Rapid detection and identification of nontuberculous mycobacterial pathogens in fish by using high-resolution melting analysis

Mycobacterial infections in fish are commonly referred to as piscine mycobacteriosis, irrespectively of the specific identity of the causal organism. They usually cause a chronic disease and sometimes may result in high mortalities and severe economic losses. Nearly 20 species of Mycobacterium have been reported to infect fish. Among them, Mycobacterium marinum, M. fortuitum, and M. chelonae are generally considered the major agents responsible for fish mycobacteriosis. As no quick and inexpensive diagnostic test exists, we tested the potential of high-resolution melting analysis (HRMA) to rapidly identify and differentiate several Mycobacterium species involved in fish infections. By analyzing both the melting temperature and melting profile of the 16S-23S rRNA internal transcribed spacer (ITS), we were able to discriminate 12 different species simultaneously. Sensitivity tests conducted on purified M. marinum and M. fortuitum DNA revealed a limit of detection of 10 genome equivalents per reaction. The primers used in this procedure did not lead to any amplification signal with 16 control non-Mycobacterium species, thereby demonstrating their specificity for the genus Mycobacterium

Biogenic volatile organic compounds as a potential stimulator for organic contaminant degradation by soil microorganisms

The effects of monoterpenes on the degradation of 14C-2,4-dichlorophenol (DCP) were investigated in soils collected from areas surrounding monoterpene and non-monoterpene emitting vegetation. Indigenous microorganisms degraded 14C-2,4-DCP to 14CO2, after 1 d contact time. Degradation was enhanced by prior exposure of the soils to 2,4-DCP for 32 d, increasing mineralization extents up to 60%. Monoterpene amendments further enhanced 2,4-DCP degradation, but only following pre-exposure to both 2,4-DCP and monoterpene, with total 2,4-DCP mineralisation extents of up to 71%. Degradation was greatest at the higher monoterpene concentrations (≥ 1 μg kg-1). Total mineralisation extents were similar between concentrations, but higher than the control and the 0.1 μg kg-1 amendment, indicating that increases in monoterpene concentration has a diminishing enhancing effect. We suggest that monoterpenes can stimulate the biodegradation of 2,4-DCP by indigenous soil microorganisms and that monoterpene amendment in soils is an effective strategy for removing organic contaminants

Evaluation of denaturing gradient gel electrophoresis in the detection of 16S rDNA sequence variation in rhizobia and methanotrophs

The ability of denaturing gradient gel electrophoresis (DGGE) technique to resolve 16S rDNA products generated from two different collections of bacteria using universal 16S primers was investigated. Alignments of 16S rDNA sequences of known species of rhizobia and methanotrophs were performed in order to determine the genetic variations within a 200 bp product obtained with PCR primers which amplify the 16S rRNA encoding genes from Eubacteria. Theoretical DNA melting curves were obtained with the Melt87 program and found to correlate with the ability to resolve fragments by DGGE. In the case of the rhizobia, the inability of DGGE analysis to resolve the PCR products from closely related species was in accordance with the low polymorphism observed amongst the sequences in the amplified area. In the case of the methanotrophs, the PCR products were surprisingly difficult to resolve given the high degree of sequence polymorphism of the amplified area in some distantly related species. The difference in sequence divergence within the two groups members allowed therefore to scale the resolution ability of the DGGE technique

Microbial communities in natural rubber coagula during maturation: Impacts on technological properties of dry natural rubber

Aim: To characterize microbial communities present in natural rubber (NR) coagula from Hevea brasiliensis latex during maturation and identify microbial taxa (bacteria and fungi) having an impact on dry NR properties. Methods and Results: Microbial community dynamics in NR coagula maturated under controlled conditions were compared and related with the evolution of dry NR properties. The pyrosequencing of 16S (119 837 effective reads) and 18S (131 879 effective reads) rRNA gene regions was performed on 21 samples covering different maturation times and two aeration conditions. Results showed a relatively high bacterial richness (Chao1 estimates of 200–1000) associated with significant bacterial dynamics. Lactic acid bacteria (LAB) were dominant in the first days of maturation. Then, in aerobic conditions, development of Actinobacteria represented by the family Microbacteriaceae was associated with alkalinization of the samples and a higher sensitivity of NR to thermo-oxidation as evaluated by its plasticity retention index (PRI). In anaerobiosis, the reduced development of bacteria, mostly LAB present, was associated with improved NR properties (higher initial plasticity P0 and PRI). Conclusions: The involvement of micro-organisms in the evolution of dry NR properties during the maturation of NR coagula was confirmed. The importance of the structure and dynamics of microbial communities is specifically highlighted. Significance and Impact of the Study: Natural rubber is a key elastomer for the tyre industry and for a variety of other applications. The majority of raw NR is obtained by natural coagulation of H. brasiliensis latex under the activity of micro-organisms. An improved understanding of the microbial communities involved in the maturation of NR coagula may lead to an improvement in the production process of raw NR to provide a better consistency in NR quality. (Résumé d'auteur

Genomic diversity and evolution within the species Streptococcus agalactiae

Streptococcus agalactiae is a leading cause of invasive infections in neonates, and responsible for bovine mastitis. It is also a commensal bacterium adapted to asymptomatic colonization of the mammalian gut and of the genitourinary tract. Here, we report the analysis of a collection of 75 strains of human and animal origin by using serotyping, multilocus sequence typing, whole genome DNA-array hybridizations and sequence comparison of putatively virulence-associated loci. Although the most variable parts of the genome are the previously predicted genomic islands, significant genetic variations were present in the genome backbone. Evolution within genes encoding surface and secreted proteins and those involved in the biosynthesis of different capsular types is mainly due to recombination events leading to the replacement of a locus of several genes or to the allelic exchange of the internal part of a gene. These two processes, which led to a broad diversity of surface protein patterns, are probably involved in the diversity of interactions with the host and its immune system. According to gene content comparisons and phylogeny, recent gene replacements by horizontal gene transfer may occur but are rare events. Although specific gene patterns, with respect to the origin of the strains and the epidemiological characteristics, were not identified, we show that the recently described hypervirulent ST-17 lineage is a homogeneous group. The study highlights for the first time that this lineage contains a specific and conserved set of surface proteins, probably accounting for its high capacity to cause infections in newborns

A European epidemiological survey of Vibrio splendidus clade shows unexplored diversity and massive exchange of virulence factors

The Vibrio splendidus clade has previously been associated with epidemic outbreaks of various aquatic animals, as in the case of the cupped oyster, Crassostrea gigas. To investigate whether involved strains could present a clonal origin and to identify possible alternative background carriage animals or zooplankton, a large epidemiological survey was conducted on isolates of the splendidus clade. For this purpose, Vibrio strains were isolated from various samples including oysters, mussels, sediments, zooplankton, and sea water on the basis of a North/South gradient of the European sea water zone (Ireland, The Netherlands, France, Italy, and Spain). A total of 435 isolates were successfully associated to the V. splendidus clade using real time polymerase chain reaction with 16S specific primers and probes. A multiple-locus variable-number tandem-repeat analysis (VNTR) was conducted on all isolates based on a multiplex PCR-VNTR with a set of primer pairs designed from the V. tasmaniensis LGP32 genome. Preliminary validation of the primers on a set of collection strains from the V. splendidus clade confirmed that the former V. splendidus-related LGP32 and relative strains were related to V. tasmaniensis rather than to the type strain V. splendidus LMG 4042. The VNTR analysis was then successfully conducted on 335 isolates which led to the characterization of 87 different profiles. Our results showed that (1) the high diversity of VNTR did not enlighten significant correlation between a specific pattern and the origin of collected samples. However, populations isolated from animal samples tend to differ from those of the background environment; (2) oyster mortality events could not be linked to the clonal proliferation of a particular VNTR type. However, few different patterns seemed successively associated with samples collected during peaks of oyster's mortality. (3) Finally, no correlation could be seen between specific VNTR patterns and sequence phylogeny of the virulence factors vsm and ompU that were detected among strains isolated during as well as outside mortality events. These results, combined with incongruence observed between the ompU and vsm phylogenetic trees, suggested both large diffusion of strains and massive lateral gene transfer within the V. splendidus clade

Genome Sequence of the Edible Cyanobacterium Arthrospira sp. PCC 8005▿

We determined the genome sequence of Arthrospira sp. PCC 8005, a cyanobacterial strain of great interest to the European Space Agency for its nutritive value and oxygenic properties in the Micro-Ecological Life Support System Alternative (MELiSSA) biological life support system for long-term manned missions into space