High natural PHA production from acetate in Cobetia sp. MC34 and Cobetia marina DSM 4741T and in silico analyses of the genus specific PhaC2 polymerase variant

Background Several members of the bacterial Halomonadacea family are natural producers of polyhydroxyalkanoates (PHA), which are promising materials for use as biodegradable bioplastics. Type-strain species of Cobetia are designated PHA positive, and recent studies have demonstrated relatively high PHA production for a few strains within this genus. Industrially relevant PHA producers may therefore be present among uncharacterized or less explored members. In this study, we characterized PHA production in two marine Cobetia strains. We further analyzed their genomes to elucidate pha genes and metabolic pathways which may facilitate future optimization of PHA production in these strains. Results Cobetia sp. MC34 and Cobetia marina DSM 4741T were mesophilic, halotolerant, and produced PHA from four pure substrates. Sodium acetate with- and without co-supplementation of sodium valerate resulted in high PHA production titers, with production of up to 2.5 g poly(3-hydroxybutyrate) (PHB)/L and 2.1 g poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV)/L in Cobetia sp. MC34, while C. marina DSM 4741T produced 2.4 g PHB/L and 3.7 g PHBV/L. Cobetia marina DSM 4741T also showed production of 2.5 g PHB/L from glycerol. The genome of Cobetia sp. MC34 was sequenced and phylogenetic analyses revealed closest relationship to Cobetia amphilecti. PHA biosynthesis genes were located at separate loci similar to the arrangement in other Halomonadacea. Further genome analyses revealed some differences in acetate- and propanoate metabolism genes between the two strains. Interestingly, only a single PHA polymerase gene (phaC2) was found in Cobetia sp. MC34, in contrast to two copies (phaC1 and phaC2) in C. marina DSM 4741T. In silico analyses based on phaC genes show that the PhaC2 variant is conserved in Cobetia and contains an extended C-terminus with a high isoelectric point and putative DNA-binding domains. Conclusions Cobetia sp. MC34 and C. marina DSM 4741T are natural producers of PHB and PHBV from industrially relevant pure substrates including acetate. However, further scale up, optimization of growth conditions, or use of metabolic engineering is required to obtain industrially relevant PHA production titers. The putative role of the Cobetia PhaC2 variant in DNA-binding and the potential implications remains to be addressed by in vitro- or in vivo methods.

Biofilm Formation by Marine Cobetia marina alex and Pseudoalteromonas spp: Development and Detection of Quorum Sensing N-Acyl Homoserine Lactones (AHLs) Molecules

Surfaces submerged in seawater are colonized by various microorganisms, resulting in the formation of heterogenic marine biofilms. This work aims to evaluate the biofilm formation by Cobetia marina alex and doing a comparative study between this promising strain with the two bacterial strains isolated previously from the Mediterranean seawater, Alexandria, Egypt. Three strains; Cobetia marina alex, Pseudoalteromonas sp. alex, and Pseudoalteromonas prydzensis alex were screened for biofilm formation using the crystal violet (CV) quantification method in a single culture. The values of biofilm formed were OD600= 3.0, 2.7, and 2.6, respectively leading to their selection for further evaluation. However, factors affecting biofilm formation by C. marina alex were investigated. Biofilm formation was evaluated in single and multispecies consortia. Synergistic and antagonistic interactions proved in this work lead to the belief that these bacteria have the capability to produce some interesting signal molecules N-acyl Homoserine Lactones (AHLs)

Viral degradation of marine bacterial exopolysaccharides

The identification of the mechanisms by which marine dissolved organic matter (DOM) is produced and regenerated is critical to develop robust prediction of ocean carbon cycling. Polysaccharides represent one of the main constituents of marine DOM and their degradation is mainly attributed to polysaccharidases derived from bacteria. Here, we report that marine viruses can depolymerize the exopolysaccharides (EPS) excreted by their hosts using five bacteriophages that infect the notable EPS producer, Cobetia marina DSMZ 4741. Degradation monitorings as assessed by gel electrophoresis and size exclusion chromatography showed that four out of five phages carry structural enzymes that depolymerize purified solution of Cobetia marina EPS. The depolymerization patterns suggest that these putative polysaccharidases are constitutive, endo-acting and functionally diverse. Viral adsorption kinetics indicate that the presence of these enzymes provides a significant advantage for phages to adsorb onto their hosts upon intense EPS production conditions. The experimental demonstration that marine phages can display polysaccharidases active on bacterial EPS lead us to question whether viruses could also contribute to the degradation of marine DOM and modify its bioavailability. Considering the prominence of phages in the ocean, such studies may unveil an important microbial process that affects the marine carbon cycle.

Antimicrobial Properties of Transition Metals Substituted Chitosan against Cobetia Marina

This research work offers the new organic compound which has potential to replace the use of tributylin (TBT) in antifouling application. All modified chitosan powders were synthesized using in-house powder processing techniques. Pure and modified chitosan were evaluated for fouling properties via antimicrobial activity test against Cobetia Marina (C. Marina). The results indicated that copper substituted chitosan (CS-Cu) showed the highest inhibition zone and lowest number of C. Marina survival. It can be implied that CS-Cu showed the best antimicrobial ability against C. Marina.