Gerrity, Raul Gonzalez, John Griffith, Pradip Gyawali, Charles Haas, Kerry Hamilton, Chanditha Hapuarachchi, Valerie Harwood, Rehnuma Haque, Greg Jackson, Stuart Khan, Wesaal Khan, Masaaki Kitajima, Asja Korajkic, Giuseppina La Rosa, Blythe Layton, Erin Lipp, Sandra McLellan, Brian McMinn, Gertjan Medema, Suzanne Metcalfe, Wim Meijer, Jochen Mueller, Heather Murphy, Colleen Naughton, Rachel Noble, Sudhi Payyappat, Susan Petterson, Tarja Pitkanen, Veronica Rajal, Brandon Reyneke, Fernando Roman, Joan Rose, Marta Rusinol, Michael Sadowsky, Laura Sala-Comorera, Yin Xiang Setoh, Samendra Sherchan, Kwanrawee Sirikanchana, Wendy Smith, Joshua Steele, Rosalie Sabburg, Erin Symonds, Phong Thai, Kevin Thomas, Josh Tynan, Simon Toze, Janelle Thompson, Andy Whiteley, Judith Wong, Daisuke Sano, Stefan Wuertz, Irene Xagoraraki, Qian Zhang, Amity Zimmer-Faust, Orin Shanks

Subject: Environmental And Earth Sciences, Waste Management And Disposal Keywords: COVID-19; SARS-CoV-2; Wastewater; Surveillance; False-positive; False-negative; RT-PCR

Online: 19 April 2021 (13:08:13 CEST)

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Wastewater surveillance for pathogens using the reverse transcription-polymerase chain reaction (RT-PCR) is an effective, resource-efficient tool for gathering additional community-level public health information, including the incidence and/or prevalence and trends of coronavirus disease-19 (COVID-19). Surveillance of SARS-CoV-2 in wastewater may provide an early-warning signal of COVID-19 infections in a community. The capacity of the world’s environmental microbiology and virology laboratories for SARS-CoV-2 RNA characterization in wastewater is rapidly increasing. However, there are no standardized protocols nor harmonized quality assurance and quality control (QA/QC) procedures for SARS-CoV-2 wastewater surveillance. This paper is a technical review of factors that can lead to false-positive and -negative errors in the surveillance of SARS-CoV-2, culminating in recommendations and strategies that can be implemented to identify and mitigate these errors. Recommendations include, stringent QA/QC measures, representative sampling approaches, effective virus concentration and efficient RNA extraction, amplification inhibition assessment, inclusion of sample processing controls, and considerations for RT-PCR assay selection and data interpretation. Clear data interpretation guidelines (e.g., determination of positive and negative samples) are critical, particularly during a low incidence of SARS-CoV-2 in wastewater. Corrective and confirmatory actions must be in place for inconclusive and/or potentially significant results (e.g., initial onset or reemergence of COVID-19 in a community). It will also be prudent to perform inter-laboratory comparisons to ensure results are reliable and interpretable for ongoing and retrospective analyses. The strategies that are recommended in this review aim to improve SARS-CoV-2 characterization for wastewater surveillance applications. A silver lining of the COVID-19 pandemic is that the efficacy of wastewater surveillance was demonstrated during this global crisis. In the future, wastewater will play an important role in the surveillance of a range of other communicable diseases.

Preprint ARTICLE | doi:10.20944/preprints201912.0093.v1

Genome-Wide Association Study for Resistance to the Meloidogyne javanica Causing Root-Knot Nematode in Soybean

Jean Carlos Alekcevetch, André Luiz de Lima Passianotto, Everton Geraldo Capote Ferreira, Adriana Brombini dos Santos, Danielle Cristina Gregório da Silva, Waldir Pereira Dias, François Belzile, Ricardo Vilela Abdelnoor, Francismar Corrêa Marcelino-Guimarães

Subject: Biology And Life Sciences, Biochemistry And Molecular Biology Keywords: mixed linear model; genotyping-by-sequencing; functional validation; RT-qPCR; resistance genes; GWAS

Online: 7 December 2019 (12:41:39 CET)

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Preprint ARTICLE | doi:10.20944/preprints202402.0088.v1

Features and Advantages of the Recurrent Approximations of Retention Times in Reversed Phase High-Performance Kiquid Chromatography

Igor Georgievich Zenkevich, Abdennour Derouiche

Subject: Chemistry And Materials Science, Analytical Chemistry Keywords: Reversed-phase HPLC, retention times, recurrent approximation, detection of anomalies, chemical transformations of analytes.

Online: 1 February 2024 (15:27:36 CET)

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Preprint ARTICLE | doi:10.20944/preprints202103.0675.v1

Identification of Compounds with Potential Therapeutic Uses from Sweet Pepper (Capsicum annuum L.) Fruits and Their Modulation by Nitric Oxide (NO)

Lucía Guevara, María Ángeles Domínguez-Anaya, Alba Ortigosa, Salvador González-Gordo, Caridad Díaz, Francisca Vicente-Pérez, Francisco J. Corpas, José Pérez del Palacio, José M. Palma

Subject: Biology And Life Sciences, Anatomy And Physiology Keywords: Fruit ripening; gingerglycolipid A; HPLC-HRMS; melatonin; nitric oxide; phytosphingosin; quercetin; transcriptomics; L-tryptophan

Online: 26 March 2021 (15:44:51 CET)

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Plant species are precursors of a wide variety of secondary metabolites that, besides having useful activity for themselves, can also be used by humans for their consumption and economic benefit. Pepper (Capsicum annuum L.) fruit is not only a common food and spice source, but it also stands out for containing high amounts of antioxidants (such as vitamins C and A), polyphenols and capsaicinoids. Particular attention has been paid to capsaicin, whose anti-inflammatory, antiproliferative and analgesic activities, have been proven. Due to the potential interest in pepper metabolites for human use, in this project, we carried out an investigation to identify new bioactive compounds of this crop is carried out. To achieve this, we developed a metabolomic approach, using an HPLC (high-performance liquid chromatography) separative technique coupled to metabolite identification by high resolution mass spectrometry (HRMS). After chromatographic analysis and data processing against metabolic databases, 12 differential bioactive compounds were identified in sweet pepper fruits, including quercetin and its derivatives, L-tryptophan, phytosphingosin, FAD, gingerglycolipid A, tetrahydropentoxylin, blumenol C glucoside, colnelenic acid and capsoside A. The abundance of these metabolites varied depending on the ripening stage of the fruits, either immature green or ripe red. We also studied the variation of these 12 metabolites upon treatment with exogenous nitric oxide (NO), a free radical gas involved in a good number of physiological processes in higher plants such as germination, growth, flowering, senescence, and fruit ripening, among others. Overall, it was found that the content of the analysed metabolites was modulated by the ripening stage and by the presence of NO. The metabolic pattern followed by quercetin and its derivatives, as a consequence of the ripening stage and NO treatment, was also corroborated by transcriptomic analysis of genes involved in the synthesis of these compounds. This opens new research windows on the pepper fruit’s bioactive compounds with nutraceutical potentiality, where biotechnological strategies can be applied for optimizing the level of these beneficial compounds.

Preprint ARTICLE | doi:10.20944/preprints202311.1062.v1

Identification of Phylogenetically Conserved Nitrogen-Dependent Small RNA across Pseudomonas

Irina Yu Pozdnyakova-Filatova, Evgeniya V Ivanova, Kirill V Petrikov, Ainur Rezhepova, Alina A Frolova, Marina V Zakharova

Subject: Biology And Life Sciences, Biochemistry And Molecular Biology Keywords: small RNA; Pseudomonas; nitrogen; NtrC; gene expression; RT-qPCR; nitrogen metabolism; carbon/nitrogen homeostasis

Online: 16 November 2023 (10:34:33 CET)

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Preprint ARTICLE | doi:10.20944/preprints202311.0621.v1

Occurrence of Picobirnavirus in Domestic and Wild Animals from Three Cities of Brazilian Amazon

Elaine Hellen Nunes Chagas, Julia Rezende Da Silva, Bruno de Cássio Veloso de Barros, José Wandilson Barboza Duarte Júnior, Fabiolla da Silva dos Santos, Edivaldo Costa Sousa Júnior, Delana Andreza Melo Bezerra, Helder Henrique Costa Pinheiro, Ceyla Maria Oeiras de Castro, Yashpal Singh Malik, Joana D’Arc Pereira Mascarenhas

Subject: Biology And Life Sciences, Animal Science, Veterinary Science And Zoology Keywords: enteric virus; picobirnavirus; brazilian animals; page, rt-pcr; rdrp gene; phylogenetic analysis; genogroup I

Online: 9 November 2023 (11:24:00 CET)

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Preprint ARTICLE | doi:10.20944/preprints202103.0292.v1

Ke Zhou, Audrey Blanc-Lapierre, Valérie Seegers, Michèle Boisdron-Celle, Frédéric Bigot, Marianne Bourdon, Hakim Mahammedi, Aurélien Lambert, Mario Campone, Martine Bellanger, Jean-Luc Raoul

Subject: Medicine And Pharmacology, Immunology And Allergy Keywords: cancer; COVID-19; symptoms; healthcare workers; anosmia; dysgeusia; ageusia; France; serological test; RT-PCR

Online: 10 March 2021 (16:17:48 CET)

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Preprint REVIEW | doi:10.20944/preprints202006.0284.v1

Factors Affecting SARS-CoV-2 (COVID-19) Pandemic, including Zoonotic, Human Transmission and Chain of Infection. Reducing Public Health Risk by Serum Antibody Testing, Avoiding Screening in Unhygienic Places and False PCR Reporting. A Scientific Review

Kamran Mahmood Ahmed Aziz, Abdullah Othman, Waleed Alqahtani, Sumaiya Azhar

Subject: Biology And Life Sciences, Virology Keywords: SARS-CoV-2; RT-PCR; antibody; zoonotic; animal transmission; genomics; asymptomatic fraction; herd immunity

Online: 23 June 2020 (13:30:11 CEST)

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Since December 2019, a rapid increase in the number of SARS-CoV-2 (COVID-19) cases was reported worldwide, despite strict infection control and lock down measures. Current paper investigated the actual facts behind this rapid increase in the number of cases. Study of genomic sequence reveals that domestic and wild animals were likely ancestors and zoonotic source for SARS-CoVs, MERS-CoVs, and SARS-CoV-2. Strong evidence suggest that these viruses already existed and replicated in animals and humans during past several decades, exhibiting diverse mutations, evolutions and self-limiting diseases, except during outbreaks. Serious zoonotic reservoir investigations are required to investigate animal transmission of SARS-CoVs and SARS-CoV-2 to limit current pandemic. This might be the reason of increasing number of cases via animals. SARS-CoV-2 has been retrospectively isolated in different studies in August 2019, several months before Wuhan announced. Hence, there is a possibility that viruses existed, went undetected, infecting subclinically, in past several years, and SARS-CoV-2 antigens and neutralizing antibodies may have been present in humans since long time. This might be another reason of increasing number of cases by screening as mass screening and antigen or antibody testing was not carried out in the past years. Randomized controlled trials are required to investigate human to human transmission by touch, as the current evidence is limited with conflicting results. As all SARS-CoVs are basically respiratory viruses, droplet precautions and infection control measures are essential, especially for hospital staff. Increased number of SARS-CoV-2 asymptomatic, or subclinical cases are detected worldwide. This silent phase of transmission can be beneficial for humans. Lack of symptoms eventually lessen virus transmission and reduce the pathogen's long-term survival and provide humoral herd immunity up to several years. Hence, seropositivity with diverse antibodies develops against mutating SARS-CoVs which will confer strong immunity during epidemics. Strategies such as identification, contact tracing and quarantine are costly and practically difficult. Hence, asymptomatic persons can continue their work with droplet precautions and standard infection control procedures, while symptomatic or sick persons can isolate themselves in their homes without the need for strict quarantine until clinical recovery, with reduced hospital visits and minimizing chances of hospital acquired infections. RT-PCR has low sensitivity and specificity, carries a high risk of handling live virus antigens, and requires difficult protocols. As viral load also sharply declines after few days of onset of infection, this technique might overlook infection. Furthermore, SARS-CoV-2 infection may be present in blood when oropharyngeal swabs are negative by RT-PCR. Additionally, RT-PCR usually gives false negative and false positive results and must be interpreted cautiously. This might be again a reason of increasing number of cases by false positive RT-PCR reporting. Moreover, antibodies against SARS-CoVs develop robustly in serum even by reduced amount of antigens. In contrast to RT-PCR, ELISA for diagnosing antibodies against SARS-CoV-2 demonstrates 100% specificity and 100% sensitivity, even in clinically asymptomatic individuals. These antibodies can be used for serologic surveys, monitoring and screening. However, screening tests for SARS-COV-2 should be avoided in unhygienic public places by nasopharyngeal swabs, which carry a high risk of further transmission, co-infection or superinfection. Such highly infectious virus must be isolated and tested in highly sterilized laboratory. Further strict international laws and policies are required to stop the possible spread of experimental viruses, biological warfare and bioterrorism.

Preprint ARTICLE | doi:10.20944/preprints202004.0216.v1

Development of a Field-deployable RT-qPCR Workflow for COVID-19 Detection

Raphael Nyaruaba, Bo Zhang, Caroline Muema, Elishiba Muturi, Greater Oyejobi, Jin Xiong, Bei Li, Zhengli Shi, Caroline Mwaliko, Junping Yu, Xiaohong Li, Hongping Wei

Subject: Biology And Life Sciences, Immunology And Microbiology Keywords: COVID-19; SARS-CoV-2; field work; community; diagnosis; rapid detection; inactivation; RT-qPCR

Online: 14 April 2020 (08:41:04 CEST)

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Preprint ARTICLE | doi:10.20944/preprints202307.1907.v1

Solid-State Fermentation as a Sustainable Tool for Extracting Phenolic Compounds from Cascalote Pods

Francisco López-Cárdenas, Emilio Ochoa-Reyes, Ramiro Baeza-Jiménez, Julio C. Tafolla-Arellano, Juan A. Ascacio-Valdés, Jose J. Buenrostro-Figueroa

Subject: Biology And Life Sciences, Biology And Biotechnology Keywords: condensed tannins; hydrolysable tannins; ABTS; DPPH; solid-state fermentation; improved extraction; RP-HPLC-ESI-MS

Online: 27 July 2023 (10:46:06 CEST)

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Preprint ARTICLE | doi:10.20944/preprints202305.1364.v1

Exploring the Multi-Faceted Potential of Carob Leaves from Morocco: A Comprehensive Analysis of Polyphenols Profile, Antimicrobial Activity, Cytotoxicity Against Breast Cancer Cell Lines, and Genotoxicity

Elbouzidi Amine, Mohamed Taibi, Sabir Ouahhoud, Sabir Ouahhoud, Douâae Ou-Yahia, El Hassania Loukili, Marouane Aherkou, Farid Mansouri, Noureddine Bencheikh, Salah Laaraj, Reda Bellaouchi, Ennouamane Saalaoui, Kaoutar Elfazazi, Abdelbasset Berrichi, Malika Abid, Mohamed Addi

Subject: Biology And Life Sciences, Plant Sciences Keywords: Ceratonia siliqua L.; Morocco; HPLC-DAD; Phenolics; Antioxidant activity; Antimicrobial activitiy; cytotoxicity; breast cancer; genotoxicity

Online: 19 May 2023 (03:23:35 CEST)

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The botanical species Ceratonia siliqua L., commonly referred to as the Carob tree, and locally as “L’Kharrûb”, holds significance as an agro-sylvo-pastoral species, and is traditionally utilized in Morocco for treating a variety of ailments. This current investigation aims to ascertain the antioxidant, antimicrobial, and cytotoxic properties of the ethanolic extract of C. siliqua leaves (CSEE). Initially, we analyzed the chemical composition of CSEE through high-performance liquid chromatography with Diode-Array Detection (HPLC-DAD). Subsequently, we conducted various assessments, including DPPH scavenging capacity, β-carotene bleaching assay, ABTS scavenging, and total antioxidant capacity assays, to evaluate the antioxidant activity of the extract. In this study, we investigated the antimicrobial properties of CSEE against five bacterial strains (two gram-positive, Staphylococcus aureus, and Enterococcus faecalis; and three gram-negative bacteria, Escherichia coli, Escherichia vekanda, and Pseudomonas aeruginosa) and two fungi (Candida albicans, and Geotrichum candidum). Additionally, we evaluated the cytotoxicity of CSEE on three human breast cancer cell lines (MCF-7, MDA-MB-231, and MDA-MB-436) and assessed the potential genotoxicity of the extract using the comet assay. Through HPLC-DAD analysis, we determined that phenolic acids and flavonoids were the primary constituents of the CSEE extract. The results of the DPPH test indicated a potent scavenging capacity of the extract with an IC50 of 302.78 ± 7.55 µg/mL, which was comparable to that of ascorbic acid with an IC50 of 260.24 ± 6.45 µg/mL. Similarly, the β-carotene test demonstrated an IC50 of 352.06 ± 12.16 µg/mL, signifying the extract's potential to inhibit oxidative damage. The ABTS assay revealed IC50 values of 48.13 ± 3.66 TE µmol/mL, indicating a strong ability of CSEE to scavenge ABTS radicals, and the TAC assay demonstrated an IC50 value of 165 ± 7.66 µg AAE/mg. The results suggest that the CSEE extract had potent antioxidant activity. Regarding its antimicrobial activity, the CSEE extract was effective against all five tested bacterial strains, indicating its broad-spectrum antibacterial properties. However, it only showed moderate activity against the two tested fungal strains, suggesting it may not be as effective against fungi. The CSEE exhibited a noteworthy dose-dependent inhibitory activity against all the tested tumor cell lines in vitro. The extract did not induce DNA damage at the concentrations of 6.25, 12.5, 25, and 50 µg/ml, as assessed by the comet assay. However, the 100 µg/ml concentration of CSEE resulted in a significant genotoxic effect compared to the negative control. A computational analysis was conducted to determine the physicochemical and pharmacokinetic characteristics of the constituent molecules present in the extract. The Prediction of Activity Spectra of Substances (PASS) test was employed to forecast the potential biological activities of these molecules. Additionally, the toxicity of the molecules was evaluated using the Protox II webserver.

Preprint ARTICLE | doi:10.20944/preprints202312.0322.v1

Digital Twin of Microgrid for Predictive Power Control to Building

Hao Jiang, Rudy Tjandra, Chew Beng Soh, Shuyu Cao, Donny Cheng Lock Soh, Kuan Tak Tan, King Jet Tseng, Sivaneasan B. Krishnan

Subject: Engineering, Energy And Fuel Technology Keywords: Matlab/Simulink; load demands; microgrid; DER, OPAL-RT; digital twin; energy optimization; Gurobi; sustainable building

Online: 6 December 2023 (05:43:41 CET)

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Preprint ARTICLE | doi:10.20944/preprints202308.0372.v3

Machine Learning Tools Applied for Training Optimization in Handball

Radu Bucea Manea Tonis, Petre Sorin Savin, Laurentiu-Gabriel Talaghir, Veronica Mindrescu

Subject: Social Sciences, Tourism, Leisure, Sport And Hospitality Keywords: complex indicator; isolated action; weighting system; simultaneous action; machine learning(ML); decision tree(C-RT)

Online: 16 October 2023 (08:35:18 CEST)

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Preprint ARTICLE | doi:10.20944/preprints202307.1532.v1

Molecular and Clinical Epidemiology of SARS-CoV-2 Infection Among Vaccinated and Unvaccinated Individuals in a Large Healthcare Organization from New Jersey

José R. Mediavilla, Tara Lozy, Annie Lee, Justine Kim, Veronica W. Kan, Elizabeth Titova, Ashish Amin, Michael Zody, André Corvelo, Dayna M. Oschwald, Amy Baldwin, Samantha Fennessey, Jerry M. Zuckerman, Thomas Kirn, Liang Chen, Yanan Zhao, Kar Fai Chow, Tom Maniatis, David S. Perlin, Barry N. Kreiswirth

Subject: Biology And Life Sciences, Virology Keywords: SARS-CoV-2; COVID-19; pandemic; variants; RT-PCR; sequencing; surveillance; vaccination; vaccine breakthrough; comorbidity.

Online: 21 July 2023 (13:55:29 CEST)

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New Jersey was among the first states impacted by the COVID-19 pandemic, with one of the highest overall death rates in the nation. Nevertheless, relatively few reports have been published focusing specifically on New Jersey. Here we report on molecular, clinical, and epidemiologic observations from the largest healthcare network in the state, in a cohort of vaccinated and unvaccinated individuals with laboratory-confirmed SARS-CoV-2 infection. We conducted molecular surveillance of SARS-CoV-2-positive nasopharyngeal swabs collected in nine hospitals from December 2020 through June 2022, using both whole genome sequencing (WGS) and a real-time RT-PCR screening assay targeting spike protein mutations found in variants of concern (VOC) within our region. De-identified clinical data were obtained retrospectively, including demographics, COVID-19 vaccination status, ICU admission, ventilator support, mortality, and medical history. Statistical analyses were performed to identify associations between SARS-CoV-2 variants, vaccination status, clinical outcomes, and medical risk factors. A total of 5,007 SARS-CoV-2-positive nasopharyngeal swabs were successfully screened and/or sequenced. Variant screening identified three predominant VOC, including Alpha (n =714), Delta (n =1,877), and Omicron (n =1,802). Omicron isolates were further sub-typed as BA.1 (n =899), BA.2 (n =853), and BA.4/BA.5 (n =50); the remaining 614 isolates were classified as “Other”. Approximately 31.5% (1,577/5,007) of the samples were associated with vaccine breakthrough infections, which increased in frequency following the emergence of Delta and Omicron. Severe clinical outcomes included ICU admission (336/5007 = 6.7%), ventilator support (236/5007 = 4.7%), and mortality (430/5007 = 8.6%), with increasing age being the most significant contributor to each (p <0.001). Unvaccinated individuals accounted for 79.7% (268/336) of ICU admissions, 78.3% (185/236) of ventilator cases, and 74.4% (320/430) of deaths. Highly significant (p <0.001) increases in mortality were observed in individuals with cardiovascular disease, hypertension, cancer, diabetes, and hyperlipidemia, but not with obesity, thyroid disease, or respiratory disease. Significant differences (p <0.001) in clinical outcomes were also noted between SARS-CoV-2 variants, including Delta, Omicron BA.1, and Omicron BA.2. Vaccination was associated with significantly improved clinical outcomes in our study, despite an increase in breakthrough infections associated with waning immunity, greater antigenic variability, or both. Underlying comorbidities contributed significantly to mortality in both vaccinated and unvaccinated individuals, with increasing risk based on the total number of comorbidities. Real-time RT-PCR-based screening facilitated timely identification of predominant variants using a minimal number of spike protein mutations, with faster turnaround time and reduced cost compared to WGS. Continued evolution of SARS-CoV-2 variants will likely require ongoing surveillance for new VOCs, with real-time assessment of clinical impact.

Preprint ARTICLE | doi:10.20944/preprints202203.0040.v1

Augmented Sensitivity of At-Home Rapid SARS-CoV-2 Antigen Test (RAT) Kits with Computer Vision

Kim Miikki, Lars Miikki, Jenny Wiklund, Alp Karakoç

Subject: Biology And Life Sciences, Virology Keywords: SARS-CoV-2; rapid antigen test; RT-PCR test; COVID-19; image processing; Raspberry Pi

Online: 2 March 2022 (08:06:49 CET)

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Preprint ARTICLE | doi:10.20944/preprints202105.0381.v1

Building-Level Wastewater Monitoring for COVID-19 Using Tampon Swabs and RT-LAMP for Rapid SARS-Cov-2 RNA Detection

Aaron Bivins, Megan Lott, Marlee Shaffer, Zhenyu Wu, Devin North, Erin Lipp, Kyle Bibby

Subject: Environmental And Earth Sciences, Water Science And Technology Keywords: SARS-CoV-2; wastewater monitoring; environmental surveillance; RT-LAMP; building-level; near-source; passive sampling

Online: 17 May 2021 (10:07:04 CEST)

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Preprint ARTICLE | doi:10.20944/preprints202104.0688.v1

Direct Quantitation of SARS-CoV-2 Using Droplet Digital PCR in Suspected Samples With Very Low Viral Load

Michela Deiana, Chiara Piubelli, Antonio Mori, Gian Paolo Chiecchi, Giulia La Marca, Martina Leonardi, Davide Treggiari, Anna Beltrame, Lucia Moro, Zeno Bisoffi, Elena Pomari

Subject: Biology And Life Sciences, Biochemistry And Molecular Biology Keywords: droplet digital PCR; real time RT-PCR; SARS-CoV-2; false negative; viral load; diagnosis

Online: 26 April 2021 (20:09:26 CEST)

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Preprint REVIEW | doi:10.20944/preprints202004.0201.v1

Detecting the COVID-19

Pravin Pokhrel, Changpeng Hu, Hanbin Mao

Subject: Biology And Life Sciences, Virology Keywords: diagnosis; detection kits; RT-PCR; immunoassay; false negative; false positive; sensitivity; point-of-care (POC)

Online: 12 April 2020 (16:50:05 CEST)

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Preprint ARTICLE | doi:10.20944/preprints202109.0071.v1

Lysophospholipids, Lysophosphatidic Acids and Monoacylglycerols: New Therapeutic Targets in Cardiovascular Diseases?

Thomas Duflot, Ly Tu, Matthieu Leuillier, Hind Messaoudi, Déborah Groussard, Saïda Azhar, Raphaël Thuillet, Marc Humbert, Vincent Richard, Christophe Guignabert, Jérémy Bellien

Subject: Medicine And Pharmacology, Pharmacology And Toxicology Keywords: lysophospholipids; lysophosphatidic acids; cardiovascular diseases; HPLC-MS/MS; rodent models; pulmonary hypertension; chronic heart failure; hypertension

Online: 3 September 2021 (15:19:54 CEST)

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Preprint ARTICLE | doi:10.20944/preprints201906.0275.v1

Untargeted Metabolomics Reveals Molecular Effects of Ketogenic Diet on Healthy and Tumor Xenograft Mouse Models

David Licha, Silvia Vidali, Sepideh Aminzadeh-Gohari, Oliver Alka, Leander Breitkreuz, Oliver Kohlbacher, Roland Reischl, Roland Feichtinger, Barbara Kofler, Christian G. Huber

Subject: Biology And Life Sciences, Biochemistry And Molecular Biology Keywords: ketogenic diet; breast cancer; xenograft; untargeted metabolomics; HPLC-MS; reversed phase chromatography; hydrophilic interaction liquid chromatography

Online: 27 June 2019 (05:35:28 CEST)

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Preprint ARTICLE | doi:10.20944/preprints202304.0574.v1

Insights into the Genes Involved in ABA Biosynthesis and Perception during Development and Ripening of the Chilean Strawberry Fruit

Maria Alejandra Moya-León, Yazmina Stappung, Elena Mattus-Araya, Raúl Herrera

Subject: Biology And Life Sciences, Plant Sciences Keywords: ABA biosynthesis; ABA perception; Fragaria chiloensis; fruit ripening; NCED; PYR/PYL receptors; phylogenic analysis; RT-qPCR

Online: 19 April 2023 (08:43:03 CEST)

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Working Paper ARTICLE

Fast Detection of SARS-CoV-2 RNA Directly from Respiratory Samples Using a Loop-Mediated Isothermal Amplification (LAMP) Test

Olympia E. Anastasiou, Caroline Holtkamp, Miriam Schäfer, Frieda Schön, Anna Maria Eis-Hübinger, Andi Krumbholz

Subject: Medicine And Pharmacology, Immunology And Allergy Keywords: SARS-CoV-2; COVID-19; RT-PCR; nucleic acids; direct testing; loop-mediated isothermal amplification; LAMP

Online: 5 April 2021 (12:21:19 CEST)

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Preprint ARTICLE | doi:10.20944/preprints202209.0026.v1

Characterization of Polyphenolic Compounds from Bacopa Procumbens and Their Effects on Wound Healing Process

Adriana Martinez-Cuazitl, María del Consuelo Gómez-García, Oriana Hidalgo-Alegria, Olivia Medel-Flores, José Alberto Núñez-Gastélum, Eduardo San Martín Martínez, Ada María Ríos-Cortés, Mario Garcia-Solis, David Guillermo Pérez-Ishiwara

Subject: Chemistry And Materials Science, Medicinal Chemistry Keywords: Bacopa procumbens HPLC characterization; effect of polyphenolic compounds; in vitro and in vivo skin wound; collagen organization

Online: 1 September 2022 (15:44:09 CEST)

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Preprint ARTICLE | doi:10.20944/preprints201811.0548.v1

Characterization of Carotenoid and Carotenoid Esters of Astringent Persimmon Tissues (Diospyros kaki Thunb. var. Rojo Brillante). Effects of Thermal and High Pressure Non-Thermal Processing

M. Pilar Cano, Andrea Gómez-Maqueo, Jorge Welti-Chanes, Tomás García-Cayuela

Subject: Chemistry And Materials Science, Food Chemistry Keywords: Persimmon, Diospyrus kaki Thunb., var. Rojo Brillante, astringent, carotenoids, carotenoid esters, HPLC-PDA-MS (APCI+), processing effects

Online: 22 November 2018 (14:26:40 CET)

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Preprint BRIEF REPORT | doi:10.20944/preprints202105.0526.v1

Circulating SARS-CoV-2 Variants in Italy, October 2020-March 2021

Alessia Lai, Annalisa Bergna, Stefano Menzo, Gianguglielmo Zehender, Sara Caucci, Valeria Ghisetti, Francesca Rizzo, Fabrizio Maggi, Francesco Cerutti, Giorgio Giurato, Alessandro Weisz, Chiara Turchi, Bianca Bruzzone, Francesca Ceccherini-Silberstein, Nicola Clementi, Annapaola Callegaro, Fabio Sagradi, Daniela Francisci, Emmanuele Venanzi Rullo, Ilaria Vicenti, Massimo Clementi, Massimo Galli

Subject: Biology And Life Sciences, Biochemistry And Molecular Biology Keywords: SARS-CoV-2 virus; complete genome sequencing; COVID-19 RT-PCR testing; Spike protein; vi-ral variants

Online: 21 May 2021 (15:12:17 CEST)

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Preprint REVIEW | doi:10.20944/preprints202009.0526.v1

Nucleic Acid and Immunological Diagnostics for SARS-CoV-2: Processes, Platforms and Pitfalls

Avinash Premraj, Abi George Aleyas, Binita Nautiyal, Thaha Jamal Rasool

Subject: Biology And Life Sciences, Virology Keywords: COVID-19 testing; molecular diagnostics; immunological testing; RT-qPCR; ELISA; pool PCR; lateral flow assay; rapid assay

Online: 23 September 2020 (03:33:12 CEST)

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Preprint ARTICLE | doi:10.20944/preprints202006.0275.v1

Analysis of the Primary Presenting Symptoms and Hematological Findings of COVID-19 Patients in Bangladesh

Abu Taiub Mohammed Mohiuddin Chowdhury, Md Rezaul Karim, H.M. Hamidullah Mehedi, Mohammad Shahbaz, Md Wazed Chowdhury, Guo Dan, Shuixiang He

Subject: Medicine And Pharmacology, Hematology Keywords: SARS-CoV-2; COVID-19; real-time RT-PCR; COVID-19 symptoms; COVID-19 hematological findings; Bangladesh

Online: 21 June 2020 (14:47:03 CEST)

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Preprint ARTICLE | doi:10.20944/preprints202402.0181.v1

Whiter and Greener RP-HPLC Method for Simultaneous Determination of Dorzolamide, Brinzolamide, and Timolol Using Isopropanol as a Sustainable Organic Solvent in the Mobile Phase

Sami El Deeb, Khalid Abdelsamad, Maria Kristina Parr

Subject: Chemistry And Materials Science, Analytical Chemistry Keywords: green analytical chemistry; white analytical chemistry; sustainability; sustainable solvent; greenness; isopropanol; RGB12; AGREE; RP-HPLC; dorzolamide; brinzolamide; timolol

Online: 8 February 2024 (09:39:11 CET)

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Preprint ARTICLE | doi:10.20944/preprints202311.0120.v1

Agrobacterium Is a Proposed Suitable System for Bioassays and Resistance Studies on Antimicrobial Peptides with Plant Protection Potential

Ghazala M. Furgani, László Makrai, Dávid Vozik, János Kiss, Ferenc Olasz, Michael Gardner Klein, Muhamad-Akbar Bin-Abdul Ghaffar, László Szabados, Ahmed Noureldeen, Katalin Bélafi-Bakó, László Fodor, Steven A. Forst, András Fodor

Subject: Biology And Life Sciences, Biology And Biotechnology Keywords: Agrobacterium; Xenorhabdus; NR-AMP; T-DNA keyword; TI-plasmid; Intact/Cured/T-DNA Deleted; Sensitive/Resistant; EMA_PF2; HPLC

Online: 2 November 2023 (07:42:14 CET)

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Preprint ARTICLE | doi:10.20944/preprints202102.0211.v1

Polymer Identification and Specific Analysis (PISA) of Microplastic Total Mass in Sediments of the Protected Marine Area of the Meloria Shoals

Valter Castelvetro, Andrea Corti, Jacopo La Nasa, Francesca Modugno, Alessio Ceccarini, Stefania Giannarelli, Virginia Vinciguerra

Subject: Chemistry And Materials Science, Analytical Chemistry Keywords: microplastics; marine sediment; pet; nylon 6; nylon 6,6; reversed-phase HPLC; polyolefin; polystyrene; Pyr-GC/MS; polymer degradation

Online: 8 February 2021 (13:13:37 CET)

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Preprint ARTICLE | doi:10.20944/preprints202304.0971.v1

Diagnostic of Patients With COVID-19 Pneumonia Using Passive Medical Microwave Radiometry (MWR)

Berik Emilov, Aleksander Sorokin, Meder Seiitov, Tulegen Chubakov, Sergey Vesnin, Igor Goryanin

Subject: Medicine And Pharmacology, Pulmonary And Respiratory Medicine Keywords: 2019-nCoV; COVID-19; RT-PCR; SARS-CoV-2; Community-acquired pneumonia; Chest CT; microwave radiometry; temperature measurement

Online: 26 April 2023 (08:02:32 CEST)

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Preprint ARTICLE | doi:10.20944/preprints202308.2086.v1

Greener and Whiter Analytical Chemistry Using Cyrene as a More Sustainable and Eco-Friendlier Mobile Phase Solvent in Chromatography

Sami El Deeb, Khalid Abdelsamad, Maria Kristina Parr

Subject: Chemistry And Materials Science, Analytical Chemistry Keywords: green analytical chemistry; white analytical chemistry; global climate change; sustibability; sus-tinable solvent; greeneess; Cyrene, RGB12; AGREE; RP-HPLC

Online: 31 August 2023 (03:21:07 CEST)

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Preprint ARTICLE | doi:10.20944/preprints201804.0309.v1

Development of a Fabric Phase Sorptive Extraction-High Performance Liquid Chromatography-Ultraviolet Detection Method for the Analysis of Phenyltin Compounds in Environmental Water and Canned Food Samples

Heena N/A, Ashok Kumar Malik, Abuzar Kabir, Kenneth G. Furton

Subject: Chemistry And Materials Science, Analytical Chemistry Keywords: phenyltin derivatives; tetraphenyltin; triphenyltin chloride; triphenyltin acetate; triphenyltin hydroxide; fabric phase sorptive extraction (FPSE), high performance liquid chromatography (HPLC)

Online: 24 April 2018 (07:45:23 CEST)

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Preprint ARTICLE | doi:10.20944/preprints201804.0073.v1

Amino Acid Content in Onions as Potential Fingerprints of Geographical Origin: The Case of Rossa da Inverno sel. Rojo Duro

Federica Ianni, Antonella Lisanti, Maura Marinozzi, Emidio Camaioni, Lucia Pucciarini, Andrea Massoli, Roccaldo Sardella, Luciano Concezzi, Benedetto Natalini

Subject: Chemistry And Materials Science, Food Chemistry Keywords: Rossa da inverno sel. Rojo Duro onion cultivar; geographical origin; amino acids content; HPLC analysis; statistical evaluations; food traceability

Online: 6 April 2018 (09:04:47 CEST)

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Preprint ARTICLE | doi:10.20944/preprints202309.1546.v1

Real-Time Extension of TAO-DFT

Hung-Yi Tsai, Jeng-Da Chai

Subject: Physical Sciences, Chemical Physics Keywords: RT-TAO-DFT; TAO-DFT; multi-reference character; real-time electron dynamics; time-dependent properties; high-order harmonic generation

Online: 22 September 2023 (11:40:32 CEST)

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Preprint ARTICLE | doi:10.20944/preprints202208.0392.v1

Comparison of Four Real-time Polymerase Chain Reaction Assays for the Detection of SARS-CoV-2 on respiratory samples from Tunja, Boyacá - Colombia

Lorenzo Hernando Salamanca Neita, Óscar Leonardo Carvajal Rojas, Juan Pablo Carvajal Rojas, Ruth Maribel Forero, Niya Alexandra Segura

Subject: Biology And Life Sciences, Virology Keywords: Severe Acute Respiratory Syndrome-CoV-2 (SARS-CoV2), COVID-19, molecular diagnostics, real-time polymerase chain reaction (RT-qPCR)

Online: 23 August 2022 (03:55:44 CEST)

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Background: Coronavirus disease (COVID-19) is an infectious disease caused by the SARS-CoV-2. In Colombia, many commercial methods are now available to perform the RT-qPCR assays, and the laboratories must evaluate its diagnostic accuracy to ensure reliable results to suspected COVID-19 patients. The purpose of the study was to compare four commercial RT-qPCR assays for detection of SARS-CoV2 virus, from nasopharyngeal swab samples referred to Laboratorio Carvajal IPS, SAS of Tunja, Boyacá - Colombia. Methods: This prospective study was conducted on 152 samples of respiratory tract samples (Nasopharyngeal Swab) from patients with suspected SARS-CoV-2 infection. Diagnostic accuracy of GeneFinderTM COVID-19 Plus RealAmp (In Vitro diagnostic), One-Step Real-Time RT-PCR (Vitro Master diagnostica), Berlin modified protocol and gold standard Berlin protocol (Berlin Charite Probe One-Step RT-qPCR Kit, New England Biolabs) as reference was assessed. Operational characteristics were estimated in terms of sensitivity, specificity, agreement, and predictive values. Results: Using Berlin Charite Probe One-Step RT-qPCR Kit as reference, the sensitivity/specificity for the diagnostic tests were found to be GeneFinderTM COVID-19 Plus RealAmp Kit 100%/92.7%, One-Step Real-Time RT-PCR, One-Step Real-Time RT-PCR 92.75%/67.47%, and Berlin modified protocol 100%/96.39%. The results of four commercially available methods were found to be consistent with those obtained from Berlin Modified protocol analysis for % of the samples and showed good agreement (κ= 0.96). Concordant SARS-CoV2 negative and positive RT-qPCR results were reported for xxx and xxx samples, respectively. Summarize something about the Ct. Conclusion: Our data demonstrate that all commercially available methods are rapid and reliable for the identification of SARS-CoV-2 virus associated with COVID-19. One-Step RT-qPCR Kit and GeneFinderTM COVID-19 Plus RealAmp assay show optimal sensitivity compared with Belin modified protocol. In addition, there is no significant correlation between xxxxx

Preprint ARTICLE | doi:10.20944/preprints202309.1037.v1

Evaluation of Standard M10 Assay for Rapid Molecular Diagnosis of SARS-CoV-2, Influenza A/B virus and Respiratory Syncytial Virus (RSV)

Azwani Abdullah, I-Ching Sam, Yin Jie Ong, Chun Hao Theo, Muhammad Harith Pukhari, Yoke Fun Chan

Subject: Medicine And Pharmacology, Clinical Medicine Keywords: respiratory syncytial virus; influenza A virus; influenza B virus; SARS-CoV-2; rapid RT-PCR; diagnostic accuracy; point-of-care testing

Online: 15 September 2023 (05:38:13 CEST)

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Preprint ARTICLE | doi:10.20944/preprints202308.1465.v1

Pfizer-BioNTech mRNA Vaccine Protection among Children and Adolescents Aged 12–17 Years against COVID-19 Infection in Qatar

Khadieja Osman Hassan Elhag, Jesha Mohammedali Mundodan, Juel Bikhash Chowdhry, Rejoice Puthuchira Ravi, Rekaya houda Baaboura, Jeevan Albuquerque, Bilal Riaz, Reem Yusuf Emran, Khatija Batoul, Abdul Mahmood Esameldin, Zinah Al Tabatabaee, Hayat Salaheldin Khogali, Soha Shawqi Albayat

Subject: Public Health And Healthcare, Public Health And Health Services Keywords: Adolescent; children; 12-17 years COVID-19 vaccine; fully vaccinated; partially vaccinated; vaccination status; RT-PCR-positive; Pfizer; BNT162b2; vaccine effectiveness

Online: 22 August 2023 (03:02:19 CEST)

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Preprint ARTICLE | doi:10.20944/preprints201809.0054.v1

THD Reduction in Wind Energy System Using Type-4 Wind Turbine/PMSG Applying the Active Front-End Converter Parallel Operation

Nadia Maria Salgado-Herrera, David Campos-Gaona, Olimpo Anaya-Lara, Aurelio Medina-Rios, Roberto Tapia-Sanchez, Juan Ramon Rodriguez-Rodriguez

Subject: Engineering, Electrical And Electronic Engineering Keywords: Active Front-End converter; back-to-back converter; PMSG; THD; Type-4 wind turbine; wind energy system; Opal-RT Technologies®

Online: 4 September 2018 (05:02:15 CEST)

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Preprint ARTICLE | doi:10.20944/preprints202402.0098.v1

HPLC-qTOF-MS/MS Based Profiling Reveals Anthocyanin Profile Alterations in Berries of Hybrid Muscadine Variety FLH 13-11 in Two Continuous Cropping Seasons

Seyit YUZUAK, James Ballington, Gui Li, Deyu Xie

Subject: Biology And Life Sciences, Biology And Biotechnology Keywords: Anthocyanidins; anthocyanidin-3,5-O-diglucoside; anthocyanidin-3,5-O-monoglucoside; anthocyanins; HPLC-quadrupole time-of-flight tandem mass spectrometer; Vitis rotundifolia; metabolic profiling

Online: 2 February 2024 (09:18:46 CET)

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Preprint ARTICLE | doi:10.20944/preprints202310.0519.v1

Optimization of a protocol for launching grapevine infection with the biologically active cDNA clones of a virus

Mehdi Shabanian, Caihong Li, Ali Ebadi, Valerian Dolja, Baozhong Meng

Subject: Biology And Life Sciences, Virology Keywords: Grapevine leafroll disease; Closteroviridae; GLRaV-3cDNA clones; RT-qPCR; Western blot; vacuum agro-infiltration; agro-pricking; agro-drenching; agro-injection; Koch’s postulates

Online: 9 October 2023 (15:17:00 CEST)

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Grapevine leafroll disease (GLRD) is the most globally prevalent and destructive disease complex responsible for significant reductions in grape yield and quality as well as wine production. GLRD is associated with several positive-strand RNA viruses of the family Closteroviridae, designated grapevine leafroll-associated viruses (GLRaVs). However, the specific etiological role of any of these GLRaVs in GLRD has not been demonstrated. Even though GLRaV-3 is considered the chief GLRD agent, little is known about the molecular, cellular and pathological properties of this virus. Such knowledge gap is due to multiple factors, including unavailability of biologically active virus cDNA clones and the lack of reliable experimental systems for launching grapevine infection using such clones. In this work, we have tested four methods for inoculating tissue cultured grapevine plantlets with cDNA clones of GLRaV-3: i) vacuum agro-infiltration; ii) agro-pricking; iii) agro-drenching and iv) agro-injection. We have shown that vacuum agro-infiltration was the most effective among these methods. Further, we examined the impacts of different experimental conditions on the survival and infectivity rate of grapevines after infiltration. To verify the infectivity rate for different treatments, we used RT-PCR, RT-qPCR, and Western blotting. We have found that humidity plays a critical role in the survival of plantlets after agro-infiltration and that the use of RNA silencing suppressor, and dormancy treatment both had strong effects on the infection rates. To our best knowledge, the experimental protocol reported here is the most effective system for launching infection of grapevine using cDNA clones of grapevine viruses featuring up to 70% infection rate. This system has strong potential to facilitate grapevine virology research including fulfillment of Koch’s postulates for GLRD and other major virus diseases as well as the molecular, cellular, and pathological properties of GLRaVs and potentially other important grapevine viruses.

Preprint ARTICLE | doi:10.20944/preprints202109.0415.v2

A Routine Sanger Sequencing Target Specific Mutation Assay For SARS-CoV-2 Variants Of Concern And Interest

Sin Hang Lee

Subject: Biology And Life Sciences, Virology Keywords: Delta variant; Variants of concern; Variants of interest; SARS-CoV-2; Spike protein; Nested RT-PCR; Sanger sequencing; Amino acid mutations; ACE2 RBD; N-terminal domain (NTD)

Online: 2 November 2021 (10:40:46 CET)

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Preprint ARTICLE | doi:10.20944/preprints202103.0298.v1

Sintered Glass Monoliths as New Supports for Affinity Columns

Marco Wilke, Bettina Röder, Martin Paul, Michael G. Weller

Subject: Biology And Life Sciences, Biochemistry And Molecular Biology Keywords: Affinity chromatography; matrix; solid support; resin; support materials; glass filter; glass frit; high-pressure; HPLC; FPLC; antibodies; immunoglobulins; purification; downstream processing; protein purification; preparative; analytical; separation; clean-up; automation

Online: 11 March 2021 (08:37:01 CET)

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A novel stationary phase for affinity separations is presented. This material is based on sintered borosilicate glass readily available as semi-finished filter plates with defined porosity and surface area. The material shows fast binding kinetics and excellent long-term stability under real application conditions due to lacking macropores and high mechanical rigidity. The glass surface can be easily modified with standard organosilane chemistry to immobilize selective binders or other molecules used for biointeraction. In this paper, the manufacturing of the columns and their respective column holders by 3D printing is shown in detail. The model system protein A/IgG was chosen as an example to examine the properties of such monolithic columns under realistic application conditions. Several specifications, such as (dynamic) IgG capacity, pressure stability, long-term performance, productivity, non-specific binding, and peak shape, are presented. It could be shown that due to the very high separation speed, 250 mg antibody per hour and column can be collected, which surpasses the productivity of most standard columns of the same size. The total IgG capacity of the shown columns is around 4 mg (5.5 mg/mL), which is sufficient for most tasks in research laboratories. The cycle time of an IgG separation can be less than 1 minute. Due to the glass material's excellent pressure resistance, these columns are compatible with standard HPLC systems. This is usually not the case with standard affinity columns, limited to manual use or application in low-pressure systems. The use of a standard HPLC system also improves the ability for automation, which enables the purification of hundreds of cell supernatants in one day. The sharp peak shape of the elution leads to an enrichment effect, which might increase the concentration of IgG by a factor of 3. The final concentration of IgG can be around 7.5 mg/mL without the need for an additional nanofiltration step. The purity of the IgG was > 95% in one step and nearly 99% with a second polishing run.

Preprint ARTICLE | doi:10.20944/preprints202307.1155.v1

Blood Plasma Small Non-Coding RNAs as Diagnostic Molecules of Progesterone Receptor-Negative Phenotype of Serous Ovarian Tumors

Angelika V. Timofeeva, Ivan S. Fedorov, Alexandra V. Asaturova, Maya V. Sannikova, Anna V. Tregubova, Oleg A. Mayboroda, Grigory N. Khabas, Vladimir E. Frankevich, Gennady T. Sukhikh

Subject: Biology And Life Sciences, Biochemistry And Molecular Biology Keywords: miRNA; piRNA; mRNA; CA125; progesterone receptor (PGR); new-generation sequencing (NGS); quantitative RT-PCR; serous ovarian carcinoma; borderline cystadenoma; benign cystadenoma; formalin-fixed paraffin-embedded (FFPE) blocks; blood plasma; cytoreduction

Online: 18 July 2023 (09:58:12 CEST)

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Progesterone receptor (PGR) expression level determines biological characteristics of the serous ovarian carcinoma, and low PGR expression appears to be associated with chemoresistance and worse outcome. In this study, we aimed to find relationships between tumor progesterone receptor level and RNA-profile (miRNAs, piwiRNAs, and mRNAs), determining its biological characteristics and behavior. For this purpose, we applied next generation sequencing of small noncoding RNAs, quantitative RT-PCR, and immunohistochemistry to analyze FFPE and frozen tumor samples as well as blood plasma from patients with benign cystadenoma (BSC), serous borderline tumour (SBT), low-grade and high-grade serous ovarian carcinoma (LGSOC and HGSOC, respectively). We found significant upregulation of MMP7 and MUC16 and downregulation of PGR in LGSOC and HGSOC in comparison with BSC. The tissue content of miR-199a-5p, miR-214-3p, miR-424-3p, miR-424-5p, miR-125b-5p significantly inversely correlated with the expression level of MUC16 and blood serum CA125 concentration, and significantly directly correlated with the PGR expression level in tumor tissue. On the contrary, the tissue content of miR-16-5p, miR-17-5p, miR-20a-5p, miR-93-5p, responsible for epithelial-mesenchymal transition (EMT) of the cell, significantly directly correlated with the blood serum CA125 concentration and significantly inversely correlated with the PGR expression level in the tumor tissue. Levels of the EMT-associated miRNAs significantly directly correlated with the content of hsa_piR_022437, hsa_piR_009295, hsa_piR_020813, hsa_piR_004307, hsa_piR_019914 in tumor tissues. Among them, expression level of hsa_piR_004307 significantly inversly correlated with the PGR expression level in the tumor. Two optimal logistic regression models were developed based on the quantitation of hsa_piR_020813, miR-16-5p, hsa_piR_022437 or hsa_piR_004307, hsa_piR_019914, miR-93-5p in the tumor tissue, both of which significantly diagnose PGR-negative tumor phenotype with 93% sensitivity. According to FunRich3.1.3 functional enrichment analysis tool, 72 gene-targets of miRNA and piRNA identified here as markers of PGR-negative ovarian tumor phenotype were proven to be mutated in different cancers such as ovarian, breast, colorectal, liver, stomach, lung, endometrial, thyroid cancer. Among them, the blood plasma levels of miR-16-5p and hsa_piR_022437 can be used to diagnose PGR-negative tumor phenotype with 86% sensitivity before surgery and chemotherapy to choose the treatment strategy for this most aggressive type of ovarian cancer (for instance, neoadjuvant chemotherapy followed by cytoreduction in combination with hyperthermic intraperitoneal chemotherapy) to increase the effectiveness of treatment and longevity of the patient.

Preprint ARTICLE | doi:10.20944/preprints201911.0042.v1

In Spino-Cerebellar Tissue, Ataxin-2 Expansion Affects Ceramide-Sphingomyelin Metabolism

Nesli-Ece Sen, Aleksandar Arsovic, David Meierhofer, Susanne Brodesser, Carola Oberschmidt, Júlia Canet-Pons, Zeynep-Ece Kaya, Melanie V. Halbach, Suzana Gispert, Konrad Sandhoff, Georg Auburger

Subject: Biology And Life Sciences, Endocrinology And Metabolism Keywords: olivo-ponto-cerebellar atrophy (OPCA); amyotrophic lateral sclerosis (ALS); tauopathy; leukodystrophy; mass spectrometry; RT-qPCR; Ceramide Synthase (CERS2/CERS1); Serine Palmitoyltransferase 2 (Sptlc2); neutral Sphingomyelinase (Smpd3); neutral Ceramidase (Asah2); Fatty Acid Elongase (Elovl1/4/5); SCA34; SCA38; acid Sphingomyelinase (ASMase, Smpd1)

Online: 5 November 2019 (03:04:02 CET)

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Ataxin-2 (ATXN2) acts during stress-responses, modulating mRNA translation and nutrient metabolism. Atxn2 knockout mice exhibit progressive obesity, dyslipidemia and insulin resistance. Conversely, the progressive ATXN2 gain-of-function due to polyGlutamine (polyQ) expansions leads to a dominantly inherited neurodegenerative process named spinocerebellar ataxia type 2 (SCA2), with early adipose tissue loss and late muscle atrophy. We tried to understand lipid dysregulation in a SCA2 patient brain and in an authentic mouse model. Thin layer chromatography of a patient cerebellum was compared to the lipid metabolome of Atxn2-CAG100-KnockIn (KIN) mouse spinocerebellar tissue. The human pathology caused deficits of sulfatide, galactosylceramide, cholesterol, C22/24-sphingomyelin and gangliosides GM1a/GD1b, despite quite normal levels of C18-sphingomyelin. Cerebellum and spinal cord from the KIN mouse showed a consistent decrease of various ceramides, with a significant elevation of sphingosine in the more severely affected spinal cord. Deficiency of C24/26-sphingomyelins contrasted with excess C18/20-sphingomyelin. Spinocerebellar expression profiling revealed consistent reductions of CERS protein isoforms, of Sptlc2 and Smpd3, but upregulation of Cers2 mRNA, as prominent anomalies in the ceramide-sphingosine metabolism. Reduction of Asah2 mRNA correlated to deficient S1P levels. In addition, downregulations for the elongase Elovl1, Elovl4, Elovl5 mRNAs and ELOVL4 protein explain the deficit of very-long-chain sphingomyelin. Reduced ASMase protein levels correlated to the accumulation of long-chain sphingomyelin. Overall, a deficit of myelin lipids was prominent in SCA2 nervous tissue at prefinal stage, not compensated by transcriptional adaptation of several metabolic enzymes. Myelination is controlled by mTORC1 signals, so our human and murine observations are in agreement with the known role of ATXN2 yeast, nematode and mouse orthologs as mTORC1 inhibitors and autophagy promoters.

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