Entrapment of anaerobic thermophilic and hyperthermophilic marine micro-organisms in a gellan/xanthan matrix.

AIMS: The aims of this study were (i) to develop a protocol for the entrapment of anaerobic (hyper)thermophilic marine micro-organisms; (ii) to test the use of the chosen polymers in a range of physical and chemical conditions and (iii) to validate the method with batch cultures. METHODS AND RESULTS: The best conditions for immobilization were obtained at 80°C with gellan and xanthan gums. After 5-week incubation, beads showed a good resistance to all tested conditions except those simultaneously including high temperature (100°C), low NaCl (<0∙5 mol l(-1) ) and extreme pH (4/8). To confirm the method efficiency, batch cultures with immobilized Thermosipho sp. strain AT1272 and Thermococcus kodakarensis strain KOD1 showed an absence of detrimental effect on cell viability and a good growth within and outside the beads. CONCLUSION: This suggests that entrapment in a gellan-xanthan matrix could be employed for the culture of anaerobic (hyper)thermophilic marine micro-organisms. SIGNIFICANCE AND IMPACT OF THE STUDY: (Hyper)thermophilic marine micro-organisms possess a high biotechnological potential. Generally microbial cells are grown as free-cell cultures. The use of immobilized cells may offer several advantages such as protection against phage attack, high cell biomass and better production rate of desired metabolites.

Cultivation of an immobilized (hyper)thermophilic marine microbial community in a bioreactor.

Cultivation in a bioreactor of immobilized deep-sea hydrothermal microbial community was tested in order to assess the stability and reactivity of this new system. A community composed of eight hydrothermal strains was entrapped in a polymer matrix that was used to inoculate a continuous culture in a gas-lift bioreactor. The continuous culture was performed for 41 days at successively 60°C, 55°C, 60°C, 85°C and 60°C, at pH 6.5, in anaerobic condition and constant dilution rate. Oxic stress and pH variations were tested at the beginning of the incubation. Despite these detrimental conditions, three strains including two strict anaerobes were maintained in the bioreactor. High cell concentrations (3 × 10(8) cells mL(-1)) and high ATP contents were measured in both liquid fractions and beads. Cloning-sequencing and qPCR revealed that Bacillus sp. dominated at the early stage, and was later replaced by Thermotoga maritima and Thermococcus sp. Acetate, formate and propionate concentrations varied simultaneously in the liquid fractions. These results demonstrate that these immobilized cells were reactive to culture conditions. They were protected inside the beads during the stress period and released in the liquid fraction when conditions were more favorable. This confirms the advantage of immobilization that highlights the resilience capacity of certain hydrothermal microorganisms after a stress period.

Diversity, dynamics and activity of bacterial populations in 'Registered Designation of Origin' Salers cheese by single-strand conformation polymorphism analysis of 16S rRNA genes.

AIMS: The aim of this work was to measure the dynamic global metabolic activities of the microbial community during ripening of RDO Salers cheese by using a direct molecular approach. METHODS AND RESULTS: A culture-independent approach including PCR, reverse transcriptase PCR (RT-PCR) and single strand conformation polymorphism (SSCP) analysis of 16S rRNA genes was applied on 'Registered Designation of Origin' Salers cheese samples collected in three farms. The evolution of the global structure of the microbial community in terms of structure or global activities was assessed using ecological indices. The diversity of the global population was higher on RNA patterns than on DNA patterns, because of less dominance and greater richness. Comparison of the SSCP patterns derived from RNA and DNA analysis indicated that the dominant population was not necessarily the most active. The metabolic activities of each bacterial group changed significantly during ripening. Besides lactic acid bacteria that were dominant on both DNA and RNA patterns, the dynamics of the presence and activity of microbial groups rarely studied in the core of cheese, such as corynebacteria, or of unidentified peaks were reported. CONCLUSIONS: By using SSCP RNA analysis, we were able to obtain information about the activity of bacterial population in cheese, which varied a lot between cheeses and was changing perpetually during ripening. SIGNIFICANCE AND IMPACT OF THE STUDY: Double DNA-RNA SSCP analysis opens up future prospects in the microbial ecology of cheeses. It will have many applications for controlling of microbial community during cheese processing.

Competence regulation by oxygen availability and by Nox is not related to specific adjustment of central metabolism in Streptococcus pneumoniae.

In Streptococcus pneumoniae oxygen availability is a major determinant for competence development in exponentially growing cultures. NADH oxidase activity is required for optimal competence in cultures grown aerobically. The implication of oxidative metabolism and more specifically of Nox on central metabolism has been examined. Glycolytic flux throughout exponential growth revealed homolactic fermentation with a lactate production/glucose utilization ratio close to 2, whatever the aerobiosis level of the culture. Loss-of-function mutations in nox, which encodes NADH oxidase, did not change this trait. Consistently, mRNA levels of glyceraldehyde-3-phosphate dehydrogenase, L-lactate dehydrogenase, pyruvate oxidase, and NADH oxidase remained comparable to wild-type levels, as did the specific activities of key enzymes which control central metabolism. Competence regulation by oxygen involving the NADH oxidase activity is not due to significant modification of carbon flux through glycolysis. Failure to obtain loss-of-function mutation in L-ldh, which encodes the L-lactate dehydrogenase, indicates its essential role in pneumococci whatever their growth status.