RESUMO
The genus Listeria comprises two pathogenic species, L. monocytogenes and L. ivanovii, as well as four nonpathogenic species, L. innocua, L. weishimeri, L. seeligeri, and L. grayi. Within L. monocytogenes, lineages I and II are responsible for most listeriosis cases, while lineage III strains are rarely associated with human morbidity but providing important clues for Listeria evolution. The gene lmo0038, belonging to the peptidylarginine deiminase family, was involved in the optimal growth under stress conditions, including low pH and heat shock (52 degrees C), and virulence potential. Further, this gene was specific to L. monocytogenes lineages I and II and L. ivanovii with significant similarities at nucleotide and amino acid levels. A novel multiplex PCR, based on lmo0038 in combination with optimized iap migration profiles, was developed for simultaneous identification of Listeria species and discrimination of L. monocytogenes lineage III, with a detection limit down to 1.0-9.0 x 10(2) CFU/mL. This assay was evaluated by 119 suspected Listeria food-related isolates and corrected 4 and 5 misidentifications by Listeria selective agar plate screening and API system, respectively. Therefore, this one-step molecular assay provides a rapid, reliable, and inexpensive screening test to detect Listeria species-particularly, the pathogenic species in surveillance programs concerning food safety and foodborne disease cases.
Assuntos
Temperatura Alta , Hidrolases/genética , Listeria monocytogenes/classificação , Listeria monocytogenes/enzimologia , Listeria/classificação , Listeria/enzimologia , Sequência de Aminoácidos , Sequência de Bases , DNA Bacteriano/análise , Microbiologia de Alimentos , Humanos , Concentração de Íons de Hidrogênio , Hidrolases/química , Hidrolases/fisiologia , Listeria/patogenicidade , Listeria monocytogenes/patogenicidade , Listeriose/microbiologia , Reação em Cadeia da Polimerase , Desiminases de Arginina em Proteínas , RNA Ribossômico/genéticaRESUMO
In this study, twenty L. monocytogenes food-related isolates collected from eastern China Zhejiang province were compared by in vivo LD50 assays as well as in vitro cytopathic plaque forming assay. Nineteen L. monocytogenes isolates (19/20) were as virulent as reference strain 10403S, while the isolate M4 had low pathogenicity. The unique isolate M4 fell into lineage III based on the partial nucleotide variations of actA, while the other isolates belonged to the more common lineages I and II. L. monocytogenes isolates were grouped in 17 to 19 subtypes using pulsed-field gel electrophoresis (PFGE) with SmaI digestion, and multilocus sequence typing (MLST) based on three virulence genes (actA, inlA and inlB) and four housekeeping genes (betL, dat, recA and sigB). The virulence genes based MLST had better discriminatory power than that targeting the housekeeping genes (0.990 vs 0.895), similar to PFGE (0.976). An isolate from the processing desk was found having the same pulsotype as the two isolates from final shrimp products in the same plant, indicating that process contamination could be the source of Listeria contamination.
Assuntos
Proteínas de Bactérias/genética , Contaminação de Alimentos/análise , Microbiologia de Alimentos , Listeria monocytogenes/patogenicidade , Fatores de Virulência/genética , Animais , Sequência de Bases , Embrião de Galinha , China , Análise por Conglomerados , DNA Bacteriano/química , DNA Bacteriano/genética , Eletroforese em Gel de Campo Pulsado , Indústria de Processamento de Alimentos , Doenças Transmitidas por Alimentos/microbiologia , Genótipo , Humanos , Dose Letal Mediana , Listeria monocytogenes/classificação , Listeria monocytogenes/genética , Listeriose/microbiologia , Proteínas de Membrana , Camundongos , Dados de Sequência Molecular , Polimorfismo Genético , Virulência/genéticaRESUMO
This study was aimed to examine the genetic characteristics of 44 Streptococcus suis type 2 (SS2) isolates and the virulence attributes of 23 representative isolates. Multilocus sequence typing revealed five sequence types (ST1, ST7, ST28, ST86, and ST162) with 19 isolates assigned to ST7 (43.2%), 14 to ST1 (31.8%), and 9 to ST28 (20.5%). PCR typing of the pilus gene clusters classified the isolates into three types: A (72.7%), B (22.7%), and N (4.5%). All isolates of pilus types A and N were assigned to the ST1 complex containing ST1, ST7, and ST86, while those of type B belonged to the ST27 complex comprising ST28 and ST162. Only two strains had the putative pathogenicity island 89-kb cluster (89K) and were of type N. The type B strains had a significantly lower adhesion, were more readily killed by macrophage, and had lower virulence to mice than those of types A and N. We conclude that SS2 strains of both ST1 and ST27 complexes, parallel to pilus types A and B, were prevalent in the pig populations in Zhejiang Province, and ST7 and ST1 strains were the predominant genotypes in the diseased pigs with pneumonia.