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1.
PLoS One ; 15(7): e0235641, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32614888

RESUMO

We sequenced 25 isolates of phenotypically multidrug-resistant Salmonella Indiana (n = 11), Typhimurium (n = 8), and Enteritidis (n = 6) using both MinION long-read [SQK-LSK109 and flow cell (R9.4.1)] and MiSeq short-read (Nextera XT and MiSeq Reagent Kit v2) sequencing technologies to determine the advantages of each approach in terms of the characteristics of genome structure, antimicrobial resistance (AMR), virulence potential, whole-genome phylogeny, and pan-genome. The MinION reads were base-called in real-time using MinKnow 3.4.8 integrated with Guppy 3.0.7. The long-read-only assembly, Illumina-only assembly, and hybrid assembly pipelines of Unicycler 0.4.8 were used to generate the MinION, MiSeq, and hybrid assemblies, respectively. The MinION assemblies were highly contiguous compared to the MiSeq assemblies but lacked accuracy, a deficiency that was mitigated by adding the MiSeq short reads through the Unicycler hybrid assembly which corrected erroneous single nucleotide polymorphisms (SNPs). The MinION assemblies provided similar predictions of AMR and virulence potential compared to the MiSeq and hybrid assemblies, although they produced more total false negatives of AMR genotypes, primarily due to failure in identifying tetracycline resistance genes in 11 of the 19 MinION assemblies of tetracycline-resistant isolates. The MinION assemblies displayed a large genetic distance from their corresponding MiSeq and hybrid assemblies on the whole-genome phylogenetic tree, indicating that the lower read accuracy of MinION sequencing caused incorrect clustering. The pan-genome of the MinION assemblies contained significantly more accessory genes and less core genes compared to the MiSeq and hybrid assemblies, suggesting that although these assemblies were more contiguous, their sequencing errors reduced accurate genome annotations. Our research demonstrates that MinION sequencing by itself provides an efficient assessment of the genome structure, antimicrobial resistance, and virulence potential of Salmonella; however, it is not sufficient for whole-genome phylogenetic and pan-genome analyses. MinION in combination with MiSeq facilitated the most accurate genomic analyses.


Assuntos
Farmacorresistência Bacteriana Múltipla/genética , Genoma Bacteriano , Salmonella enterica/genética , Sequenciamento Completo do Genoma/métodos , Antibacterianos/farmacologia , Genótipo , Testes de Sensibilidade Microbiana , Fenótipo , Filogenia , Plasmídeos/genética , Plasmídeos/metabolismo , Mutação Puntual , Salmonella enterica/classificação , Salmonella enterica/efeitos dos fármacos , Salmonella enterica/patogenicidade , Salmonella enteritidis/classificação , Salmonella enteritidis/efeitos dos fármacos , Salmonella enteritidis/genética , Salmonella enteritidis/patogenicidade , Salmonella typhimurium/classificação , Salmonella typhimurium/efeitos dos fármacos , Salmonella typhimurium/genética , Salmonella typhimurium/patogenicidade , Virulência
2.
PLoS One ; 15(7): e0235291, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32609730

RESUMO

Nontyphoidal Salmonella enterica (NTS) causes a substantial health burden to human populations in Canada and worldwide. Exposure sources and demographic factors vary by location and can therefore have a major impact on salmonellosis clustering. We evaluated major NTS serotypes: S. Enteritidis (n = 620), S. Typhimurium (n = 150), S. Thompson (n = 138), and S. Heidelberg (n = 136) reported in the city of Toronto, Canada, between January 1, 2015, and December 31, 2017. Cases were analyzed at the forward sortation area (FSA)-level (an area indicated by the first three characters of the postal code). Serotype-specific global and local clustering of infections were evaluated using the Moran's I method. Spatial and space-time clusters were investigated using Poisson and multinomial scan statistic models. Case-case analyses using a multinomial logistic regression model was conducted to compare seasonal and demographic factors among the different serotypes. High infection rate FSAs clustered in the central region of Toronto for S. Enteritidis, in the south-central region for S. Typhimurium, in north-west region for S. Thompson, and in the south-east region for S. Heidelberg. The relative risk ratio of S. Enteritidis infections was significantly higher in cases who reported travel outside of Ontario. The relative risk ratio of infections was significantly higher in summer for S. Typhimurium, and in fall for S. Thompson. The relative risk ratio of infection was highest for the 0-9 age group for S. Typhimurium, and the 20-39 age group for S. Heidelberg. Our study will aid public health stakeholders in designing serotype-specific geographically targeted disease prevention programs.


Assuntos
Intoxicação Alimentar por Salmonella/epidemiologia , Intoxicação Alimentar por Salmonella/microbiologia , Salmonella enterica/classificação , Salmonella enterica/isolamento & purificação , Adolescente , Adulto , Criança , Pré-Escolar , Análise por Conglomerados , Demografia , Feminino , Geografia , Humanos , Lactente , Recém-Nascido , Masculino , Pessoa de Meia-Idade , Ontário/epidemiologia , Fatores de Risco , Estações do Ano , Sorogrupo , Conglomerados Espaço-Temporais , Adulto Jovem
3.
Appl Environ Microbiol ; 86(13)2020 06 17.
Artigo em Inglês | MEDLINE | ID: mdl-32358002

RESUMO

Food safety is a new area for novel applications of metagenomics analysis, which not only can detect and subtype foodborne pathogens in a single workflow but may also produce additional information with in-depth analysis capabilities. In this study, we applied a quasimetagenomic approach by combining short-term enrichment, immunomagnetic separation (IMS), multiple-displacement amplification (MDA), and nanopore sequencing real-time analysis for simultaneous detection of Salmonella and Escherichia coli in wheat flour. Tryptic soy broth was selected for the 12-h enrichment of samples at 42°C. Enrichments were subjected to IMS using beads capable of capturing both Salmonella and E. coli MDA was performed on harvested beads, and amplified DNA fragments were subjected to DNA library preparation for sequencing. Sequencing was performed on a portable device with real-time basecalling adaptability, and resulting sequences were subjected to two parallel pipelines for further analysis. After 1 h of sequencing, the quasimetagenomic approach could detect all targets inoculated at approximately 1 CFU/g flour to the species level. Discriminatory power was determined by simultaneous detection of dual inoculums of Salmonella and E. coli, absence of detection in control samples, and consistency in microbial flora composition of the same flour samples over several rounds of experiments. The total turnaround time for detection was approximately 20 h. Longer sequencing for up to 15 h enabled serotyping for many of the samples with more than 99% genome coverage, which could be subjected to other appropriate genetic analysis pipelines in less than a total of 36 h.IMPORTANCE Enterohemorrhagic Escherichia coli (EHEC) and Salmonella are of serious concern in low-moisture foods, including wheat flour and its related products, causing illnesses, outbreaks, and recalls. The development of advanced detection methods based on molecular principles of analysis is essential to incorporate into interventions intended to reduce the risk from these pathogens. In this work, a quasimetagenomic method based on real-time sequencing analysis and assisted by magnetic capture and DNA amplification was developed. This protocol is capable of detecting multiple Salmonella and/or E. coli organisms in the sample within less than a day, and it can also generate sufficient whole-genome sequences of the target organisms suitable for subsequent bioinformatics analysis. Multiplex detection and identification were accomplished in less than 20 h and additional whole-genome analyses of different nature were attained within 36 h, in contrast to the several days required in previous sequencing pipelines.


Assuntos
Escherichia coli/isolamento & purificação , Farinha/microbiologia , Microbiologia de Alimentos/métodos , Salmonella enterica/isolamento & purificação , Sorotipagem/métodos , Escherichia coli/classificação , Separação Imunomagnética/métodos , Fenômenos Magnéticos , Metagenômica/métodos , Sequenciamento por Nanoporos/métodos , Salmonella enterica/classificação , Triticum
4.
Int J Food Microbiol ; 328: 108660, 2020 09 02.
Artigo em Inglês | MEDLINE | ID: mdl-32450393

RESUMO

Salmonella (S.) Infantis is currently the most common serovar in broilers and boiler meat in the European Union. In the field, eradication of S. Infantis in affected poultry flocks is considered extremely difficult. Despite stringent cleaning and disinfection measures between the placement of flocks, recurrent infections are often reported. So far, the efficacy of disinfectants on S. Infantis has rarely been studied. Therefore, in the present in-vitro study the bacteriostatic and bactericidal efficacy of ten commercial disinfectants were tested against seven S. Infantis field isolates. Combinations of aldehyde and quarternary ammonium were the active compounds of five, peroxygen of three, cresol and alkylamines of one disinfectant, respectively. Investigations were performed according to standard protocols and regulations. Different concentrations of disinfectants were used to test the bacteriostatic efficacy. Different temperatures and low and high protein exposures were applied as variables to investigate the bactericidal efficacy. Following neutralization of the disinfectants an additional incubation step was introduced to investigate the revitalisation potential of S. Infantis. The bacteriostatic efficacy could be assessed for seven disinfectants. For three disinfectants a bacteriostatic effect was observed when the recommended concentration was used, whereas with four disinfectants only increased concentrations led to this effect. The bactericidal efficacy was not influenced by temperature, whereas high protein exposure decreased the efficacy of nine disinfectants. Furthermore, reactivation of S. Infantis was revealed after application of disinfectants for the majority of products. Interestingly, the strain of S. Infantis influenced the efficacy of the disinfectants. Overall, products based on aldehydes and quarternary ammonium compounds proved most efficient, followed by peroxgen, cresol and alkylamines.


Assuntos
Antibacterianos/farmacologia , Desinfetantes/farmacologia , Doenças das Aves Domésticas/prevenção & controle , Salmonelose Animal/prevenção & controle , Salmonella enterica/efeitos dos fármacos , Aldeídos/farmacologia , Compostos de Amônio/farmacologia , Animais , Galinhas/microbiologia , Cresóis/farmacologia , Microbiologia de Alimentos , Carne/microbiologia , Aves Domésticas/microbiologia , Doenças das Aves Domésticas/microbiologia , Salmonelose Animal/microbiologia , Salmonella enterica/classificação , Salmonella enterica/isolamento & purificação
5.
Int J Food Microbiol ; 325: 108640, 2020 Jul 16.
Artigo em Inglês | MEDLINE | ID: mdl-32344254

RESUMO

Although a nation-wide microbiological screening program of chicken carcasses after chilling in Taiwanese chicken abattoirs has been undertaken since 2006, little is known regarding the potential sources of the Salmonella during the slaughter process. The present study provides data on the detection and serotypes of Salmonella isolated from broilers during processing and from the environment in six abattoirs in Taiwan. Overall, Salmonella were detected in 156 of 622 samples (25.1%; 95% CI: 21.7-28.7) collected. The prevalence of Salmonella varied between sampling sites with 5.8, 17.6, 31.3 and 35.5% of cloacal swabs, environmental samples prior to processing, environmental samples during processing and carcass rinse fluid, respectively, being positive (χ2 = 51.3, p < 0.0001). A total of 15 serotypes were identified from the 156 Salmonella isolates with S. Albany (41.7%) S. Schwarzengrund (20.5%), S. Kentucky (12.8%) and S. Tennessee (5.1%) being the most commonly isolated serotypes. Characterization of 156 isolates by Pulse Field Gel Electrophoresis (PFGE) identified 50 PFGE types. Typing confirmed the presence of the same PFGE type at multiple stages during processing including plucking, evisceration, chilling and post-chilling. The abattoir environment and intestinal contents of chickens are important sources of Salmonella in broiler chicken abattoirs, with the same PFGE types detected at different stages of processing both before and during slaughtering. It is concluded that Salmonella isolates present in the environment and intestinal contents of processed birds survived in the abattoir environment resulting in subsequent carcass contamination along the processing chain including plucking, evisceration, chilling and post-chilling.


Assuntos
Galinhas/microbiologia , Carne/microbiologia , Salmonella enterica/isolamento & purificação , Matadouros , Animais , Eletroforese em Gel de Campo Pulsado , Microbiologia de Alimentos , Intestinos/microbiologia , Salmonelose Animal/microbiologia , Salmonella enterica/classificação , Sorogrupo , Taiwan
6.
Lett Appl Microbiol ; 71(2): 138-145, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32333808

RESUMO

Salmonellosis is a leading bacterial cause of foodborne illness, and numerous Salmonella enterica serovars have been responsible for foodborne outbreaks. In the United States outbreaks are often linked to poultry and poultry-related products. The prevalence of Salmonella serovar Infantis has been increasing in poultry processing facilities over the past few years and in 2018 was identified as the causative agent for a large multistate outbreak linked to raw chicken. CRISPR-typing is a subtyping approach based on PCR and the sequencing of two Salmonella loci, CRISPR1 and CRISPR2. CRISPR-typing was used to interrogate 138 recent (2018-2019) isolates and genomes of ser. Infantis. Results show that the CRISPR elements are remarkably conserved in this serovar. The most conserved spacers, and those also unique to ser. Infantis, were used as targets to develop a ser. Infantis-specific qPCR assay. This assay was able to detect ser. Infantis in mixed serovar cultures of Salmonella, down to 0·1% of the population, highlighting the utility of this molecular approach in improving surveillance sensitivity for this important food safety pathogen. SIGNIFICANCE AND IMPACT OF THE STUDY: The incidence of human salmonellosis cases caused by Salmonella enterica serovar Infantis (ser. Infantis) has been increasing, as has its prevalence in broiler chickens, which are a frequent reservoir of Salmonella. A cluster of ser. Infantis genetically linked to an outbreak strain have been identified in numerous processing facilities. A qPCR assay targeting CRISPR elements that are unique to ser. Infantis has been developed and can detect this serovar directly from mixed cultures. This assay is sensitive enough to reveal ser. Infantis within a mixed Salmonella population where it constitutes only 0·1% of the population. The rapid nature of qPCR lends this assay to high-throughput screening of poultry samples to detect this important pathogen.


Assuntos
Tipagem Molecular/métodos , Doenças das Aves Domésticas/epidemiologia , Reação em Cadeia da Polimerase em Tempo Real/métodos , Intoxicação Alimentar por Salmonella/epidemiologia , Salmonella enterica/classificação , Animais , Galinhas/microbiologia , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas/genética , Surtos de Doenças , Humanos , Aves Domésticas/microbiologia , Salmonella enterica/genética , Salmonella enterica/isolamento & purificação , Sorogrupo , Estados Unidos/epidemiologia
7.
Lett Appl Microbiol ; 71(2): 210-217, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32304584

RESUMO

The effects of soil type and temperature on the survival of a cocktail of five Salmonella enterica serotypes (Enteritidis, Infantis, Montevideo, Typhimurium and Zanzibar) in manure-amended soils under controlled laboratory conditions was assessed. Containers of clay loam or sandy soil, unaltered or amended with 2% (w/w) poultry manure, were inoculated with S. enterica (~5 log10 CFU per gram) and held at 5, 21 or 37°C for 6 weeks. Statistical analysis of the persistence of S. enterica identified a significant three-way interaction between soil type, manure amendment and temperature. Clay loam soils and lower temperatures tended to support S. enterica persistence over 6 weeks with only 1- and 2-log reductions respectively. In contrast, sand and higher temperatures resulted in a 4-log and either 3- to 4-log reductions respectively. Manure amendment had an overarching effect of reducing die-off of S. enterica in comparison with unamended soils. This study highlights that a large component of variation of the rate of S. enterica reduction in soils may be attributed to combinations of environmental factors, in particular, soil type and temperature. It further underscores the importance of risk management strategies and industry guidelines based on local data and that reflect the diversity of prevailing horticultural production environments. SIGNIFICANCE AND IMPACT OF THE STUDY: The persistence of Salmonella enterica in soil environments was shown to be significantly influenced by a range of individual and interacting environmental effects, including temperature, soil type and amendment addition. This indicates that current horticultural food safety management systems which employ a uniform prescribed exclusion period between application of manure and time of harvest may be unfit for purpose under certain conditions by either underestimating or overestimating pathogen die-off. These findings support exclusion periods that account for a range of environmental factors including temperature, soil type and growing region that may be more appropriate to manage microbiological risks associated with soil which has been amended with manure.


Assuntos
Esterco/microbiologia , Aves Domésticas/microbiologia , Salmonella enterica/crescimento & desenvolvimento , Salmonella enterica/isolamento & purificação , Animais , Temperatura Alta , Salmonella enterica/classificação , Solo/química , Microbiologia do Solo
8.
BMC Genomics ; 21(1): 202, 2020 Mar 04.
Artigo em Inglês | MEDLINE | ID: mdl-32131727

RESUMO

BACKGROUND: Salmonella enterica subsp. enterica serovar Napoli (S. Napoli) is among the top serovars causing human infections in Italy, although it is relatively uncommon in other European countries; it is mainly isolated from humans and the environment, but neither the reservoir nor its route of infection are clearly defined. This serovar is characterized by high genomic diversity, and molecular evidences revealed important similarities with typhoidal serovars. RESULTS: 179 S. Napoli genomes as well as 239 genomes of typhoidal and non-typhoidal serovars were analyzed in a comparative genomic study. Phylogenetic analysis and draft genome characterization in terms of Multi Locus Sequence Typing (MLST), plasmid replicons, Salmonella Pathogenicity Islands (SPIs), antimicrobial resistance genes (ARGs), phages, biocide and metal-tolerance genes confirm the high genetic variability of S. Napoli, also revealing a within-serovar phylogenetic structure more complex than previously known. Our work also confirms genomic similarity of S. Napoli to typhoidal serovars (S. Typhi and S. Paratyphi A), with S. Napoli samples clustering primarily according to ST, each being characterized by specific genomic traits. Moreover, two major subclades of S. Napoli can be clearly identified, with ST-474 being biphyletic. All STs span among isolation sources and years of isolation, highlighting the challenge this serovar poses to define its epidemiology and evolution. Altogether, S. Napoli strains carry less SPIs and less ARGs than other non-typhoidal serovars and seldom acquire plasmids. However, we here report the second case of an extended-spectrum ß-lactamases (ESBLs) producing S. Napoli strain and the first cases of multidrug resistant (MDR) S. Napoli strains, all isolated from humans. CONCLUSIONS: Our results provide evidence of genomic plasticity of S. Napoli, highlighting genomic similarity with typhoidal serovars and genomic features typical of non-typhoidal serovars, supporting the possibility of survival in different niches, both enteric and non-enteric. Presence of horizontally acquired ARGs and MDR profiles rises concerns regarding possible selective pressure exerted by human environment on this pathogen.


Assuntos
DNA Bacteriano/genética , Infecções por Salmonella/microbiologia , Salmonella enterica/classificação , Sequenciamento Completo do Genoma/métodos , Farmacorresistência Bacteriana Múltipla , Ilhas Genômicas , Genômica , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Itália , Filogenia , Plasmídeos/genética , Salmonella enterica/genética , Salmonella enterica/imunologia , Salmonella enterica/isolamento & purificação , Sorogrupo , Febre Tifoide/microbiologia , Resistência beta-Lactâmica
9.
Microb Genom ; 6(2)2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-32003708

RESUMO

To establish the prevalence of mobile colistin resistance (mcr) genes amongst Salmonella enterica isolates obtained through public health surveillance in England (April 2014 to September 2017), 33 205 S. enterica genome sequences obtained from human, food, animal and environmental isolates were screened for the presence of mcr variants 1 to 8. The mcr-positive genomes were assembled, annotated and characterized according to plasmid type. Nanopore sequencing was performed on six selected isolates with putative novel plasmids, and phylogenetic analysis was used to provide an evolutionary context for the most commonly isolated clones. Fifty-two mcr-positive isolates were identified, of which 32 were positive for mcr-1, 19 for mcr-3 and 1 for mcr-5. The combination of Illumina and Nanopore sequencing identified three novel mcr-3 plasmids and one novel mcr-5 plasmid, as well as the presence of chromosomally integrated mcr-1 and mcr-3. Monophasic S. enterica serovar Typhimurium accounted for 27/52 (52 %) of the mcr-positive isolates, with the majority clustering in clades associated with travel to Southeast Asia. Isolates in these clades were associated with a specific plasmid range and an additional extended-spectrum beta-lactamase genotype. Routine whole-genome sequencing for public health surveillance provides an effective screen for novel and emerging antimicrobial determinants, including mcr. Complementary long-read technologies elucidated the genomic context of resistance determinants, offering insights into plasmid dissemination and linkage to other resistance genes.


Assuntos
Antibacterianos/farmacologia , Proteínas de Bactérias/genética , Colistina/farmacologia , Genoma Bacteriano , Salmonella enterica/genética , Animais , Antibacterianos/metabolismo , Proteínas de Bactérias/metabolismo , Farmacorresistência Bacteriana , Inglaterra/epidemiologia , Microbiologia Ambiental , Microbiologia de Alimentos , Humanos , Testes de Sensibilidade Microbiana , Filogenia , Plasmídeos/genética , Plasmídeos/metabolismo , Vigilância em Saúde Pública , Infecções por Salmonella/epidemiologia , Infecções por Salmonella/microbiologia , Salmonelose Animal/epidemiologia , Salmonelose Animal/microbiologia , Salmonella enterica/classificação , Salmonella enterica/efeitos dos fármacos , Salmonella enterica/isolamento & purificação
10.
Epidemiol Infect ; 148: e51, 2020 02 13.
Artigo em Inglês | MEDLINE | ID: mdl-32052718

RESUMO

In June 2017, an outbreak of Salmonella Kottbus infection was suspected in Germany. We investigated the outbreak with whole-genome sequencing (WGS) and a case-control study. Forty-six isolates from 69 cases were subtyped. Three WGS clusters were identified: cluster 1 (n = 36), cluster 2 (n = 5) and cluster 3 (n = 3). Compared to controls, cluster 1 cases more frequently consumed raw smoked ham (odds ratio (OR) 10, 95% confidence interval (CI) 1.2-88) bought at supermarket chain X (OR 36, 95% CI 4-356; 9/10 consumed ham Y). All four cluster 2 cases interviewed had consumed quail eggs. Timely WGS was invaluable in distinguishing concurrent outbreaks of a rare Salmonella serotype.


Assuntos
Surtos de Doenças , Doenças Transmitidas por Alimentos/epidemiologia , Epidemiologia Molecular/métodos , Tipagem Molecular/métodos , Infecções por Salmonella/epidemiologia , Salmonella enterica/classificação , Sequenciamento Completo do Genoma/métodos , Adulto , Idoso , Idoso de 80 Anos ou mais , Estudos de Casos e Controles , Análise por Conglomerados , Comportamento Alimentar , Feminino , Doenças Transmitidas por Alimentos/microbiologia , Alemanha/epidemiologia , Humanos , Masculino , Pessoa de Meia-Idade , Infecções por Salmonella/microbiologia , Salmonella enterica/genética , Salmonella enterica/isolamento & purificação
11.
Intern Med ; 59(2): 289-292, 2020 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-31534082

RESUMO

Non-typhoidal Salmonella (NTS) infection is a major pathogen causing gastroenteritis among immunocompetent adults. NTS infection is mainly transmitted by contaminated food and water, but some cases are transmitted by animal contact. Salmonella enterica subsp. enterica serovar Poona (S. Poona) is an NTS usually transmitted by reptiles, and cases including outbreaks of gastroenteritis have been reported previously. However, invasive infections due to this organism among immunocompetent adults are rare. We herein report a case of a 39-year-old man who was admitted to our hospital for a fever and headache. Blood cultures were positive for S. Poona, although he did not recall any exposure to reptiles. He was treated successfully with intravenous ceftriaxone without any subsequent complications. This case implies that NTS bacteremia can occur in immunocompetent adults, and the diagnosis may be challenging since there may be no clear exposure or focal physical signs.


Assuntos
Bacteriemia/diagnóstico , Infecções por Salmonella/diagnóstico , Salmonella enterica/classificação , Adulto , Animais , Antibacterianos/uso terapêutico , Bacteriemia/tratamento farmacológico , Bacteriemia/microbiologia , Hemocultura , Ceftriaxona/uso terapêutico , Humanos , Japão , Masculino , Infecções por Salmonella/tratamento farmacológico , Sorogrupo , Tóquio
12.
Equine Vet J ; 52(1): 98-103, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30991461

RESUMO

BACKGROUND: Salmonella enterica subsp. enterica serovar Abortusequi (S. Abortusequi) is a serotype restricted to equines, which produces abortion outbreaks. Nowadays the disease is being reported in different countries including Argentina thus generating an important impact in the equine industry. Molecular characterization of the 95 kb virulence plasmid and the spvC gene of S. Abortusequi demonstrated their importance in the pathogenicity of the serotype. In the last decades, high clonality of S. Abortusequi was identified in Japan, Mongolia and Croatia. OBJECTIVES: The aim of this work was to characterize S. Abortusequi isolates obtained in Argentina between 2011 and 2016 by virulence-gene profiling and pulsed-field gel electrophoresis. STUDY DESIGN: Case report. METHODS: S. Abortusequi isolates were studied by virulence-gene profiling and pulsed-field gel electrophoresis. RESULTS: Four virulence profiles and nine pulsed-field gel electrophoresis pulsotypes were identified among the 27 isolates included in the study. Different strains were found in the same outbreak and/or farm suggesting the presence of different sources of infection or mutation of isolates. MAIN LIMITATIONS: The number of related and nonrelated strains. More isolates may be necessary for a more intensive study. CONCLUSIONS: Most strains presented the same virulence profile, being positive for all the studied genes except gipA and sopE1, which are involved in intestinal virulence. Only few isolates showed different results in the same outbreak or farm. Unlike other studies, our results demonstrate a considerable diversity of S. Abortusequi pulsed-field gel electrophoresis pulsotypes, which suggests that different sources of infection may be involved within the same outbreak.


Assuntos
Genótipo , Doenças dos Cavalos/microbiologia , Salmonelose Animal/microbiologia , Salmonella enterica/genética , Animais , Argentina/epidemiologia , Doenças dos Cavalos/epidemiologia , Cavalos , Salmonelose Animal/epidemiologia , Salmonella enterica/classificação , Salmonella enterica/patogenicidade , Transcriptoma , Virulência
13.
Appl Environ Microbiol ; 86(3)2020 01 21.
Artigo em Inglês | MEDLINE | ID: mdl-31732576

RESUMO

Salmonella enterica serovar Dublin is a cattle-adapted S. enterica serovar causing both intestinal and systemic infection in its bovine host, and it is also a serious threat to human health. The present study aimed to determine the population structure of S Dublin isolates obtained from Danish cattle herds and to investigate how cattle isolates relate to Danish human isolates, as well as to non-Danish human and bovine isolates. Phylogenetic analysis of 197 Danish cattle isolates from 1996 to 2016 identified three major clades corresponding to distinct geographical regions of cattle herds. Persistence of closely related isolates within the same herd and their circulation between epidemiologically linked herds for a period of more than 20 years were demonstrated. These findings suggest that a lack of internal biosecurity and, to some extent, also a lack of external biosecurity in the herds have played an important role in the long-term persistence of S Dublin in Danish cattle herds in the period investigated. Global population analysis revealed that Danish cattle isolates clustered separately from bovine isolates from other countries, whereas human isolates were geographically spread. Resistance genes were not commonly demonstrated in Danish bovine isolates; only the isolates within one Danish clade were found to often harbor two plasmids of IncFII/IncFIB and IncN types, the latter plasmid carrying bla TEM-1, tetA, strA, and strB antibiotic resistance genes.IMPORTANCE S Dublin causes economic losses in cattle production, and the bacterium is a public health concern. A surveillance and control program has been in place in Denmark since 2002 with the ultimate goal to eradicate S Dublin from Danish cattle herds; however, a small proportion of herds have remained positive for many years. In this study, we demonstrate that herds with persistent infection often were infected with the same strain for many years, indicating that internal biosecurity has to be improved to curb the infection. Further, domestic cases of S Dublin infection in humans were found to be caused both by Danish cattle isolates and by isolates acquired abroad. This study shows the strength of whole-genome sequencing to obtain detailed information on epidemiology of S Dublin and allows us to suggest internal biosecurity as a main way to control this bacterium in Danish cattle herds.


Assuntos
Doenças dos Bovinos/epidemiologia , Salmonelose Animal/epidemiologia , Infecções por Salmonella/epidemiologia , Salmonella enterica/isolamento & purificação , Animais , Bovinos , Doenças dos Bovinos/microbiologia , Dinamarca/epidemiologia , Humanos , Filogenia , Prevalência , Estudos Retrospectivos , Infecções por Salmonella/microbiologia , Salmonelose Animal/microbiologia , Salmonella enterica/classificação , Sorogrupo , Sequenciamento Completo do Genoma/veterinária
14.
Jpn J Infect Dis ; 73(2): 119-123, 2020 Mar 24.
Artigo em Inglês | MEDLINE | ID: mdl-31666490

RESUMO

Non-typhoidal salmonellae (NTS) are a major cause of acute diarrhea with characteristic multidrug resistance (MDR). In a hospital-based study, 81 NTS were isolated and tested for serotypes and antimicrobial resistance (AMR). Salmonella enterica isolates were classified into 7 different typable serovars, however, 19 (23%) isolates remained untypable. The most common serovars were S. Kentucky (48%), and S. Virchow (22%). Most of the NTS isolates displayed resistance to nalidixic acid (NA) (73%), ciprofloxacin (CIP) (48%), ampicillin (AM) and norfloxacin (NOR) (36% each), and gentamicin (CN) (31%). The AMR profiles for CN and NA; and AM, CIP, NA and NOR, were found to be high in S. Virchow (83%) and S. Kentucky (43%), respectively. Analysis of the pulsed-field gel electrophoresis patterns of S. Kentucky revealed 3 clusters. S. Kentucky has clones closely related to become prominent in recent years in Delhi. The AMR appears to be consistent with the change in MDR patterns during 2014-2017. The observed prevalence of S. Kentucky and S. Virchow in large numbers of diarrheal cases is novel. The NTS are mostly resistant to fluoroquinolones, which is the current drug of choice for treating diarrheal cases. MDR is very common among clonally related S. Kentucky.


Assuntos
Diarreia/epidemiologia , Farmacorresistência Bacteriana Múltipla , Infecções por Salmonella/epidemiologia , Salmonella enterica/efeitos dos fármacos , Antibacterianos/farmacologia , Criança , Pré-Escolar , Diarreia/microbiologia , Eletroforese em Gel de Campo Pulsado , Humanos , Índia/epidemiologia , Kentucky , Prevalência , Salmonella enterica/classificação , Sorogrupo
15.
Pathog Glob Health ; 113(6): 282-287, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31818238

RESUMO

Infections caused by non-typhoidal Salmonella (NTS) are common around the world, with high morbidity and mortality rates recorded annually. Salmonella serovars harbor plasmids of various sizes which may play roles in antibiotic resistance and virulence. The aim of this study was to profile and determine the role of plasmids in ciprofloxacin resistance and virulence of Salmonella serovars. Using alkaline lysis method 25 NTS serovars from food animals and humans were assayed for plasmids. Isolates ability to resist healthy human serum, bind congo red, produce hemolysin and susceptibility to ciprofloxacin before and after plasmid curing were evaluated. Mobility of plasmids was determined by conjugation. Fifteen (60%) of the 25 Salmonella serovars harbored plasmids with sizes ranging from 0.4 to 38.4 kb. S. Budapest serovars harbored 5-9 plasmids, while S. Essen and S. Mura had six plasmids each. S. Chomedey and a S. Budapest serovar were sensitive to ciprofloxacin after plasmid curing while other serovars remained resistant to ciprofloxacin after plasmid curing. All Salmonella isolates had the ability to withstand human serum before and after plasmid curing, however, some serovars lost their ability to bind congo red after plasmid curing. All Salmonella isolates that initially displayed hemolysin activity retained their ability after curing. Thirteen (86.7%) of the 15 serovars that harbored plasmids conjugatively transferred their plasmids to E. coli K-12 (DH5α). Having Salmonella serovars that harbor transferrable plasmids in the food chain can drive antibiotic resistance and enhanced virulence of otherwise less virulence strains.


Assuntos
Plasmídeos/genética , Salmonelose Animal/microbiologia , Infecções por Salmonella/microbiologia , Salmonella enterica/genética , Salmonella enterica/patogenicidade , Animais , Antibacterianos/farmacologia , Bovinos/microbiologia , Galinhas/microbiologia , Humanos , Testes de Sensibilidade Microbiana , Nigéria , Plasmídeos/metabolismo , Salmonella enterica/classificação , Salmonella enterica/isolamento & purificação , Ovinos/microbiologia , Virulência
16.
PLoS Negl Trop Dis ; 13(12): e0007917, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31790418

RESUMO

BACKGROUND: Limited epidemiological and antimicrobial resistance data are available on Salmonella enterica from sub-Saharan Africa. We determine the prevalence of resistance to antibiotics in isolates in the Central African Republic (CAR) between 2004 and 2013 and the genetic basis for resistance to third-generation cephalosporin (C3G). METHODOLOGY/PRINCIPAL FINDINGS: A total of 582 non-duplicate human clinical isolates were collected. The most common serotype was Typhimurium (n = 180, 31% of the isolates). A randomly selected subset of S. Typhimurium isolates were subtyped by clustered regularly interspaced short palindromic repeat polymorphism (CRISPOL) typing. All but one invasive isolate tested (66/68, 96%) were associated with sequence type 313. Overall, the rates of resistance were high to traditional first-line drugs (18-40%) but low to many other antimicrobials, including fluoroquinolones (one resistant isolate) and C3G (only one ESBL-producing isolate). The extended-spectrum beta-lactamase (ESBL)-producing isolate and three additional ESBL isolates from West Africa were studied by whole genome sequencing. The blaCTX-M-15 gene and the majority of antimicrobial resistance genes found in the ESBL isolate were present in a large conjugative IncHI2 plasmid highly similar (> 99% nucleotide identity) to ESBL-carrying plasmids found in Kenya (S. Typhimurium ST313) and also in West Africa (serotypes Grumpensis, Havana, Telelkebir and Typhimurium). CONCLUSIONS/SIGNIFICANCE: Although the prevalence of ESBL-producing Salmonella isolates was low in CAR, we found that a single IncHI2 plasmid-carrying blaCTX-M-15 was widespread among Salmonella serotypes from sub-Saharan Africa, which is of concern.


Assuntos
Farmacorresistência Bacteriana , Infecções por Salmonella/epidemiologia , Salmonella enterica/classificação , Salmonella enterica/efeitos dos fármacos , Sorogrupo , Antibacterianos/farmacologia , República Centro-Africana/epidemiologia , Genes Bacterianos , Genótipo , Técnicas de Genotipagem , Humanos , Plasmídeos/análise , Prevalência , Estudos Retrospectivos , Salmonella enterica/genética , Salmonella enterica/isolamento & purificação
17.
Nat Commun ; 10(1): 4927, 2019 10 30.
Artigo em Inglês | MEDLINE | ID: mdl-31666527

RESUMO

Raman optical spectroscopy promises label-free bacterial detection, identification, and antibiotic susceptibility testing in a single step. However, achieving clinically relevant speeds and accuracies remains challenging due to weak Raman signal from bacterial cells and numerous bacterial species and phenotypes. Here we generate an extensive dataset of bacterial Raman spectra and apply deep learning approaches to accurately identify 30 common bacterial pathogens. Even on low signal-to-noise spectra, we achieve average isolate-level accuracies exceeding 82% and antibiotic treatment identification accuracies of 97.0±0.3%. We also show that this approach distinguishes between methicillin-resistant and -susceptible isolates of Staphylococcus aureus (MRSA and MSSA) with 89±0.1% accuracy. We validate our results on clinical isolates from 50 patients. Using just 10 bacterial spectra from each patient isolate, we achieve treatment identification accuracies of 99.7%. Our approach has potential for culture-free pathogen identification and antibiotic susceptibility testing, and could be readily extended for diagnostics on blood, urine, and sputum.


Assuntos
Antibacterianos/uso terapêutico , Bactérias/classificação , Infecções Bacterianas/diagnóstico , Aprendizado Profundo , Análise Espectral Raman/métodos , Bactérias/química , Infecções Bacterianas/tratamento farmacológico , Infecções Bacterianas/microbiologia , Técnicas de Tipagem Bacteriana , Candida/química , Candida/classificação , Enterococcus/química , Enterococcus/classificação , Escherichia coli/química , Escherichia coli/classificação , Humanos , Klebsiella/química , Klebsiella/classificação , Modelos Logísticos , Staphylococcus aureus Resistente à Meticilina/química , Staphylococcus aureus Resistente à Meticilina/classificação , Testes de Sensibilidade Microbiana , Redes Neurais de Computação , Análise de Componente Principal , Proteus mirabilis/química , Proteus mirabilis/classificação , Pseudomonas aeruginosa/química , Pseudomonas aeruginosa/classificação , Salmonella enterica/química , Salmonella enterica/classificação , Análise de Célula Única , Staphylococcus aureus/química , Staphylococcus aureus/classificação , Streptococcus/química , Streptococcus/classificação , Máquina de Vetores de Suporte
18.
BMC Genomics ; 20(1): 870, 2019 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-31730461

RESUMO

BACKGROUND: Salmonella Infantis (S. Infantis) is one of the most frequent Salmonella serovars isolated from human cases of salmonellosis and the most detected serovar from animal and food sources in Europe. The serovar is commonly associated with poultry and there is increasing concern over multidrug resistant clones spreading worldwide, as the dominating clones are characterized by presence of large plasmids carrying multiple resistance genes. Increasing the knowledge of the S. Infantis population and evolution is important for understanding and preventing further spread. In this study, we analysed a collection of strains representing different decades, sources and geographic locations. We analysed the population structure and the accessory genome, in particular we identified prophages with a view to understand the role of prophages in relation to the evolution of this serovar. RESULTS: We sequenced a global collection of 100 S. Infantis strains. A core-genome SNP analysis separated five strains in e-Burst Group (eBG) 297 with a long branch. The remaining strains, all in eBG31, were divided into three lineages that were estimated to have separated approximately 150 years ago. One lineage contained the vast majority of strains. In five of six clusters, no obvious correlation with source or geographical locations was seen. However, one cluster contained mostly strains from human and avian sources, indicating a clone with preference for these sources. The majority of strains within this cluster harboured a pESI-like plasmid with multiple resistance genes. Another lineage contained three genetic clusters with more rarely isolated strains of mainly animal origin, possibly less sampled or less infectious clones. Conserved prophages were identified in all strains, likely representing bacteriophages which integrated into the chromosome of a common ancestor to S. Infantis. We also saw that some prophages were specific to clusters and were probably introduced when the clusters were formed. CONCLUSIONS: This study analysed a global S. Infantis population and described its genetic structure. We hypothesize that the population has evolved in three separate lineages, with one more successfully emerging lineage. We furthermore detected conserved prophages present in the entire population and cluster specific prophages, which probably shaped the population structure.


Assuntos
Farmacorresistência Bacteriana Múltipla/genética , Genoma Bacteriano , Filogenia , Polimorfismo de Nucleotídeo Único , Salmonella enterica/genética , Animais , Antibacterianos/farmacologia , Ásia/epidemiologia , Galinhas , Europa (Continente)/epidemiologia , Humanos , Família Multigênica , Filogeografia , Doenças das Aves Domésticas/epidemiologia , Doenças das Aves Domésticas/microbiologia , Prófagos , Salmonelose Animal/epidemiologia , Salmonelose Animal/microbiologia , Salmonella enterica/classificação , Salmonella enterica/efeitos dos fármacos , Salmonella enterica/isolamento & purificação , Estados Unidos/epidemiologia , Sequenciamento Completo do Genoma
19.
J Vet Diagn Invest ; 31(6): 818-827, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31646949

RESUMO

Rapid identification of the infecting Salmonella serovar from porcine diagnostic samples is vital to allow implementation of appropriate on-farm treatment and management decisions. Although identification at the serogroup level can be rapidly achieved at most veterinary diagnostic laboratories, final Salmonella serovar identification often takes several weeks because of the limited number of reference laboratories performing the complex task of serotyping. Salmonella serogroup B, currently the dominant serogroup identified from swine clinical samples in the United States, contains serovars that vary from highly pathogenic to minimally pathogenic in swine. We determined the frequency of detection of individual group B serovars at the Iowa State Veterinary Diagnostic Laboratory from 2008 to 2017, and validated a multiplex real-time PCR (rtPCR) to distinguish pathogenic serogroup B serovars from those of lesser pathogenicity. Our results indicate that, since 2014, Salmonella enterica ssp. enterica serovar 4,[5],12:i:- has been the dominant serovar identified from swine clinical samples at the ISU-VDL, with S. Typhimurium now the second most common serovar identified. We developed a rtPCR to allow rapid differentiation of samples containing S. 4,[5],12:i:- and S. Typhimurium from samples containing serovars believed to be of less pathogenicity, such as S. Agona and S. Derby. When combined with enrichment culture, this rtPCR has the ability to significantly improve the time to final serovar identification of the 2 most commonly identified pathogenic Salmonella serovars in swine, and allows rapid implementation of serovar-specific intervention strategies.


Assuntos
Reação em Cadeia da Polimerase em Tempo Real/veterinária , Salmonelose Animal/diagnóstico , Salmonella enterica/classificação , Sorotipagem/veterinária , Doenças dos Suínos/diagnóstico , Animais , Reação em Cadeia da Polimerase Multiplex/métodos , Reação em Cadeia da Polimerase Multiplex/veterinária , Reação em Cadeia da Polimerase em Tempo Real/métodos , Salmonelose Animal/microbiologia , Salmonella enterica/patogenicidade , Sorogrupo , Sorotipagem/métodos , Suínos , Doenças dos Suínos/microbiologia , Estados Unidos , Virulência
20.
Int J Food Microbiol ; 310: 108359, 2019 Nov 16.
Artigo em Inglês | MEDLINE | ID: mdl-31655448

RESUMO

A dearth of knowledge exists on the pathogen-commodity association of Salmonella enterica on cucumber, despite cucumbers being implicated in multiple salmonellosis outbreaks in recent years in the U.S. Data are lacking on cultivar susceptibility to Salmonella colonization in relation to heterogeneity in fruit surface morphology. Further, fruit waxing is a common practice in wholesale cucumber to preserve the water content of fruit and prolong shelf-life, but its impact on epiphytic microbiota is not well studied. This study investigated the survival of Salmonella Newport and S. Javiana on the surface of six cucumber cultivars of varying surface morphology and the effect of fruit waxing on the persistence of S. Newport. S. Newport and S. Javiana were spot inoculated onto fruit of cultivars 'Marketmore 97', 'Patio Snacker' and 'Corinto' (varieties with trichomes or spines), and 'Bella', 'Pepinex' and 'Summer Dance' (glabrous or smooth varieties). Cucumbers were held at room temperature for 24 h before inoculated sections of exocarp were excised for enteropathogen enumeration. S. Javiana persisted at higher populations than S. Newport (1.0 and 1.5 log CFU g-1 exocarp decline, respectively) after 24 h (P < 0.05). Fruit waxing lessened S. Newport population decline after 24 h on all cultivars tested (P < 0.01). While there was a ~2 log CFU g-1 exocarp decline in populations on unwaxed cucumbers, the reduction on waxed cucumbers was ~1 log CFU g-1 exocarp. A cultivar effect was also observed; 'Summer Dance' supported consistently higher and 'Corinto' and 'Marketmore 97' (though for S. Javiana only) consistently lower Salmonella levels. Cultivars with trichomes were associated with lower populations than glabrous cultivars (P < 0.05). Our data showed that Salmonella persistence on fruit differed for the two serotypes tested and that cultivar differences play a role in determining cucumber surface favourability for pathogen colonization. Additionally, fruit waxing may elevate the risk of Salmonella persistence on cucumber.


Assuntos
Cucumis sativus/microbiologia , Manipulação de Alimentos/métodos , Microbiologia de Alimentos , Conservantes de Alimentos/normas , Salmonella enterica/classificação , Salmonella enterica/fisiologia , Sorogrupo
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