Synechococcus sp. PCC 11901 reportedly demonstrates the highest, sustained growth of any known cyanobacterium under optimized conditions. Due to its recent discovery our knowledge of its biology, including the factors underlying sustained, fast growth, is limited. Furthermore, tools specific for genetic manipulation of PCC 11901 are not established. Here, we demonstrate that PCC 11901 shows faster growth than other model cyanobacteria, including the fast-growing species Synechococcus elongatus UTEX 2973, under optimal growth conditions for UTEX 2973. Comparative genomics between PCC 11901 and Synechocystis sp. PCC 6803 reveal conservation of most metabolic pathways but PCC 11901 has a simplified electron transport chain and reduced light-harvesting complex. This may underlie its superior light utilization, reduced photoinhibition and higher photosynthetic and respiratory rates. To aid biotechnology applications we developed a vitamin B12 auxotrophic mutant but were unable to generate unmarked knockouts using two negative selectable markers, suggesting that recombinase- or CRISPR-based approaches may be required for repeated genetic manipulation. Overall, this study establishes PCC 11901 as one of the most promising species currently available for cyanobacterial biotechnology and provides a useful set of bioinformatics tools and strains for advancing this field, in addition to insights into the factors underlying its fast growth phenotype.

Abstract: Given the increasing resistance of some bacterial strains to many common antibiotics used in the local market, the current study aimed to produce nanoscale particles capable of combating pathogenic bacteria that have developed resistance to current antibiotics. In this research, an ex-perimental study was conducted to produce nanoscale particles using the single-celled blue-green algae Synechococcus sp., which was isolated from a local environment in the Arab Republic of Egypt, specifically from a soil sample in Beni-Suef City. The aim of the study was to generate silver NPs employing different biological approaches, encompassing the direct strain powder technique, ethanol extract pellet technique, and ethanol extract technique. These synthesized nanoparticles displayed different shapes, including cubic, spherical, pellet-like, and rod-shaped particles. They were characterized by bio-functional groups containing Ag+ ions, which were detected using FT-IR, Zeta sizer, and Zeta potential measurements. High-resolution SEM imaging unveiled the mor-phological features of Silver NPs with biologically formed protective layers, spanning sizes from 20 to 430 nm for individual particles. The study evaluated the antibacterial efficiency against both G+ve and G-ve bacteria. The ethanolic extract technique exhibited the largest inhibition zone (24 mm) against G+ve bacteria, particularly Staphy. aureus. Furthermore, it was observed that the larger particles produced using the powder and pellet techniques were less effective antibacterial agents. In conclusion, this investigation emphasizes the potential of cyanobacteria-based synthesis of silver nanoparticles as a promising approach for producing innovative antimicrobial agents.

There is no doubt that seagrass beds constitute one of the most productive ecosystems in shallow coastal waters. Despite this, picoplankton in seagrass ecosystems has received relatively little attention. The purpose of this study was to compare picoplankton growth and mortality rates between seagrass and unvegetated habitats using chamber incubations. We tested two main hypotheses: (i) incubation with seagrass would result in higher bacterial growth rates due to increased DOM release from seagrass photosynthesis, and (ii) Synechococcus spp. would be lower in the presence of seagrass due to competition for inorganic nutrients. Our results showed bacterial growth rates of 2.31 and 2.44 d–1 without and with seagrass, respectively. In seagrass chambers, bacterial growth was higher than that in non-seagrass chambers, suggesting that organic carbon coming from the seagrass community may support bacterial production. Furthermore, we observed a negative Synechococcus spp. growth rate (-0.9 d–1) in the seagrass treatment, likely reflecting nutrient competition with the seagrass. Small-scale chambers proved to be a useful tool for studying the factors controlling spatial and temporal patterns of picoplankton across different habitats. Furthermore, future studies should examine picoplankton growth over a wider range of spatial scales in seagrass beds and adjacent unvegetated sediment.

Yeast two-hybrid approaches, which are based on fusion proteins that must co-localise to the nucleus to reconstitute the transcriptional activity of GAL4 have greatly contributed to understand the nitrogen interaction network of cyanobacteria, whose main hubs are the trimeric PII and the monomeric PipX regulators. The bacterial two-hybrid system, based on the reconstitution in the E. coli cytoplasm of the adenylate cyclase of Bordetella pertussis, should provide a relatively faster and presumably more physiological assay for cyanobacterial proteins than the yeast system. Here we use the bacterial two-hybrid system to gain additional insights into the cyanobacterial PipX interaction network while simultaneously assessing the advantages and limitations of the two most popular two-hybrid systems. Comprehensive mutational analysis of PipX and bacterial two-hybrid assays were performed to compare outcomes between yeast and bacterial systems. We detected interactions previously recorded in the yeast two-hybrid system as negative as well as a “false positive”, the self-interaction of PipX, which is rather an indirect interaction dependent on PII homologs from the E. coli host, a result confirmed by Western analysis with relevant PipX variants. This is, to our knowledge, the first report of the molecular basis of a false positive in the bacterial two-hybrid system.

Cyanobacteria, microorganisms performing oxygenic photosynthesis, must adapt their metabolic processes to environmental challenges such as day and night changes. PipX, a unique regulatory protein from cyanobacteria provides a mechanistic link between signalling protein PII, a widely conserved (in bacteria and plants) transducer of carbon/nitrogen/energy richness, and the transcriptional regulator NtcA, which controls a large regulon involved in nitrogen assimilation. PipX is also involved in translational regulation by interaction with the ribosome-assembly GTPase EngA. However, increases of the PipX/PII ratio are toxic, presumably by abnormally increased binding of PipX to other partner(s). Here we present mutational and structural analyses of reported PipX-PII and PipX-NtcA complexes, leading to identification of single amino acid changes that decrease or abolish PipX toxicity. Four out of 11 mutations decreasing toxicity did not decrease PipX levels, suggesting that the targeted residues (F12, D23, L36, R54) provide toxicity determinants. In addition, one of those four mutations (D23A) argued against the over-activation of NtcA as the cause of PipX toxicity. Since most mutations at residues contacting PII decreased PipX levels, PipX stability would depend on its ability to bind to PII, a conclusion supported by the light-induced decrease in PipX levels observed in Synechococcus elongatus PCC7942.

Terpenes are high-value chemicals which can be produced by engineered cyanobacteria from sustainable resources, solar energy, water and CO2. We previously reported that the euryhaline unicellular cyanobacteria Synechocystis PCC 6803 (S.6803) and Synechococcus PCC 7002 (S.7002) produce farnesene and limonene, respectively, more efficiently than other terpenes. In the present study, we attempted at enhancing farnesene production in S.6803 and limonene production in S.7002. Practically, we tested the influence of key cyanobacterial enzymes acting in carbon fixation (RubisCO, PRK, CcmK3 and CcmK4), utilization (CrtE, CrtR and CruF) and storage (PhaA and PhaB) on terpene production in S.6803, and we compared some of the findings with the data obtained in S.7002. We report that the overproduction of RubisCO from S.7002 and PRK from Cyanothece PCC 7425 increased farnesene production in S.6803, but not limonene production in S.7002. The overexpression of the crtE genes (synthesis or terpene precursors) from S.6803 or S.7002 did not increased farnesene production in S.6803. In contrast, the overexpression of the crtE gene from S.6803, but not S.7002, increased farnesene production in S.7002, emphasizing the physiological difference between these two model cyanobacteria. Furthermore, the deletion of the crtR and cruF genes (carotenoid synthesis) and phaAB genes (carbon storage) did not increase the production of farnesene in S.6803. Finally, as a containment strategy of genetically modified (GMO) strains of S.6803 we report that the deletion of the ccmK3K4 genes (carboxysome for CO2 fixation) did not affect the production of limonene but decreased the production of farnesene in S.6803.