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1.
Int J Food Microbiol ; 391-393: 110151, 2023 Apr 16.
Artigo em Inglês | MEDLINE | ID: mdl-36871395

RESUMO

Salmonella is the main human pathogen present in the poultry chain. Salmonella Heidelberg is one of the most important serovars for public health since it has been frequently isolated in broiler chickens from different countries and may present multidrug resistance (MDR). This study was carried out with 130 S. Heidelberg isolates collected from pre-slaughter broiler farms in 2019 and 2020 in 18 cities from three Brazilian states to study relevant aspects regarding their genotypic and phenotypic resistance. The isolates were tested and identified using somatic and flagellar antiserum (0:4, H:2, and H:r), and an antimicrobial susceptibility test (AST) was performed against 11 antibiotics for veterinary use. The strains were typed by Enterobacterial Repetitive Intergenic Consensus (ERIC)-PCR, and representatives of the main clusters of the identified profiles were sequenced by Whole Genome Sequencing (WGS). AST results showed that all isolates were resistant to sulfonamide, 54 % (70/130) were resistant to amoxicillin, and only one was sensitive to tetracycline. Twelve isolates (15.4 %) were MDR. The dendrogram obtained from the ERIC-PCR showed that the strains were grouped into 27 clusters with similarity above 90 %, with some isolates showing 100 % similarity but with different phenotypic profiles of antimicrobial resistance. Identical strains collected on the same farm on other dates were identified, indicating that they were residents. WGS identified 66 antibiotic-resistance genes. The sul2 (present in all sequenced samples) and tet(A) genes were highlighted and validated in the experimental analysis. The fosA7 gene was also identified in all sequenced samples, but resistance was not observed in the phenotypic test, possibly due to the heteroresistance of the S. Heidelberg strains evaluated. Considering that chicken meat is one of the most consumed meats in the world, the data obtained in the present study can corroborate the mapping of the origin and trends of antimicrobial resistance.


Assuntos
Galinhas , Farmacorresistência Bacteriana Múltipla , Animais , Humanos , Brasil , Farmacorresistência Bacteriana Múltipla/genética , Galinhas/microbiologia , Testes de Sensibilidade Microbiana , Salmonella , Antibacterianos/farmacologia
2.
Food Microbiol ; 110: 104152, 2023 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-36462834

RESUMO

The importance of thermotolerant Campylobacter spp. as a food-borne pathogen continues to increase and there is a great need for rapid quantitative results in routine diagnostics. However, currently, only the culture-based ISO method is authorized for use in the context of official food control. The present study therefore aimed to assess the suitability of a qPCR method for a rapid quantitative determination of Campylobacter spp. at different stages in the poultry production chain and its equivalence with the culture-based method. Samples from two processors were collected and evaluated both separately and together. Censored regression (Tobit) models have been used to establish a relationship between Campylobacter qPCR counts on the carcasses and explanatory variables of processor and meat counts. Further, correlations of qPCR Campylobacter spp. counts at the different stages of production were calculated. In addition, the comparative data between microbiological enumeration and qPCR results were statistically analyzed. In the correlation calculation of the qPCR results, a highly significant relationship between the Campylobacter spp. counts of the neck skin samples to breast fillet and leg samples could be calculated, indicating a good prediction of Campylobacter spp. loads in these samples. The intercalating dye ethidium monoazide (EMA) was used to see whether the correlations between microbiological counts and qPCR results were improved by pretreating fecal and cecal samples before qPCR analysis. It was shown that the observed values of scatter plots between the qPCR-based and the culture-based methods were strongly correlated. However, on average, the qPCR results were two log10 CFU/mL levels higher than the microbiological counts. However, the classical culture-based method for food hygiene risk assessment cannot be replaced one-to-one by the qPCR or EMA-qPCR. The qPCR method can rather be used for the rapid identification of particularly highly contaminated flocks.


Assuntos
Campylobacter , Cadeia Alimentar , Animais , Reação em Cadeia da Polimerase em Tempo Real , Galinhas , Campylobacter/genética , Fezes
3.
Microbiol Spectr ; 10(5): e0247922, 2022 10 26.
Artigo em Inglês | MEDLINE | ID: mdl-36194132

RESUMO

High-resolution and efficient typing for the bacterial pathogen is essential for tracking the sources, detecting or diagnosing variants, and conducting a risk assessment. However, a systematic in-field investigation of Salmonella along the food chain has not been documented. This study assessed 12 typing methods, such as antimicrobial-resistance (AMR) gene profile typing, Core Genome Multilocus Sequence Typing (cgMLST), and CRISPR multi-virulence locus sequence typing (CRISPR-MVLST), to evaluate their effectiveness for use in routine monitoring of foodborne Salmonella transmission along the poultry production chain. During 2015-16, a total of 1,064 samples were collected from poultry production chain, starting from breeding farms and slaughterhouses to the markets of Zhejiang province in China. A total of 61 consecutive unique Salmonella isolates recovered from these samples were selected for genome sequencing and further comparative typing analysis. Traditional typing methods, including serotyping, AMR phenotype-based typing, as well as modern genotyping approaches, were evaluated and compared by their discrimination index (DI). The results showed that the serotyping method identified nine serovars. The gold standard cgMLST method indicated only 18 different types (DI = 0.8541), while the CRISPR-MVLST method detected 30 types (DI = 0.9628), with a higher DI than all examined medium-resolution WGS-based genotyping methods. We demonstrate that the CRISPR-MVLST might be used as a tool with high discriminatory power, comparable ease of use, ability of tracking the source of Salmonella strains along the food chain and indication of genetic features especially virulence genes. The available methods with different purposes and laboratory expertise were also illustrated to assist in rational implementation. IMPORTANCE In public health field, high-resolution and efficient typing of the bacterial pathogen is essential, considering source-tracking and risk assessment are fundamental issues. Currently, there are no recommendations for applying molecular characterization methods for Salmonella along the food chain, and a systematic in-field investigation comparing subtyping methods in the context of routine surveillance was partially addressed. Using 1,064 samples along a poultry production chain with a considerable level of Salmonella contamination, we collected representative isolates for genome sequencing and comparative analysis by using 12 typing techniques, particularly with whole-genome sequence (WGS) based methods and a recently invented CRISPR multi-virulence locus sequence typing (CRISPR-MVLST) method. CRISPR-MVLST is identified as a tool with higher discriminatory power compared with medium-resolution WGS-based typing methods, comparable ease of use and proven ability of tracking Salmonella isolates. Besides, we also offer recommendations for rational choice of subtyping methods to assist in better implementation schemes.


Assuntos
Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Salmonella , Salmonella/genética , Tipagem de Sequências Multilocus/métodos , Sorogrupo , Análise de Sequência de DNA
4.
Poult Sci ; 101(2): 101611, 2022 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-34953378

RESUMO

The study of non-typhoid Salmonella in broiler integrations has been limited by the resolution of typing techniques. Although serotyping of Salmonella isolates is used as a traditional approach, it is not of enough resolution to clearly understand the dynamics of this pathogen within poultry companies. The aim of this research was to investigate the epidemiology and population dynamics of Salmonella serotypes in 2 poultry integrations using a whole genome sequencing approach. Two hundred and forty-three Salmonella isolates recovered from the broiler production chain of 2 integrated poultry companies were whole genome sequenced and analyzed with dedicated databases and bioinformatic software. The analyses of sequences revealed that S. Infantis was the most frequent serotype (82.3%). Most isolates showed a potential for resistance against medically important antibiotics and disinfectants. Furthermore, 97.5% of isolates harbored the pESI-like mega plasmid, that plays an important role in the global dissemination of AMR. SNP tree analysis showed that there were clones that are niche-specific while other ones were distributed throughout the broiler production chains. In this study, we demonstrated the potential of whole genome sequencing analysis for a comprehensive understanding of Salmonella distribution in integrated poultry companies. Data obtained with these techniques allow determination of the presence of genetic factors that play an important role in the environmental fitness and pathogenicity of Salmonella.


Assuntos
Galinhas , Aves Domésticas , Animais , Antibacterianos , Genoma Bacteriano , Salmonella/genética , Sequenciamento Completo do Genoma/veterinária
5.
Antonie Van Leeuwenhoek ; 108(5): 1227-38, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-26337044

RESUMO

Antimicrobial resistance profiles and presence of resistance determinants and integrons were evaluated in Salmonella enterica strains from Brazilian poultry. The analysis of 203 isolates showed that those from the poultry environment (88 isolates) were significantly more resistant to antimicrobials than isolates from other sources, particularly those isolated from poultry by-product meal (106 isolates). Thirty-seven isolates were resistant to at least three antimicrobial classes. Class 1 integrons were detected in 26 isolates, and the analysis of the variable region between the 5' conserved segment (CS) and 3' CS of each class 1 integron-positive isolate showed that 13 contained a typical 3' CS and 14 contained an atypical 3' CS. One Salmonella Senftenberg isolate harbored two class 1 integrons, showing both typical and atypical 3' CSs. The highest percentage of resistance was found to sulfonamides, and sul genes were detected in the majority of the resistant isolates. Aminoglycoside resistance was detected in 50 isolates, and aadA and aadB were present in 28 and 32 isolates, respectively. In addition, strA and strB were detected in 78.1 and 65.6% isolates resistant to streptomycin, respectively. Twenty-one isolates presented reduced susceptibility to ß-lactams and harbored bla(TEM), bla(CMY), and/or bla(CTX-M). Forty isolates showed reduced susceptibility to tetracycline, and most presented tet genes. These results highlight the importance of the environment as a reservoir of resistant Salmonella, which may enable the persistence of resistance determinants in the poultry production chain, contributing, therefore, to the debate regarding the impacts that antimicrobial use in animal production may exert in human health.


Assuntos
Anti-Infecciosos/farmacologia , Farmacorresistência Bacteriana , Microbiologia de Alimentos , Produtos da Carne/microbiologia , Salmonella enterica/classificação , Salmonella enterica/efeitos dos fármacos , Animais , Brasil , Farmacorresistência Bacteriana/genética , Genes Bacterianos , Integrons , Testes de Sensibilidade Microbiana , Aves Domésticas , Salmonella enterica/genética , Salmonella enterica/isolamento & purificação
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