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1.
Sci Rep ; 9(1): 11460, 2019 Aug 07.
Artigo em Inglês | MEDLINE | ID: mdl-31391518

RESUMO

The clinical use of the antibiotic erythromycin (ery) is hampered owing to the spread of resistance genes that are mostly mutating rRNA around the ery binding site at the entrance to the protein exit tunnel. Additional effective resistance mechanisms include deletion or insertion mutations in ribosomal protein uL22, which lead to alterations of the exit tunnel shape, located 16 Å away from the drug's binding site. We determined the cryo-EM structures of the Staphylococcus aureus 70S ribosome, and its ery bound complex with a two amino acid deletion mutation in its ß hairpin loop, which grants the bacteria resistance to ery. The structures reveal that, although the binding of ery is stable, the movement of the flexible shorter uL22 loop towards the tunnel wall creates a wider path for nascent proteins, thus enabling bypass of the barrier formed by the drug. Moreover, upon drug binding, the tunnel widens further.

2.
Int J Food Microbiol ; 300: 14-21, 2019 Jul 02.
Artigo em Inglês | MEDLINE | ID: mdl-30991234

RESUMO

It is well established, that certain bacteria within the Brochothrix, Carnobacterium, Lactobacillus, Lactococcus, and Leuconostoc genera have an important role in the spoilage of chill stored poultry meat packaged in modified atmosphere. However, little is known about the role of microorganisms that are difficult to culture and the microbiota during poultry spoilage. We combined traditional cultivation and culture-independent 16S rRNA amplicon sequencing to investigate the microbiota encompassing putative bacteria of whole broiler meat, packaged in modified atmosphere, during and exceeding shelf-life. Samples were taken from 6 flocks during independent slaughter days. Additional samples were analysed from the production line. There was a significant difference in the microbial community structure of 80%O2/20%CO2 retail packaged broiler meat during different times of shelf-life, mainly due to an increase of species within the Brochothrix, Carnobacterium, Vagococcus, and Janthinobacterium genera. These genera were already detected four to eight days after slaughter. However, no significant difference between flocks with respect to the microbiota encompassing putative spoilage bacteria was observed when examined in retail packaged broilers, slaughtered at the same abattoir on different days. Our study also showed that lactic acid bacteria within the Vagococcus genus can constitute a dominating part of the later shelf-life microbiota in fresh whole broiler meat packaged in 80%O2/20%CO2 modified atmosphere. A single operational taxonomic unit (OTU) assigned as Janthinobacterium lividum, an occasional spoiler of meat products, was identified as a major part of the microbiota in late shelf life broiler meat and swab samples in the cooling facility at the slaughter house production line. The combination of traditional cultivation and culture-independent methods provided a great insight into the microbiota of broiler meat during shelf-life and identified a potential point of contamination in the production line for cold tolerant Janthinobacterium.


Assuntos
Fenômenos Fisiológicos Bacterianos , Galinhas/microbiologia , Microbiologia de Alimentos , Carne/microbiologia , Microbiota/fisiologia , Matadouros , Animais , Bactérias/genética , Bactérias/crescimento & desenvolvimento , Carnobacterium/genética , Carnobacterium/fisiologia , Galinhas/genética , Embalagem de Alimentos , Microbiota/genética , RNA Ribossômico 16S/genética
3.
Nat Commun ; 10(1): 1124, 2019 03 08.
Artigo em Inglês | MEDLINE | ID: mdl-30850636

RESUMO

Antimicrobial resistance (AMR) is a serious threat to global public health, but obtaining representative data on AMR for healthy human populations is difficult. Here, we use metagenomic analysis of untreated sewage to characterize the bacterial resistome from 79 sites in 60 countries. We find systematic differences in abundance and diversity of AMR genes between Europe/North-America/Oceania and Africa/Asia/South-America. Antimicrobial use data and bacterial taxonomy only explains a minor part of the AMR variation that we observe. We find no evidence for cross-selection between antimicrobial classes, or for effect of air travel between sites. However, AMR gene abundance strongly correlates with socio-economic, health and environmental factors, which we use to predict AMR gene abundances in all countries in the world. Our findings suggest that global AMR gene diversity and abundance vary by region, and that improving sanitation and health could potentially limit the global burden of AMR. We propose metagenomic analysis of sewage as an ethically acceptable and economically feasible approach for continuous global surveillance and prediction of AMR.


Assuntos
Bactérias/efeitos dos fármacos , Farmacorresistência Bacteriana Múltipla/genética , Genes Bacterianos , Metagenoma , Esgotos/microbiologia , África , Ásia , Bactérias/classificação , Bactérias/genética , Bactérias/isolamento & purificação , Monitoramento Epidemiológico , Europa (Continente) , Humanos , Metagenômica/métodos , Consórcios Microbianos/efeitos dos fármacos , Consórcios Microbianos/genética , América do Norte , Oceania , Saúde da População , Fatores Socioeconômicos , América do Sul
4.
Euro Surveill ; 23(6)2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29439754

RESUMO

Background and aimPlasmid-mediated colistin resistance mechanisms have been identified worldwide in the past years. A multiplex polymerase chain reaction (PCR) protocol for detection of all currently known transferable colistin resistance genes (mcr-1 to mcr-5, and variants) in Enterobacteriaceae was developed for surveillance or research purposes. Methods: We designed four new primer pairs to amplify mcr-1, mcr-2, mcr-3 and mcr-4 gene products and used the originally described primers for mcr-5 to obtain a stepwise separation of ca 200 bp between amplicons. The primer pairs and amplification conditions allow for single or multiple detection of all currently described mcr genes and their variants present in Enterobacteriaceae. The protocol was validated testing 49 European Escherichia coli and Salmonella isolates of animal origin. Results: Multiplex PCR results in bovine and porcine isolates from Spain, Germany, France and Italy showed full concordance with whole genome sequence data. The method was able to detect mcr-1, mcr-3 and mcr-4 as singletons or in different combinations as they were present in the test isolates. One new mcr-4 variant, mcr-4.3, was also identified. Conclusions: This method allows rapid identification of mcr-positive bacteria and overcomes the challenges of phenotypic detection of colistin resistance. The multiplex PCR should be particularly interesting in settings or laboratories with limited resources for performing genetic analysis as it provides information on the mechanism of colistin resistance without requiring genome sequencing.


Assuntos
Antibacterianos/farmacologia , Colistina/farmacologia , Enterobacteriaceae/efeitos dos fármacos , Enterobacteriaceae/genética , Proteínas de Escherichia coli/genética , Plasmídeos/genética , Salmonella/efeitos dos fármacos , Salmonella/genética , Enterobacteriaceae/isolamento & purificação , Infecções por Enterobacteriaceae/tratamento farmacológico , Infecções por Enterobacteriaceae/microbiologia , Proteínas de Escherichia coli/metabolismo , Humanos , Proteínas de Membrana , Testes de Sensibilidade Microbiana , Reação em Cadeia da Polimerase Multiplex , Plasmídeos/metabolismo , Salmonella/isolamento & purificação , Transferases (Outros Grupos de Fosfato Substituídos)
5.
Euro Surveill ; 22(50)2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-29258647

RESUMO

In August 2017, an outbreak of six listeriosis cases in Denmark was traced to cold-smoked salmon, using epidemiological investigations and whole-genome sequencing (WGS) analyses. Exchange of genome sequences allowed identification in France of a food isolate from a salmon-derived product and a human isolate from 2016 within the same cgMLST cluster as the Danish isolates (L2-SL8-ST8-CT771). The salmon product came from a third European Union country. WGS can rapidly link human cases and food isolates across Europe.


Assuntos
Surtos de Doenças/estatística & dados numéricos , Doenças Transmitidas por Alimentos , Genoma Bacteriano/genética , Listeria monocytogenes/genética , Listeriose/epidemiologia , Salmão/microbiologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Dinamarca/epidemiologia , Emigração e Imigração , Feminino , Doenças Transmitidas por Alimentos/epidemiologia , Doenças Transmitidas por Alimentos/microbiologia , França/epidemiologia , Humanos , Listeria monocytogenes/isolamento & purificação , Listeriose/diagnóstico , Listeriose/microbiologia , Masculino , Pessoa de Meia-Idade , Epidemiologia Molecular , Tipagem de Sequências Multilocus , Análise de Sequência de DNA , Sequenciamento Completo do Genoma
6.
Structure ; 25(8): 1233-1241.e3, 2017 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-28689968

RESUMO

Erythromycin is a clinically useful antibiotic that binds to an rRNA pocket in the ribosomal exit tunnel. Commonly, resistance to erythromycin is acquired by alterations of rRNA nucleotides that interact with the drug. Mutations in the ß hairpin of ribosomal protein uL22, which is rather distal to the erythromycin binding site, also generate resistance to the antibiotic. We have determined the crystal structure of the large ribosomal subunit from Deinococcus radiodurans with a three amino acid insertion within the ß hairpin of uL22 that renders resistance to erythromycin. The structure reveals a shift of the ß hairpin of the mutated uL22 toward the interior of the exit tunnel, triggering a cascade of structural alterations of rRNA nucleotides that propagate to the erythromycin binding pocket. Our findings support recent studies showing that the interactions between uL22 and specific sequences within nascent chains trigger conformational rearrangements in the exit tunnel.


Assuntos
Proteínas de Bactérias/química , Proteínas Ribossômicas/química , Antibacterianos/química , Antibacterianos/farmacologia , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Sítios de Ligação , Deinococcus/química , Eritromicina/química , Eritromicina/farmacologia , Mutação , Ligação Proteica , RNA Ribossômico/química , RNA Ribossômico/metabolismo , Proteínas Ribossômicas/genética , Proteínas Ribossômicas/metabolismo
7.
Int J Food Microbiol ; 249: 72-76, 2017 May 16.
Artigo em Inglês | MEDLINE | ID: mdl-28324679

RESUMO

Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) is increasingly related to human infections. Farmers and veterinarians have the highest risk, but infections have also occurred in individuals without prior contact to livestock. Clonal complex (CC) 398 is the predominant LA-MRSA lineage causing human infections, and although pigs are the major source of CC398 worldwide, poultry and other animals are also reservoirs. This raises concern for transmission of MRSA via meat. In this study, the occurrence and characteristics of S. aureus isolated from Danish retail meat were examined with main focus on chicken meat. A total of 145 meat samples from Danish supermarkets were examined, including chicken (Danish, n=102), turkey (non-Danish origin; n=23), and pork (Danish, n=20). S. aureus was detected in 69% of the meat samples. MRSA was detected in 19 meat samples (13%), resulting in MRSA prevalence of 4% of chicken, 52% of turkey, and 15% of pork. Three MRSA positive samples were obtained by direct plating (Brilliance MRSA2), whereas 16 MRSA positive samples were detected only after enrichment (TSB+6.5% NaCl and Brilliance MRSA2). Based on spa typing, 68% of MRSA isolates belonged to CC398 (spa t034, t011, t2582, t108), and hereof one isolate derived from chicken (1%). Further findings were spa type t1430 (CC9) in turkey samples (16%) and the human-associated t008 (CC8) in chicken samples (16%). In conclusion, S. aureus was readily detected in Danish retail meat, but presence of MRSA in chicken meat is rare and it is unlikely to be an important transmission factor of MRSA to humans.


Assuntos
Contaminação de Alimentos/análise , Staphylococcus aureus Resistente à Meticilina/isolamento & purificação , Carne Vermelha/microbiologia , Animais , Técnicas de Tipagem Bacteriana , Galinhas/microbiologia , Dinamarca , Microbiologia de Alimentos , Humanos , Testes de Sensibilidade Microbiana , Aves Domésticas/microbiologia , Infecções Estafilocócicas/epidemiologia , Sus scrofa/microbiologia , Suínos/microbiologia , Perus/microbiologia
8.
BMC Res Notes ; 6: 322, 2013 Aug 13.
Artigo em Inglês | MEDLINE | ID: mdl-23941403

RESUMO

BACKGROUND: Arcobacter spp. have in recent years received increasing interest as potential emerging enteropathogens and zoonotic agents. They are associated with various animals including poultry and can be isolated from meat products. The possibilities of persistence and cross-contamination in slaughterhouses during meat processing are not well established. We have evaluated the occurrence and persistence of Arcobacter spp. in a Danish slaughterhouse and determined the sensitivity of isolates to sodium hypochlorite, a commonly used biocide. RESULTS: Arcobacter contamination was examined in a broiler slaughterhouse by selective enrichment of 235 swabs from the processing line during two production days and after sanitizing in between. In total 13.6% of samples were positive for A. butzleri with the majority (29 of 32 isolates) originating from the evisceration machine. No Arcobacter spp. was isolated after cleaning. A. butzleri isolates confirmed by PCR were typed by multilocus sequence typing (MLST) resulting in 10 new sequence types (STs). Two sequence types were isolated on both processing days. Minimum inhibitory concentration (MIC) to sodium hypochlorite was determined to 0.5% hypochlorite biocide (500 ppm chlorine) for most isolates, which allows growth of A. butzleri within the working concentration of the biocide (0.2 - 0.5%). CONCLUSIONS: A. butzleri was readily isolated from a Danish broiler slaughterhouse, primarily in the evisceration machine. Typing by MLST showed high strain variability but the recurrence of two STs indicate that some persistence or cross-contamination takes place. Importantly, the isolates tolerated sodium hypochlorite, a biocide commonly employed in slaughterhouse sanitizing, at levels close to the disinfection concentration, and thus, A. butzleri may survive the disinfection process although this was not observed in our study.


Assuntos
Arcobacter/efeitos dos fármacos , Arcobacter/genética , Técnicas de Tipagem Bacteriana , Galinhas/microbiologia , Desinfetantes/farmacologia , Farmacorresistência Bacteriana , Contaminação de Alimentos , Microbiologia de Alimentos , Carne/microbiologia , Tipagem de Sequências Multilocus , Hipoclorito de Sódio/farmacologia , Matadouros/instrumentação , Animais , Arcobacter/classificação , DNA Bacteriano/análise , Dinamarca , Contaminação de Equipamentos/prevenção & controle , Desenho de Equipamento , Contaminação de Alimentos/prevenção & controle , Humanos , Testes de Sensibilidade Microbiana , Reação em Cadeia da Polimerase Multiplex , Filogenia
9.
mBio ; 3(5): e00190-12, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22930338

RESUMO

UNLABELLED: Fermented sausages, although presumed safe for consumption, sometimes cause serious bacterial infections in humans that may be deadly. Not much is known about why and when this is the case. We tested the hypothesis that residual veterinary antibiotics in meat can disrupt the fermentation process, giving pathogenic bacteria a chance to survive and multiply. We found that six commercially available starter cultures were susceptible to commonly used antibiotics, namely, oxytetracycline, penicillin, and erythromycin. In meat, statutorily tolerable levels of oxytetracycline and erythromycin inhibited fermentation performance of three and five of the six starter cultures, respectively. In model sausages, the disruption of meat fermentation enhanced survival of the pathogens Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium compared to successful fermentations. Our work reveals an overlooked risk associated with the presence of veterinary drugs in meat. IMPORTANCE: Antibiotics have for a long time been used as growth promoters in farm animals, and while they are banned as such in Europe, their clinical use in farm animals still accounts for the majority of consumption. Here, we examined how acceptable levels of antibiotics in meat influence fermentation. Our results show that commonly used bacterial starter cultures are sensitive to residual antibiotics at or near statutorily tolerable levels, and as a result, processed sausages may indeed contain high levels of pathogens. Our findings provide a possible explanation for outbreaks and disease cases associated with consumption of fermented sausages and offer yet another argument for limiting the use of antimicrobials in farm animals.


Assuntos
Antibacterianos/farmacologia , Bactérias/efeitos dos fármacos , Bactérias/crescimento & desenvolvimento , Carne/microbiologia , Drogas Veterinárias/farmacologia , Animais , Bactérias/patogenicidade , Infecções Bacterianas , Eritromicina/farmacologia , Europa (Continente) , Fermentação , Doenças Transmitidas por Alimentos , Humanos , Testes de Sensibilidade Microbiana , Oxitetraciclina/farmacologia , Penicilinas/farmacologia
10.
Appl Environ Microbiol ; 77(23): 8456-8, 2011 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-21984236

RESUMO

We describe a simple method for stabilizing and extracting high-quality prokaryotic RNA from meat. Heat and salt stress of Escherichia coli and Salmonella spp. in minced meat reproducibly induced dnaK and otsB expression, respectively, as observed by quantitative reverse transcription-PCR (>5-fold relative changes). Thus, the method is applicable in studies of bacterial gene expression in a meat matrix.


Assuntos
Escherichia coli/efeitos dos fármacos , Escherichia coli/efeitos da radiação , Perfilação da Expressão Gênica/métodos , Carne/microbiologia , Salmonella/efeitos dos fármacos , Salmonella/efeitos da radiação , Escherichia coli/genética , Manipulação de Alimentos/métodos , Temperatura Alta , RNA Bacteriano/genética , RNA Bacteriano/isolamento & purificação , Reação em Cadeia da Polimerase em Tempo Real , Salmonella/genética , Sais/toxicidade
11.
Int J Food Microbiol ; 131(2-3): 256-9, 2009 May 31.
Artigo em Inglês | MEDLINE | ID: mdl-19297052

RESUMO

Arcobacter butzleri is prevalent on chicken products. Arcobacter spp. are generally isolated in only low numbers from the chicken gut, so chicken carcasses may be contaminated by A. butzleri that proliferate in the slaughterhouse environment. To address this issue, we examined the behaviour of A. butzleri ATCC 49616 and newly isolated A. butzleri strains under conditions likely to prevail in the slaughterhouse environment using a chicken meat juice medium (CMJ). CMJ supported growth of A. butzleri at 15 degrees C, the recognised minimal growth temperature of this organism, and at 10 degrees C. At 5 degrees C, CMJ enhanced survival of A. butzleri as compared with survival in Brain Heart Infusion with less than a one log reduction after 77 days incubation. Lastly, we examined the ability of A. butzleri to form biofilms and found that the organism produces biofilm at temperatures ranging from 5 to 37 degrees C. Given the ability to survive, multiply and form biofilm under chilled conditions A. butzleri appears well suited for establishment in food processing and slaughterhouse environments.


Assuntos
Arcobacter/crescimento & desenvolvimento , Biofilmes/crescimento & desenvolvimento , Galinhas/microbiologia , Temperatura Baixa , Microbiologia de Alimentos , Carne/microbiologia , Viabilidade Microbiana , Matadouros , Animais , Arcobacter/genética , Arcobacter/isolamento & purificação
12.
Int J Food Microbiol ; 124(3): 250-9, 2008 Jun 10.
Artigo em Inglês | MEDLINE | ID: mdl-18456355

RESUMO

Thirteen storage trials and ten challenge tests were carried out to examine microbial changes, spoilage and the potential growth of Listeria monocytogenes in brined shrimp (Pandalus borealis). Shrimp in brine as well as brined and drained shrimp in modified atmosphere packaging (MAP) were produced and studied. Different recipes were used to study the effect of preserving parameters (organic acids, pH and NaCl) on growth of microorganisms and shelf life at 7-8 degrees C or 12 degrees C. Particularly, brines with different concentrations of (i) benzoic, citric and sorbic acids or (ii) acetic, citric and lactic acids were studied. Furthermore, the effect of adding diacetate to brined shrimp was evaluated. A single batch of cooked and peeled shrimp was used to study both industrially and manually processed brined shrimp with respect to the effect of process hygiene on microbial changes and the shelf life of products. Concentrations of microorganisms on newly produced brined shrimp from an industrial scale processing line were 1.0-2.3 log (CFU g(-1)) higher than comparable concentrations in manually processed samples. This resulted in a substantially shorter shelf life and a more diverse spoilage microflora of the industrially processed brined shrimp. In addition, shelf life of brined shrimp was affected by the types and concentrations of organic acids and by the storage temperature as expected. The effect of MAP was less pronounced. Eighty-two isolates from the spoilage microflora of brined shrimp were identified and they included 53 lactic acid bacteria, 6 coagulase negative Staphylococcus spp., 18 Pseudomonas fluorescens and 5 yeast isolates. After storage at 7 degrees C, P. fluorescens, Enterococcus-like isolates, E. malodoratus, Carnobacterium maltaromaticum, coagulase negative Staphylococcus spp. and Lactobacillus sakei constituted the dominating microflora of shrimp in brines that contained benzoic, citric and sorbic acids as preservatives. L. sakei dominated the spoilage microflora of brined and drained MAP shrimp, and of brined shrimp preserved using acetic, citric and lactic acids, irrespective of packaging conditions. Shrimp in brine with benzoic, citric and sorbic acids prevented growth of L. monocytogenes during more than 40 days at 7 degrees C when the preserving parameters resembled those of commercial products. However, small changes in the preserving parameters and, particularly, reduced concentrations of benzoic acid led to growth of L. monocytogenes in brined shrimp. The present study provides significant new information on microbial changes, shelf life and growth of L. monocytogenes in brined shrimp. This information can facilitate development of new and safe brined shrimp products.


Assuntos
Contaminação de Alimentos/análise , Embalagem de Alimentos/métodos , Conservação de Alimentos/métodos , Listeria monocytogenes/crescimento & desenvolvimento , Pandalidae/microbiologia , Frutos do Mar/microbiologia , Animais , Contagem de Colônia Microbiana , Qualidade de Produtos para o Consumidor , Manipulação de Alimentos/métodos , Microbiologia de Alimentos , Humanos , Concentração de Íons de Hidrogênio , Higiene , Refrigeração , Fatores de Tempo , Vácuo
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