RESUMO
Vibrio bacteria, and particularly members of the Harveyi clade, are the causative agents of vibriosis. This disease is responsible for mass mortality events and important economic losses on aquaculture farms. Improvements in surveillance and diagnosis are needed to successfully manage vibriosis outbreaks. 16S rRNA gene sequencing is generally considered to be the gold standard for bacterial identification but the cost and long processing time make it difficult to apply for routine identification. In contrast, MALDI-TOF MS offers rapid diagnosis and is commonly used in veterinary laboratories today. The major limiting factor for using this technique is the low environmental bacterial diversity in the commonly available databases. Here, we demonstrate that the sole use of the commercially available Bruker BioTyper database is not fully adequate for identifying Vibrio bacteria isolated from aquaculture farms. We therefore developed a new in-house database named Luvibase, composed of 23 reference MALDI-TOF mass spectra profiles obtained from Vibrio collection strains, mostly belonging to the Harveyi clade. The comparison of the accuracy of MALDI-TOF MS profiling and 16S rRNA gene sequencing revealed a lack of resolution for 16S rRNA gene sequencing. In contrast, MALDI-TOF MS profiling proved to be a more reliable tool for resolving species-level variations within the Harveyi clade. Finally, combining the Luvibase with the Bruker ver.9.0.0.0 database, led to successful identification of 47 Vibrio isolates obtained from moribund abalone, seabass and oysters. Thus, the use of Luvibase allow for increased confidence in identifying Vibrio species belonging to the Harveyi clade.
RESUMO
Seafood products are widely consumed all around the world and play a significant role on the economic market. Bacteria of the Vibrio genus can contaminate seafood and thus pose a risk to human health. Three main Vibrio species, V. cholerae, V. parahaemolyticus and V. vulnificus, are potentially pathogenic to humans. These species are responsible for a dramatic increase of seafood-borne infections worldwide. Hence, early detection of total and pathogenic Vibrio is needed and should rely on quick and effective methods. This review aims to present the standard methods FDA-BAM, ISO/TS 21872-1:2007 and TS 21872-2:2007 and compare them to recent molecular biology methods including endpoint PCR, quantitative real-time PCR (qPCR) and PCR-derived methods with a focus on LAMP (loop-mediated isothermal amplification). The available methods presented here are dedicated to the detection and identification of the Vibrio species of interest in seafood.
Assuntos
Microbiologia de Alimentos/métodos , Doenças Transmitidas por Alimentos/microbiologia , Alimentos Marinhos/microbiologia , Vibrio/isolamento & purificação , DNA Bacteriano/análise , Surtos de Doenças , Humanos , Reação em Cadeia da Polimerase/métodos , Reação em Cadeia da Polimerase em Tempo Real/métodos , Vibrio/genética , Vibrio cholerae/classificação , Vibrio cholerae/genética , Vibrio cholerae/isolamento & purificação , Vibrio parahaemolyticus/classificação , Vibrio parahaemolyticus/genética , Vibrio parahaemolyticus/isolamento & purificação , Vibrio vulnificus/classificação , Vibrio vulnificus/genética , Vibrio vulnificus/isolamento & purificaçãoRESUMO
BACKGROUND: Enterococcus faecalis, generally considered as a saprophytic bowel commensal, has recently emerged as an important nosocomial pathogen causing severe urinary tract infections, surgical wound infections, bacteremia, and bacterial endocarditis. This bacterium is capable of forming biofilms on various surfaces and its high level of antibiotic resistance contributes to its pathogenicity. The aim of this study was to evaluate the effect on E. faecalis, of Substance P (SP), an antimicrobial peptide that is produced in the gut and skin. RESULTS: We found that SP did not have antibacterial activity against E. faecalis V583 (MIC >1000 µg/ml). Conversely, SP stimulated aggregation, hydrophobicity, lactic acid and tyramine production in this bacterium. The cytotoxicity and bacterial translocation were also accelerated when E. faecalis V583 were pretreated with SP before infection of intestinal Caco-2/TC7 cells. CONCLUSION: SP can modulate the physiology of E. faecalis. Extensive studies are now needed to screen within the human microbiota which bacteria are responsive to host molecules, and to identify their sensors.
RESUMO
Various phenotypes ranging from biofilm formation to pigment production have been shown to be regulated by quorum sensing (QS) in many bacteria. However, studies of the regulation of pigments produced by marine bacteria in saline conditions and of biofilm-associated phenotypes are scarcer. This study focuses on the demonstration of the existence of a QS communication system involving N-acylhomoserine lactones (AHLs) in the Mediterranean Sea strain Pseudoalteromonas ulvae TC14. We have investigated whether TC14 produces the violacein pigment, and whether intrinsic or exogenous AHLs could influence its production and modulate biofilm-associated phenotypes. Here, we demonstrate that the purple pigment produced by TC14 is violacein. The study shows that in planktonic conditions, TC14 produces more pigment in the medium in which it grows less. Using different approaches, the results also show that TC14 does not produce intrinsic AHLs in our conditions. When exogenous AHLs are added in planktonic conditions, the production of violacein is upregulated by C6-, C12-, 3-oxo-C8 and 3-oxo-C12-HSLs (homoserine lactones), and downregulated by 3-oxo-C6-HSL. In sessile conditions, 3-oxo-C8-HSL upregulates the production of violacein. The study of the biofilm-associated phenotypes shows that oxo-derived-HSLs decrease adhesion, swimming and biofilm formation. While 3-oxo-C8 and 3-oxo-C12-HSLs decrease both swimming and adhesion, 3-oxo-C6-HSLs decrease not only violacein production in planktonic conditions but also swimming, adhesion and more subtly biofilm formation. Therefore, TC14 may possess a functional LuxR-type QS receptor capable of sensing extrinsic AHLs, which controls violacein production, motility, adhesion and biofilm formation.
Assuntos
Acil-Butirolactonas/metabolismo , Biofilmes/crescimento & desenvolvimento , Indóis/metabolismo , Pigmentos Biológicos/metabolismo , Pseudoalteromonas/efeitos dos fármacos , Pseudoalteromonas/fisiologia , Percepção de Quorum , Organismos Aquáticos/efeitos dos fármacos , Organismos Aquáticos/fisiologia , Aderência Bacteriana , Locomoção , Mar MediterrâneoRESUMO
BACKGROUND: Biogenic amines are molecules with allergenic properties. They are found in fermented products and are synthesized by lactic acid bacteria through the decarboxylation of amino acids present in the food matrix. The concentration of biogenic amines in fermented foodstuffs is influenced by many environmental factors, and in particular, biogenic amine accumulation depends on the quantity of available precursors. Enological practices which lead to an enrichment in nitrogen compounds therefore favor biogenic amine production in wine. Free amino acids are the only known precursors for the synthesis of biogenic amines, and no direct link has previously been demonstrated between the use of peptides by lactic acid bacteria and biogenic amine synthesis. RESULTS: Here we demonstrate for the first time that a Lactobacillus plantarum strain isolated from a red wine can produce the biogenic amine tyramine from peptides containing tyrosine. In our conditions, most of the tyramine was produced during the late exponential growth phase, coinciding with the expression of the tyrDC and tyrP genes. The DNA sequences of tyrDC and tyrP in this strain share 98% identity with those in Lactobacillus brevis consistent with horizontal gene transfer from L. brevis to L. plantarum. CONCLUSION: Peptides amino acids are precursors of biogenic amines for Lactobacillus plantarum strain IR BL0076.
Assuntos
Lactobacillus plantarum/metabolismo , Tiramina/metabolismo , Vinho/microbiologia , Proteínas de Bactérias/biossíntese , Proteínas de Bactérias/genética , DNA Bacteriano/química , DNA Bacteriano/genética , Perfilação da Expressão Gênica , Lactobacillus plantarum/isolamento & purificação , Dados de Sequência Molecular , Análise de Sequência de DNA , Homologia de Sequência de AminoácidosRESUMO
Ornithine decarboxylase (ODC) is responsible for the production of putrescine, the major biogenic amine found in wine. Oenococcus oeni is the most important lactic acid bacterium in the winemaking process and is involved in malolactic fermentation. We report here the characterization of ODC from an O. oeni strain isolated from wine. Screening of 263 strains isolated from wine and cider from all over the world revealed that the presence of the odc gene appears to be strain specific in O. oeni. After cloning, heterologous expression in Escherichia coli, and characterization, the enzyme was found to have a molecular mass of 85 kDa and a pI of 6.2 and revealed maximal activity at pH 5.5 and an optimum temperature of 35°C. Kinetic studies showed that O. oeni ODC is specific for L-ornithine with a K(m) value of 1 mM and a V(max) of 0.57 U·mg(-1). The hypothesis that cadaverine, which results from lysine decarboxylation, may be linked to putrescine production is not valid since O. oeni ODC cannot decarboxylate L-lysine. As no lysine decarboxylase was detected in any of the O. oeni genomes sequenced, cadaverine synthesis may result from another metabolic pathway. This work is the first characterization of an ODC from a lactic acid bacterium isolated from a fermented product.