RESUMO
Staphylococcus aureus are extremely important microorganisms, either from an epidemiological point of view or as a pathogen, responsible for causing a series of infectious processes, whether simple, restricted to the skin, or invasive infections such as bacteremia. The emergence of Oxacillin Sensitive-Methicillin Resistant S.aureus (OS-MRSA) isolates has imposed difficulties in the treatment of patients with staphylococcal infection, as such isolates can be mistakenly classified as sensitive and lead to failure of the therapy used. Thus, the objective of this study is to evaluate the prevalence, and genotypic and phenotypic characteristics, of OS-MRSA isolates, from bloodstream infections, collected from patients admitted to a hospital in southern Brazil, as well as to evaluate the treatment used. For this, 801 unique isolates of S. aureus, collected from blood cultures, between January 2011 and December 2020 were evaluated. Of these, 96 isolates were classified as sensitive to oxacillin. The isolates were identified and had their sensitivity profile performed by manual and automated methods. The minimum inhibitory concentration for vancomycin, daptomycin, oxacillin, linezolid and teicoplanine was performed by e-test. The mecA, vanA genes, typing of the SCCmec elements, as well as the search for the icaA, tst-1 and pvl virulence genes were performed by PCR. Biofilm formation was performed using the crystal violet technique. The Sequence Type (ST), as well as the Clonal Complex (CC) of the isolates was evaluated by the RTq -PCR. The clinical characteristics of the patients were evaluated through an active search in medical records. After investigating the mecA gene, 27.1% (26/96) of the isolates were considered OS-MRSA, with SCCmec type I being the most prevalent, 46.1% (12/26). Among the evaluated isolates, 41% (9/22) were included in CC5 and ST9. As for virulence, all isolates were positive for the icaA gene and characterized as strong biofilm formers. The pvl gene was found in 92.3% (24/26) of the isolates and the toxic shock syndrome toxin was present in 61.5% of the isolates (16/26). All isolates were negative for the presence of the van A gene. As for the clinical outcome, 73% (19/26) of the patients were discharged from the hospital and 27% (7/26) died. It was possible to observe a high frequency of OS-MRSA isolates causing bloodstream infections. Furthermore, such isolates contain several virulence genes, which may contribute to a worse clinical outcome.
Assuntos
Antibacterianos , Bacteriemia , Proteínas de Bactérias , Genótipo , Staphylococcus aureus Resistente à Meticilina , Testes de Sensibilidade Microbiana , Oxacilina , Proteínas de Ligação às Penicilinas , Infecções Estafilocócicas , Centros de Atenção Terciária , Humanos , Oxacilina/farmacologia , Brasil , Antibacterianos/farmacologia , Infecções Estafilocócicas/microbiologia , Centros de Atenção Terciária/estatística & dados numéricos , Bacteriemia/microbiologia , Proteínas de Bactérias/genética , Proteínas de Ligação às Penicilinas/genética , Staphylococcus aureus Resistente à Meticilina/genética , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Staphylococcus aureus Resistente à Meticilina/isolamento & purificação , Staphylococcus aureus Resistente à Meticilina/classificação , FenótipoRESUMO
The presence of Staphylococcus spp. resistant to methicillin in the nasal cavity of swine has been previously reported. Considering the possible occurrence of bacterial resistance and presence of resistance genes in intensive swine breeding and the known transmissibility and dispersion potential of such genes, this study aimed to investigate the prevalence of resistance to different antibiotics and the presence of the mecA resistance gene in Staphylococcus spp. from piglets recently housed in a nursery. For this, 60 nasal swabs were collected from piglets at the time of their housing in the nursery, and then Staphylococcus spp. were isolated and identified in coagulase-positive (CoPS) and coagulase-negative (CoNS) isolates. These isolates were subjected to the disk-diffusion test to evaluate the bacterial resistance profile and then subjected to molecular identification of Staphylococcus aureus and analyses of the mecA gene through polymerase chain reaction. Of the 60 samples collected, 60 Staphylococcus spp. were isolated, of which 38 (63.33%) were classified as CoNS and 22 (36.67%) as CoPS. Of these, ten (45.45%) were identified as Staphylococcus aureus. The resistance profile of these isolates showed high resistance to different antibiotics, with 100% of the isolates resistant to chloramphenicol, clindamycin, and erythromycin, 98.33% resistant to doxycycline, 95% resistant to oxacillin, and 85% resistant to cefoxitin. Regarding the mecA gene, 27 (45%) samples were positive for the presence of this gene, and three (11.11%) were phenotypically sensitive to oxacillin and cefoxitin. This finding highlights the importance of researching the phenotypic profile of resistance to different antimicrobials and resistance genes in the different phases of pig rearing to identify the real risk of these isolates from a One Health perspective. The present study revealed the presence of samples resistant to different antibiotics in recently weaned production animal that had not been markedly exposed to antimicrobials as growth promoters or even as prophylactics. This information highlights the need for more research on the possible sharing of bacteria between sows and piglets, the environmental pressure within production environments, and the exposure of handlers during their transport, especially considering the community, hospital, and political importance of the presence of circulating resistant strains.
Assuntos
Infecções Estafilocócicas , Doenças dos Suínos , Animais , Suínos , Feminino , Cefoxitina , Coagulase , Cavidade Nasal/química , Proteínas de Bactérias/genética , Proteínas de Ligação às Penicilinas/genética , Testes de Sensibilidade Microbiana/veterinária , Antibacterianos/farmacologia , Oxacilina , Infecções Estafilocócicas/microbiologia , Infecções Estafilocócicas/veterinária , Staphylococcus aureus/genética , Staphylococcus/genética , Doenças dos Suínos/microbiologiaRESUMO
Infections caused by Staphylococcus aureus are increasingly prevalent, and treatment has become more difficult due to the emergence of strains that are resistant to multiple drugs, such as methicillin-resistant Staphylococcus aureus (MRSA). Penicillin-binding proteins (PBPs) are essential enzymes in peptidoglycan biosynthesis. Only found in bacteria, they are an excellent target for the development of bacterial control strategies. S. aureus has 4 PBPs, and only PBP2 has transglycosylation activity, making it a good model to evaluate whether the inactivation of the transglycosylase domain (PBP2t) could lead to bacterial death. (His6)-tagged PBP2t was purified from the E. coli cell lysate using Ni-charged resin, and ELISA and immunoblotting assays demonstrated that PBP2t is immunogenic. Flow cytometry analysis was performed to verify the binding of polyclonal antibodies to the bacterial cell surface. In order to verify the ability to provide protection, immunized mice were challenged with a sublethal dose of MRSA, and the bacterial loads in kidneys and spleen were evaluated. A reduction of 2-2.5 logs was seen in organs from immunized mice compared with the negative controls in two independent assays (p < 0.01). Our results demonstrate that the PBP2t is a promising target for the development of novel antimicrobial strategies, but further testing should be performed to validate the protection conferred by immunization with this protein.
Assuntos
Staphylococcus aureus Resistente à Meticilina , Staphylococcus aureus , Animais , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Proteínas de Bactérias , Escherichia coli , Imunoterapia , Camundongos , Proteínas de Ligação às Penicilinas/genéticaRESUMO
Oxacillin-susceptible mecA-positive Staphylococcus aureus (OS-MRSA) isolates have been described worldwide, but data regarding dogs and their owners have not been reported. This study investigated the occurrence of OS-MRSA and MRSA isolates in the nasal mucosa of 241 healthy dogs and 208 owners in the community. S. aureus isolates were characterized by susceptibility testing, detection of the mecA and the Panton-Valentine leukocidin (PVL) genes, staphylococcal chromosome cassette (SCC)mec typing and rep-PCR-RW3A. We report an unprecedented detection of nasal carriage of OS-MRSA in 5.3 % (2/38) of healthy dogs and 1.75 % (1/57) of their owners. We also found MRSA in 2.6 % (1/38) of the dogs and 3.5 % (2/57) of the owners. Only the human isolate was SCCmec IV and PVL-positive. Molecular typing revealed that the same cluster of S. aureus was present in owners and dogs from the same or different families attended at the same veterinary clinic. The three OS-MRSA isolates did not show genetic similarity to each other. Detection of OS-MRSA in this context alerts us to the role of dogs and owners as possible silent reservoirs of this microorganism in the community, which may potentially be misidentified as methicillin-sensitive S. aureus (MSSA) in the laboratory routine, representing an additional threat in antimicrobial therapy for staphylococcal infections.
Assuntos
Doenças do Cão , Infecções Estafilocócicas , Staphylococcus aureus/isolamento & purificação , Animais , Proteínas de Bactérias/genética , Brasil/epidemiologia , Doenças do Cão/epidemiologia , Doenças do Cão/microbiologia , Cães , Humanos , Testes de Sensibilidade Microbiana/veterinária , Oxacilina/farmacologia , Proteínas de Ligação às Penicilinas/genética , Infecções Estafilocócicas/epidemiologia , Infecções Estafilocócicas/veterinária , Staphylococcus aureus/genéticaRESUMO
Aim: Genomic analysis of a methicillin-resistant Staphylococcus aureus (MRSA) strain cultured from a non-migratory seabird at Fernando de Noronha Archipelago (Brazilian oceanic islands) was carried out to investigate the potential origin of MRSA genetic determinants in an ecological setting with minimal or absent antimicrobial selective pressure, and minimal interaction with humans and domestic animals. Results: The study determined mecA gene homology and the phylogenetic relatedness with mecA described in Staphylococcus sciuri, which was the major Staphylococcus spp. cultured from the birds. Our findings corroborate in silico assumptions that the mecA gene in MRSA strains clinically relevant for humans and animals originates from S. sciuri ancestors. Conclusion: Coagulase-negative staphylococci seem to be natural reservoirs of methicillin-resistant genes to S. aureus, even in environments with very low antimicrobial selection pressure.
Assuntos
Proteínas de Bactérias/genética , Aves/microbiologia , Staphylococcus aureus Resistente à Meticilina/genética , Proteínas de Ligação às Penicilinas/genética , Staphylococcus/genética , Animais , Brasil , Ilhas , Sequenciamento Completo do GenomaRESUMO
Staphylococcus aureus is a versatile and highly adaptable pathogen associated with a wide range of infectious diseases in humans and animals. In the last decades, concern has increased worldwide due to the emergence and spread of methicillin-resistant S. aureus (MRSA) strains shortly after this drug became a therapeutic option. In this study, we report the genomic features of the first mecC-mediated, ß-lactam resistant MRSA strain associated with livestock in Brazil and in the American continent. Three clonally related phenotypic MRSA isolates originated from a dairy herd were confirmed by polymerase chain reaction as mecC-harbouring MRSA isolates. Whole-genome sequencing was performed by Illumina Miseq platform. Downstream analyses showed that the strain was identified as the sequence type 126 (ST126) and spa type t605. In silico analysis revealed a mecC homolog gene in the orfX region associated with different penicillin-binding proteins. Moreover, genes encoding for efflux pump systems (arlR, mepR, LmrS, norA and mgrA), and antibiotic inactivation enzymes (blaZ and FosB) were also detected. Virulence analyses revealed that the strain harbours genes encoding for exoenzymes (aur, splA, splB and splE), toxin (hlgA, hlgB, hlgC, lukD and lukE) and enterotoxin (sea). The epidemiologic and genomic information provided by this study will support further epidemiological and evolutionary investigations to understand the origin and dissemination of mecC-MRSA among animals and its impact on public health.
Assuntos
Doenças dos Bovinos , Staphylococcus aureus Resistente à Meticilina , Infecções Estafilocócicas/veterinária , Animais , Brasil/epidemiologia , Bovinos , Doenças dos Bovinos/diagnóstico , Doenças dos Bovinos/epidemiologia , Feminino , Staphylococcus aureus Resistente à Meticilina/genética , Staphylococcus aureus Resistente à Meticilina/isolamento & purificação , Staphylococcus aureus Resistente à Meticilina/patogenicidade , Proteínas de Ligação às Penicilinas/genética , Infecções Estafilocócicas/diagnóstico , Infecções Estafilocócicas/epidemiologia , Virulência/genéticaRESUMO
Context: Nosocomial infections arise from many microorganisms, including Staphylococcus aureus. Aims: The aim of this study is to determine the molecular epidemiology of circulating methicillin-resistant S. aureus (MRSA) clones among patients attending community and health-care facilities in Nova Friburgo, RJ, Brazil. Methods: A total of 1002 nasal swab samples were collected from May 2010 to September 2015. S. aureus isolates were identified through phenotypic tests, submitted to antimicrobial susceptibility tests and genotypic analysis to detect mecA, panton-valentine leucocidin (PVL) genes, SCCmec, SPA and multilocus sequencing typing (MLST) typing. Results: We identified 294 (29.3%) isolates as S. aureus and 91 (9.1%) as MRSA. A total of 17 isolates did not present a correlation between phenotypic and genotypic resistance profiles. Among MRSA isolates, 17 (18.7%) carried PVL genes. A total of 20 different SPA types were determined, being grouped by MLST into eight different sequence types. ST5/t002 was the most prevalent genotype found among these isolates. Conclusions: There is a gradual colonisation shift happening in the infection pattern by S. aureus in Brazil. The Brazilian Epidemic Clone (ST239-SCCmec IIIa-PVL-) seems to be substituted by isolates from different clonal complexes, such as ST5, ST8 and ST30. The non-correlation between phenotypic/genotypic resistance profile observed in some isolates suggests the presence of other methicillin resistance mechanisms different from mecA presence or a difference in the nucleotide sequence, which prevents the primers to identify the specific region during polymerase chain reaction reactions. MRSA identification should be based on phenotypic and genotypic testing to ensure the various types of resistance mechanisms.
Assuntos
Antibacterianos/farmacologia , Portador Sadio/epidemiologia , Staphylococcus aureus Resistente à Meticilina/isolamento & purificação , Nariz/microbiologia , Oxacilina/farmacologia , Infecções Estafilocócicas/epidemiologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Proteínas de Bactérias/genética , Portador Sadio/microbiologia , Infecção Hospitalar/epidemiologia , Infecção Hospitalar/microbiologia , Estudos Transversais , Testes de Sensibilidade a Antimicrobianos por Disco-Difusão , Farmacorresistência Bacteriana Múltipla , Genes Bacterianos , Variação Genética , Instalações de Saúde , Humanos , Staphylococcus aureus Resistente à Meticilina/classificação , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Staphylococcus aureus Resistente à Meticilina/genética , Pessoa de Meia-Idade , Epidemiologia Molecular , Tipagem Molecular , Tipagem de Sequências Multilocus , Proteínas de Ligação às Penicilinas/genética , Prevalência , Infecções Estafilocócicas/microbiologia , Proteína Estafilocócica A/genética , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/isolamento & purificação , Adulto JovemRESUMO
INTRODUCTION: Staphylococci are the most important agents associated with bovine mastitis. This study aimed at characterizing resistance factors to antimicrobials in Staphylococcus spp. isolated from the milk of cows with subclinical mastitis. METHODOLOGY: In vitro resistance of 243 Staphylococcus spp. isolates to antimicrobials commonly used in clinical practice was evaluated. The detection and expression of genes encoding resistance mecA (gene encoding penicillin binding protein 2a) mecALGA251 (mecA homologue), blaZ (gene encoding penicillin resistance), femA and femB (genes encoding essential factors - A and B - for the expression of methicillin resistance) and aacA-aphD (gene encoding for a bifunctional enzyme that confers resistance to gentamicin) using PCR and RT-PCR was investigated. RESULTS: One or more genes encoding resistance to different antimicrobials were detected in 184 Staphylococcus spp. SAMPLES: The femA and femB genes were the most frequent. Regarding the variables' detection (N = number of strains) and expression (% of strains), the following results were obtained: blaZ (N = 40 - 82.5%), femA (N = 147 - 47.6%), aacAaphD (N = 30 - 43.3%), femB (N = 138 - 29.7%), mecA (N = 33 - 27.3%), mecALGA251 (N = 01 - 0.0%). There was a higher occurrence of phenotypic resistant strains for amoxicillin, ampicillin and penicillin in isolates positive for detection and/or expression of blaZ gene when compared with the other genes. CONCLUSIONS: The present study provides new information on genotypic traits of Staphylococcus isolates from bovine subclinical mastitis especially regarding the evaluation of expression of genes associated with antimicrobial resistance in Staphylococcus spp. using molecular tools.
Assuntos
Farmacorresistência Bacteriana/genética , Mastite Bovina/microbiologia , Leite/microbiologia , Infecções Estafilocócicas/veterinária , Staphylococcus/efeitos dos fármacos , Staphylococcus/genética , Animais , Antibacterianos/farmacologia , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Brasil , Bovinos , DNA Bacteriano , Feminino , Resistência a Meticilina , Testes de Sensibilidade Microbiana , Resistência às Penicilinas , Proteínas de Ligação às Penicilinas/genética , Proteínas de Ligação às Penicilinas/metabolismo , Infecções Estafilocócicas/tratamento farmacológico , Infecções Estafilocócicas/microbiologia , Staphylococcus/isolamento & purificaçãoRESUMO
ABSTRACT The increasing rates of nosocomial infection associated with coagulase-negative staphylococci (CoNS) were the rationale for this study, aiming to categorize oxacillin-resistant CoNS species recovered from blood culture specimens of inpatients at the UNESP Hospital das Clínicas in Botucatu, Brazil, over a 20-year period, and determine their sensitivity to other antimicrobial agents. The mecA gene was detected in 222 (74%) CoNS samples, and the four types of staphylococcal chromosomal cassette mec (SCCmec) were characterized in 19.4%, 3.6%, 54.5%, and 14.4% of specimens, respectively, for types I, II, III, and IV. Minimal inhibitory concentration (MIC) values to inhibit 50% (MIC50) and 90% (MIC90) of specimens were, respectively, 2 and >256 µL/mL for oxacillin, 1.5 and 2 µL/mL for vancomycin, 0.25 and 0.5 µL/mL for linezolid, 0.094 and 0.19 µL/mL for daptomycin, 0.19 and 0.5 µL/mL for quinupristin/dalfopristin, and 0.125 and 0.38 µL/mL for tigecycline. Resistance to oxacillin and tigecycline and intermediate resistance to quinupristin/dalfopristin were observed. Eight (2.7%) of all 300 CoNS specimens studied showed reduced susceptibility to vancomycin. Results from this study show high resistance rates of CoNS to antimicrobial agents, reflecting the necessity of using these drugs judiciously and controlling nosocomial dissemination of these pathogens.
Assuntos
Humanos , Infecções Estafilocócicas/microbiologia , Staphylococcus/efeitos dos fármacos , Coagulase/metabolismo , Farmacorresistência Bacteriana/genética , Antibacterianos/farmacologia , Staphylococcus/genética , Staphylococcus/química , Proteínas de Bactérias/genética , Testes de Sensibilidade Microbiana , Reação em Cadeia da Polimerase , Farmacorresistência Bacteriana/efeitos dos fármacos , Proteínas de Ligação às Penicilinas/genética , Genes Bacterianos/genética , Hospitais de EnsinoRESUMO
The increasing rates of nosocomial infection associated with coagulase-negative staphylococci (CoNS) were the rationale for this study, aiming to categorize oxacillin-resistant CoNS species recovered from blood culture specimens of inpatients at the UNESP Hospital das Clínicas in Botucatu, Brazil, over a 20-year period, and determine their sensitivity to other antimicrobial agents. The mecA gene was detected in 222 (74%) CoNS samples, and the four types of staphylococcal chromosomal cassette mec (SCCmec) were characterized in 19.4%, 3.6%, 54.5%, and 14.4% of specimens, respectively, for types I, II, III, and IV. Minimal inhibitory concentration (MIC) values to inhibit 50% (MIC50) and 90% (MIC90) of specimens were, respectively, 2 and >256µL/mL for oxacillin, 1.5 and 2µL/mL for vancomycin, 0.25 and 0.5µL/mL for linezolid, 0.094 and 0.19µL/mL for daptomycin, 0.19 and 0.5µL/mL for quinupristin/dalfopristin, and 0.125 and 0.38µL/mL for tigecycline. Resistance to oxacillin and tigecycline and intermediate resistance to quinupristin/dalfopristin were observed. Eight (2.7%) of all 300 CoNS specimens studied showed reduced susceptibility to vancomycin. Results from this study show high resistance rates of CoNS to antimicrobial agents, reflecting the necessity of using these drugs judiciously and controlling nosocomial dissemination of these pathogens.