RESUMEN
The genus Clostridium is a large and diverse group of species that can cause food spoilage, including late blowing defect (LBD) in cheese. In this study, we investigated the taxonomic status of strain FAM25158 isolated from Emmental cheese with LBD using a polyphasic taxonomic and comparative genomic approach. A 16S rRNA gene sequence phylogeny suggested affiliation to the Clostridium sensu stricto cluster, with Clostridium tyrobutyricum DSM 2637T being the closest related type strain (99.16% sequence similarity). Average Nucleotide Identity (ANI) analysis revealed that strain FAM25158 is at the species threshold with C. tyrobutyricum, with ANI values ranging from 94.70 to 95.26%, while the digital DNA-DNA hybridization values were below the recommended threshold, suggesting that FAM25158 is significantly different from C. tyrobutyricum at the genomic level. Moreover, comparative genomic analysis between FAM25158 and its four closest C. tyrobutyricum relatives revealed a diversity of metabolic pathways, with FAM25158 differing from other C. tyrobutyricum strains by the presence of genes such as scrA, srcB, and scrK, responsible for sucrose utilization, and the absence of many important functional genes associated with cold and osmolality adaptation, which was further supported by phenotypic analyses. Surprisingly, strain FAM25158 exhibited unique physiologic traits, such as an optimal growth temperature of 30°C, in contrast to its closest relatives, C. tyrobutyricum species with an optimal growth temperature of 37°C. Additionally, the growth of FAM25158 was inhibited at NaCl concentrations higher than 0.5%, a remarkable observation considering its origin from cheese. While the results of this study provide novel information on the genetic content of strain FAM25158, the relationship between its genetic content and the observed phenotype remains a topic requiring further investigation.
RESUMEN
Ropy defect of pasteurized fluid milk is a type of spoilage which manifests itself by an increased viscosity, slimy body, and string-like flow during pouring. This defect has, among other causes, been attributed to the growth, proliferation and exopolysaccharide production by coliform bacteria, which are most commonly introduced in milk as post-pasteurization contaminants. As we identified both Klebsiella pneumoniae ssp. pneumoniae and Rahnella inusitata that were linked to a ropy defect, the goal of this study was to characterize 3 K. pneumoniae ssp. pneumoniae strains and 2 R. inusitata for (1) their ability to grow and cause ropy defect in milk at 6°C and 21°C and to (2) probe the genetic basis for observed ropy phenotype. Although all K. pneumoniae ssp. pneumoniae and R. inusitata strains showed net growth of >4 log10 over 48 h in UHT milk at 21°C, only R. inusitata strains displayed growth during 28-d incubation period at 6°C (>6 log10). Two out of 3 K. pneumoniae ssp. pneumoniae strains were capable of causing the ropy defect in milk at 21°C, as supported by an increase in the viscosity of milk and string-like flow during pouring; these 2 strains were originally isolated from raw milk. Only one R. inusitata strains was able to cause the ropy defect in milk; this strain was able to cause the defect at both 6°C and 21°C, and was originally isolated from a pasteurized milk. These findings suggest that the potential of K. pneumoniae ssp. pneumoniae and R. inusitata to cause ropy defect in milk is a strain-dependent characteristic. Comparative genomics provided no definitive answer on genetic basis for the ropy phenotype. However, for K. pneumoniae ssp. pneumoniae, genes rffG, rffH, rfbD, and rfbC involved in biosynthesis and secretion of enterobacterial common antigen (ECA) could only be found in the 2 strains that produced ropy defect, and for R. inusitata a set of 2 glycosyltransferase- and flippase genes involved in nucleotide sugar biosynthesis and export could only be identified in the ropy strain. Although these results provide some initial information for potential markers for strains that can cause ropy milk, the relationship between genetic content and ropiness in milk remains poorly understood and merits further investigation.