Characterization of Shewanella baltica strains with usual and atypical H2S productions, isolated from a spoiled whiting (Merlangius merlangus)
Résumé
Introduction:
Shewanella putrefaciens has been described in numerous studies as the specific spoilage organism of marine fish conserved on melting ice. S. putrefaciens is more likely a bacterial group divided into four sub-groups than a single species; subgroup II being only composed by Shewanella baltica. In most cases, isolation of Shewanella strains involves the ability to produce hydrogen sulfur while growing on iron agar medium. Besides, among Shewanella, S. baltica has been described as the most H2S producing bacteria during the storage of marine fish.
Material and methods:
Four strains of Shewanellaceae were isolated from a spoiled whiting (Merlangius merlangus) using a phenotypic criterion (purple pigment production on TSA-YE medium). All the strains were subjected to growth ability, biochemical, molecular and proteomic analyses. Growth tests were performed in aerobic condition on TSA-YE NaCl medium from 4°C to 37°C. A test was also carried out anaerobically at 25°C on TSA-YE NaCl medium with 40 mM trimethylamine-N-oxide (TMAO). Biochemical patterns were observed on API 20E system incubated at 30 °C for 48H. Molecular analysis involved partial sequencing of both 16S rRNA and gyrB genes. MALDI-TOF analyses were carried out on proteins extracts with a Brüker LT Microflex device.
Results and discussion:
All strains were able to grow aerobically from 4°C to 30°C. Interestingly, they were also able to grow in anaerobic condition at 25°C with TMAO, the characteristic odor of trimethylamine (fishy off-odor) being ascertained. Biochemical characterization did not allow accurately identifying the four strains. This study allows emphasizing that a strain was unable to produce hydrogen sulfur. Two isolates synthetized an ornithine decarboxylase, making them potential producers of putrescine. Concerning the use of carbon source, different patterns were recorded among the strains. Genotypic characterization allowed identifying the strains thanks to both 16Sr RNA and gyrB sequencing. The four isolates were identified as member of Shewanella baltica species. Comparing the two sequencing approaches, gyrB sequences allowed a better discrimination of the strains. Proteomic characterization of the strains confirmed that the isolates belong to S. baltica species. MALDI-TOF study allows pointing out some slight differences between the proteome of the strains.
Conclusion:
This study allows isolating a H2S negative S. baltica from a spoiled fish. New studies will be necessary to estimate the prevalence of such a strain and its spoilage abilities. Thus H2S production might not be a suitable screening parameter both to count Shewanella or to study fish spoilage flora.