Mammaliicoccus sciuri's Pan-Immune System and the Dynamics of Horizontal Gene Transfer Among Staphylococcaceae: a One-Health CRISPR Tale.

Recently emancipated from the Staphylococcus genus due to genomic differences, Mammaliicoccus sciuri, previously classified as an occasional pathogen, emerges as a significant player in the landscape of resistance gene dissemination among Staphylococcaceae. Despite its classification, its role remained enigmatic. In this study, we delved into the genomic repertoire of M. sciuri to unravel its contribution to resistance and virulence gene transfer in the context of One Health. Through comprehensive analysis of publicly available genomes, we unveiled a diverse pan-immune system adept at defending against exogenous genetic elements, yet concurrently fostering horizontal gene transfer (HGT). Specifically, exploration of CRISPR-Cas systems, with spacer sequences as molecular signatures, elucidated a global dissemination pattern spanning environmental, animal, and human hosts. Notably, we identified the integration of CRISPR-Cas systems within SCCmecs (Staphylococcal Cassette Chromosome mec), harboring key genes associated with pathogenicity and resistance, especially the methicillin resistance gene mecA, suggesting a strategic adaptation to outcompete other mobile genetic elements. Our findings underscored M. sciuri's active engagement in HGT dynamics and evolutionary trajectories within Staphylococcaceae, emphasizing its central role in shaping microbial communities and highlighting the significance of understanding its implications in the One Health framework, an interdisciplinary approach that recognizes the interconnectedness of human, animal, and environmental health to address global health challenges.

Co-resistance to methicillin and clindamycin among coagulase-negative staphylococci isolates recovered from pet food in Brazil.

Pet food have been considered as possible vehicles of bacterial pathogens. The sudden boom of the pet food industry due to the worldwide increase in companion animal ownership calls for pet food investigations. Herein, this study aimed to determine the frequency, antimicrobial susceptibility profile, and molecular characteristics of coagulase-negative staphylococci (CoNS) in different pet food brands in Brazil. Eighty-six pet food packages were screened for CoNS. All isolates were identified at species level by MALDI-TOF MS and species-specific PCR. Antimicrobial susceptibility testing was performed by disc diffusion and broth microdilution (vancomycin and teicoplanin only) methods. The D-test was used to screen for inducible clindamycin phenotype (MLS-B). SCCmec typing and detection of mecA, vanA, vanB, and virulence-encoding genes were done by PCR. A total of 16 (18.6 %) CoNS isolates were recovered from pet food samples. Isolates were generally multidrug-resistant (MDR). All isolates were completely resistant (100 %) to penicillin. Resistances (12.5 % - 75 %) were also observed for fluoroquinolones, sulfamethoxazole-trimethoprim, tetracycline, rifampicin, erythromycin, and tobramycin. Isolates were susceptible to vancomycin (MICs <0.25-1 µg/mL) and teicoplanin (MICs <0.25-4 µg/mL). Intriguingly, 3/8 (37.5 %) CoNS isolates with the ERYRCLIS antibiotype expressed MLS-B phenotype. All isolates harboured blaZ gene. Seven (43.8 %) isolates carried mecA; and among them, the SCCmec Type III was the most frequent (n = 5/7; 71.4 %). Isolates also harboured seb, see, seg, sej, sem, etb, tsst, pvl, and hla toxin virulence-encoding genes (6.3 % - 25 %). A total of 12/16 (75 %) isolates were biofilm producers, while the icaAB gene was detected in an S. pasteuri isolate. Herein, it is shown that pet food is a potential source of clinically important Gram-positive bacterial pathogens. To the best of our knowledge, this is the first report of MLS-B phenotype and MR-CoNS in pet food in Latin America.

Staphylococcus capitis Bloodstream Isolates: Investigation of Clonal Relationship, Resistance Profile, Virulence and Biofilm Formation.

Staphylococcus capitis has been recognized as a relevant opportunistic pathogen, particularly its persistence in neonatal ICUs around the world. Therefore, the aim of this study was to describe the epidemiological profile of clinical isolates of S. capitis and to characterize the factors involved in the persistence and pathogenesis of these strains isolated from blood cultures collected in a hospital in the interior of the state of São Paulo, Brazil. A total of 141 S. capitis strains were submitted to detection of the mecA gene and SCCmec typing by multiplex PCR. Genes involved in biofilm production and genes encoding enterotoxins and hemolysins were detected by conventional PCR. Biofilm formation was evaluated by the polystyrene plate adherence test and phenotypic resistance was investigated by the disk diffusion method. Finally, pulsed-field gel electrophoresis (PFGE) was used to analyze the clonal relationship between isolates. The mecA gene was detected in 99 (70.2%) isolates, with this percentage reaching 100% in the neonatal ICU. SCCmec type III was the most prevalent type, detected in 31 (31.3%) isolates and co-occurrence of SCCmec was also observed. In vitro biofilm formation was detected in 46 (32.6%) isolates but was not correlated with the presence of the ica operon genes. Furthermore, biofilm production in ICU isolates was favored by hyperosmotic conditions, which are common in ICUs because of the frequent parenteral nutrition. Analysis of the clonal relationship between the isolates investigated in the present study confirms a homogeneous profile of S. capitis and the persistence of clones that are prevalent in the neonatal ICU and disseminated across the hospital. This study highlights the adaptation of isolates to specific hospital environments and their high clonality.

Microbial Complexity of Oral Cavity of Healthy Dogs Identified by Mass Spectrometry and Next-Generation Sequencing.

The high complexity of the oral microbiota of healthy dogs and the close exposure of humans to companion animals represent a risk of the transmission of potential zoonotic microorganisms to humans, especially through dog bites, including multidrug-resistant ones. Nonetheless, a limited number of comprehensive studies have focused on the diversity of the microorganisms that inhabit the oral cavities of healthy dogs, particularly based on modern molecular techniques. We investigated bacterial and fungal organisms in the oral cavities of 100 healthy dogs based on a combination of conventional and selective microbiological culture, mass spectrometry (MALDI-TOF MS), and next-generation sequencing. In addition, in vitro antimicrobial susceptibility patterns of isolates and mecA resistance gene were assessed. A total of 213 bacteria and 20 fungi were isolated. Staphylococcus pseudintermedius (40/100 = 40%), α-hemolytic Streptococcus (37/100 = 37%), and Pasteurella stomatis (22/100 = 22%) were the most prevalent bacteria diagnosed by microbiological culture and MALDI-TOF MS, whereas Aspergillus (10/100 = 10%) was the most common fungi identified. Based on next-generation sequencing of selected 20 sampled dogs, Porphyromonas (32.5%), Moraxella (16.3%), Fusobacterium (12.8%), Conchiformibius (9.5%), Bergeyella (5%), Campylobacter (3.8%), and Capnocytophaga (3.4%) genera were prevalent. A high multidrug resistance rate was observed in Staphylococcus pseudintermedius isolates, particularly to azithromycin (19/19 = 100%), penicillin (15/19 = 78.9%), and sulfamethoxazole/trimethoprim (15/19 = 78.9%). In addition, the mecA resistance gene was detected in 6.1% (3/49) of coagulase-positive staphylococci. Here, we highlight the microbial complexity of the oral mucosa of healthy dogs, including potential zoonotic microorganisms and multidrug-resistant bacteria, contributing with the investigation of the microbiota and antimicrobial resistance patterns of the microorganisms that inhabit the oral cavity of healthy dogs.

Beta-Lactam Antibiotic Resistance Genes in the Microbiome of the Public Transport System of Quito, Ecuador.

Multidrug-resistant bacteria present resistance mechanisms against ß-lactam antibiotics, such as Extended-Spectrum Beta-lactamases (ESBL) and Metallo-ß-lactamases enzymes (MBLs) which are operon encoded in Gram-negative species. Likewise, Gram-positive bacteria have evolved other mechanisms through mec genes, which encode modified penicillin-binding proteins (PBP2). This study aimed to determine the presence and spread of ß-lactam antibiotic resistance genes and the microbiome circulating in Quito's Public Transport (QTP). A total of 29 station turnstiles were swabbed to extract the surface environmental DNA. PCRs were performed to detect the presence of 13 antibiotic resistance genes and to identify and to amplify 16S rDNA for barcoding, followed by clone analysis, Sanger sequencing, and BLAST search. ESBL genes blaTEM-1 and blaCTX-M-1 and MBL genes blaOXA-181 and mecA were detected along QPT stations, blaTEM being the most widely spread. Two subvariants were found for blaTEM-1, blaCTX-M-1, and blaOXA-181. Almost half of the circulating bacteria found at QPT stations were common human microbiota species, including those classified by the WHO as pathogens of critical and high-priority surveillance. ß-lactam antibiotic resistance genes are prevalent throughout QPT. This is the first report of blaOXA-181 in environmental samples in Ecuador. Moreover, we detected a new putative variant of this gene. Some commensal coagulase-negative bacteria may have a role as mecA resistance reservoirs.

Genomic analysis of a multidrug methicillin-resistant staphylococcus epidermidis recovered from the urine of a guinea pig (Cavia porcellus) with suspected pyelonephritis.

Staphylococcus epidermidis is an emerging pathogen causing infant pyelonephritis. There is a lack of genomic data on Staphylococcus epidermidis as the etiology of pyelonephritis and its resistant determinants. In this study, we have conducted a genomic and microbiologic investigation of an S. epidermidis recovered from the urine of a guinea pig with suspected pyelonephritis in Brazil. The genome was sequenced using the Illumina MiSeq platform and de novo assembled using SPades. Resistome, virulome, and plasmidome were in silico predicted using bioinformatics tools. Data analysis revealed that S. epidermidis USP-LZB-G06 belonged to sequence type ST332. Strikingly, a broad resistome (antibiotics, hazardous heavy metals, and biocides) was predicted, including the presence of the clinically relevant mecA, blaZ, and qacA efflux pump genes. SNP-based analysis revealed that strain USP-LZB-G06 was clustered along mecA positive S. epidermidis strains of ST332 isolated between 2008 and 2016 from humans in Australia and the United States of America. Our results indicate that the detection of this microorganism should be considered as a urinary tract infection agent in exotic pets, particularly guinea pigs. In addition, there is an urgent need to update veterinarians regarding the detection and therapeutic management of these microorganisms.

Resistance profile of bacterial isolates from different llama microbiotas (Lama glama Linnaeus 1758) / Perfil de resistência de isolados bacterianos de diferentes microbiotas de lhamas (Lama glama Linnaeus 1758)

Este trabalho objetivou o isolamento e a identificação das bactérias de diferentes microbiotas de lhamas. Para tanto, testes de disco difusão e diagnósticos moleculares para pesquisa de genes de resistência foram realizados. Foram isolados cinco Staphylococcus spp. coagulase positiva e 19 Staphylococcus spp. coagulase negativa, 19 Escherichia coli¸ duas Pantoea agglomerans, uma Koserella trabulsii, uma Enterobacter aerogenes e uma Klebsiella Pneumoniae. Entre os isolados de Staphylococcus spp., 79,17% foram resistentes ao sulfazotrim, 45,83% resistentes à penicilina e 20,83% à ampicilina. Foi confirmada a presença do gene mecA em apenas um isolado oxacilina resistente. No teste de disco difusão, 58,3% das enterobactérias foram resistentes à amoxicilina + ácido clavulânico e à cefotaxima, 50% à ceftazidima e ceftriaxona, e 33,3% à amoxicilina. Ainda entre os isolados Gram-negativos, não houve a expressão fenotípica de isolados ESBL. O presente trabalho expôs a presença de microrganismos resistentes a antibióticos em lhamas que não tiveram contato prévio com essas drogas, além da presença do gene mecA em um dos animais. O conhecimento da microbiota bacteriana de diferentes espécies animais tem se tornado cada vez mais importante. Tal relevância se deve à possibilidade de esses microrganismos serem compartilhados entre os animais, os humanos e até mesmo o meio ambiente.

Genomic Analysis of a Community-Acquired Methicillin-Resistant Staphylococcus aureus Sequence Type 1 Associated with Caprine Mastitis.

This study aimed to investigate the genomic and epidemiological features of a methicillin-resistant Staphylococcus aureus sequence type 1 (MRSA ST1) strain associated with caprine subclinical mastitis. An S. aureus strain was isolated from goat's milk with subclinical mastitis in Paraiba, Northeastern Brazil, by means of aseptic procedures and tested for antimicrobial susceptibility using the disk-diffusion method. Whole genome sequencing was performed using the Illumina MiSeq platform. After genome assembly and annotation, in silico analyses, including multilocus sequence typing (MLST), antimicrobial resistance and stress-response genes, virulence factors, and plasmids detection were performed. A comparative SNP-based phylogenetic analysis was performed using publicly available MRSA genomes. The strain showed phenotypic resistance to cefoxitin, penicillin, and tetracycline and was identified as sequence type 1 (ST1) and spa type 128 (t128). It harbored the SCCmec type IVa (2B), as well as the lukF-PV and lukS-PV genes. The strain was phylogenetically related to six community-acquired MRSA isolates (CA-MRSA) strains associated with human clinical disease in North America, Europe, and Australia. This is the first report of a CA-MRSA strain associated with milk in the Americas. The structural and epidemiologic features reported in the MRSA ST1 carrying a mecA-SCCmec type IVa suggest highly complex mechanisms of horizontal gene transfer in MRSA. The SNP-based phylogenetic analysis suggests a zooanthroponotic transmission, i.e., a strain of human origin.

Phenotypic and genotypic characterization of oxacillin-susceptible and mecA positive Staphylococcus aureus strains isolated in Uruguay / Caracterización fenotípica y genotípica de cepas de Staphylococcus aureus sensibles a oxacilina y mecA positivas aisladas en Uruguay

Abstract The aim of this study was to characterize phenotypically and genotypically 27 mecApositive Staphylococcus aureus strains with oxacillin MICs of ≤2 g/ml by Vitek 2, isolated indifferent regions of Uruguay. Susceptibility to oxacillin and cefoxitin was studied by gradient dif-fusion, disk diffusion to cefoxitin, and Phoenix and MicroScan systems. PBP2a was determined.SCCmec typing was performed and the isolates were compared by PFGE. Twenty-six isolateswere susceptible to oxacillin; one strain was susceptible to cefoxitin by disk diffusion and 3strains by gradient diffusion. Phoenix and MicroScan panels detected methicillin resistance in25 and 27 strains, respectively. Twenty-six strains tested positive for PBP2a. Twenty-six strainscarried SCCmec V and 24 belonged to pulsotype A. One strain carried SCCmec IV and did notbelong to pulsotype A. Cefoxitin disk diffusion test and PBP2a detection correctly identified 26of these 27 strains as MRSA. PFGE results suggest the dissemination of a cluster of MRSA carryingSCCmec V.

Resumen El objetivo de este estudio fue caracterizar fenotípicamente y genotípicamente 27 cepas de Staphylococcus aureus positivas para mecA y con CIM de oxacilina <2 pg/ml según Vitek 2, obtenidas en diferentes regiones del país. La sensibilidad frente a la oxacilina y la cefoxitina se estudió por difusión en gradiente, por disco-difusión (cefoxitina) y por los sistemas Phoenix y MicroScan. Se analizó la portación de PBP2a, se realizó la tipificación de SCCmec y las cepas se compararon mediante PFGE. Resultaron sensibles a oxacilina por difusión en gradiente 26 cepas; una fue sensible a cefoxitina por disco-difusión y 3 lo fueron por difusión en gradiente. Los sistemas Phoenix y MicroScan detectaron resistencia a meticilina en 25 y 27 cepas, respectivamente. Asimismo, 26 cepas portaban PBP2a y 26 cepas mostraron presencia de SCCmec V, 24 correspondieron al pulsotipo A. Una portaba SCCmec IV y no perteneció al pulsotipo A. La prueba de disco-difusión con cefoxitina y la detección de PBP2a identificaron 26 de 27 cepas como MRSA. La PFGE sugiere la diseminación de un grupo MRSA con SCCmec V. © 2022 Asociación Argentina de Microbiología. Publicado por Elsevier Espana, S.L.U. Este es un artículo Open Access bajo la licencia CC BY-NC-ND (https://creativecommons.org/licenses/by-nc-nd/4.0/).

Phenotypic and genotypic characterization of oxacillin-susceptible and mecA positive Staphylococcus aureus strains isolated in Uruguay.

The aim of this study was to characterize phenotypically and genotypically 27 mecA positive Staphylococcus aureus strains with oxacillin MICs of ≤2µg/ml by Vitek 2, isolated in different regions of Uruguay. Susceptibility to oxacillin and cefoxitin was studied by gradient diffusion, disk diffusion to cefoxitin, and Phoenix and MicroScan systems. PBP2a was determined. SCCmec typing was performed and the isolates were compared by PFGE. Twenty-six isolates were susceptible to oxacillin; one strain was susceptible to cefoxitin by disk diffusion and 3 strains by gradient diffusion. Phoenix and MicroScan panels detected methicillin resistance in 25 and 27 strains, respectively. Twenty-six strains tested positive for PBP2a. Twenty-six strains carried SCCmec V and 24 belonged to pulsotype A. One strain carried SCCmec IV and did not belong to pulsotype A. Cefoxitin disk diffusion test and PBP2a detection correctly identified 26 of these 27 strains as MRSA. PFGE results suggest the dissemination of a cluster of MRSA carrying SCCmec V.

Resistencia de cepas de Staphylococcus aureus aislados en ambientes nosocomiales / Resistance of Staphylococcus aureus strains isolated in nosocomial environments / Resistência de cepas de Staphylococcus aureus isoladas em ambientes nosocomiais

Staphylococcus aureus es un microorganismo que posee características particulares de virulencia y resistencia a los antibióticos. Las infecciones que produce, ocurren mayormente en personas inmunodeprimidas que podrían presentar severas consecuencias, a pesar de la terapia antimicrobiana. Objetivo. Determinar la resistencia de Staphylococcus aureus aislados en ambientes nosocomiales mediante métodos convencionales y moleculares. Materiales y métodos. el estudio presenta un enfoque cuantitativo, investigación de campo, observacional de corte transversal. Se analizó 200 muestras de aislados de ambientes nosocomiales mediante métodos fenotípicos (antibiograma por la técnica de Kirby Bauer) y genotípicos (genes de resistencia blaZ, mecA, y vanA por PCR punto final). Resultados. Los resultados de las pruebas de susceptibilidad antibiótica mostraron que el 100% de las cepas de S. aureus aisladas, fueron resistentes a oxacilina y penicilina g; y sensibles a vancomicina. Conclusiones. los resultados obtenidos en este estudio evidenciaron la presencia de SARM en las diferentes áreas hospitalarias, constituyéndose en un factor de riesgo de transmisión horizontal entre el personal sanitario y los pacientes, razón por la cual es de gran importancia evaluar su prevalencia y establecer medidas rigurosas de prevención y control de su diseminación, para disminuir el riesgo de nuevas infecciones.

Staphylococcus aureus is a microorganism with particular characteristics of virulence and resistance to antibiotics. The infections it produces occur mostly in immunocompromised individuals who may present severe consequences, despite antimicrobial therapy. Objective. To determine the resistance of Staphylococcus aureus isolated in nosocomial environments by conventional and molecular methods. Materials and methods. The study presents a quantitative, field research, observational, cross-sectional approach. Two hundred samples of isolates from nosocomial environments were analyzed by phenotypic (antibiogram by Kirby Bauer technique) and genotypic (resistance genes blaZ, mecA, and vanA by endpoint PCR) methods. Results. The results of the antibiotic susceptibility tests showed that 100% of the S. aureus strains isolated were resistant to oxacillin and penicillin g; and sensitive to vancomycin. Conclusions. the results obtained in this study showed the presence of MRSA in the different hospital areas, constituting a risk factor for horizontal transmission between health personnel and patients, which is why it is of great importance to evaluate its prevalence and establish rigorous measures for the prevention and control of its dissemination, in order to reduce the risk of new infections.

O Staphylococcus aureus é um microorganismo com características particulares de virulência e resistência a antibióticos. As infecções ocorrem principalmente em indivíduos imunocomprometidos que podem ter conseqüências graves, apesar da terapia antimicrobiana. Objetivo. Para determinar a resistência de Staphylococcus aureus isolado em ambientes nosocomiais usando métodos convencionais e moleculares. Materiais e métodos. O estudo apresenta uma abordagem quantitativa, pesquisa de campo, observacional, transversal. Duzentas amostras de isolados de ambientes nosocomiais foram analisadas pelos métodos fenotípico (antibiograma pela técnica de Kirby Bauer) e genotípico (genes de resistência blaZ, mecA, e vanA pelo método de PCR de ponto final). Resultados. Os resultados dos testes de suscetibilidade aos antibióticos mostraram que 100% das cepas de S. aureus isoladas eram resistentes à oxacilina e penicilina g; e sensíveis à vancomicina. Conclusões. Os resultados obtidos neste estudo mostraram a presença de MRSA nas diferentes áreas hospitalares, constituindo um fator de risco para a transmissão horizontal entre o pessoal de saúde e os pacientes, razão pela qual é de grande importância avaliar sua prevalência e estabelecer medidas rigorosas para a prevenção e controle de sua disseminação, a fim de reduzir o risco de novas infecções.

Vigilancia epidemiológica de Staphylococcus aureus y resistencia antibiótica en ambientes nosocomiales / Epidemiological surveillance of Staphylococcus aureus and the antibiotic resistence in nosocomial environments / Vigilância epidemiológica de Staphylococcus aureus e resistência a antibióticos em ambientes nosocomiais

Uno de los microrganismos más importantes en Infecciones Asociadas a la Atención en Salud (IAAS) es Staphylococcus aureus, una bacteria aerobia Gram positiva, resistente a diferentes condiciones ambientales. Objetivo. Identificar Staphylococcus aureus y su resistencia a los principales antibióticos Betalactámicos, aislada en áreas inertes. Materiales y métodos. Se realizó análisis fenotípico, antibiograma y métodos moleculares como: Extracción de ADN mediante Lisis Alcalina, identificación molecular para la amplificación de los genes tanto de identificación de la bacteria (nucA y femB), como de resistencia de antibióticos (blaZ, mecA y vanA) mediante PCR punto final, la separación de los amplicones se realizó mediante electroforesis en gel de Agarosa, los productos de la PCR se revelaron mediante la utilización de transiluminador UV. Resultados. De 200 muestras tomadas se obtuvo dos muestras positivas (1%) para Staphylococcus aureus, con el 100% de resistencia a penicilina y sensible a todos los demás antibióticos testeados. Conclusiones. La identificación de la bacteria y su resistencia hoy en día se realiza mayormente mediante métodos moleculares, lo cual no descarta la identificación fenotípica que, en este caso, determinó resultados importantes como lo es la prueba D-test positivo. La resistencia a fármacos betalactámicos se considera como un serio problema de salud, por lo tanto, se requiere de una vigilancia epidemiológica constante.

Staphylococcus aureus is one of the most important microorganisms related to Health Care Associated Infections (HCAI), it is a Gram-positive aerobic bacterium, highly resistant to the outer environment. Objective. Identify Staphylococcus aureus and its resistance to the most common beta-lactam antibiotics in isolated, inert areas. Materials and methods. Phenotypic analysis, antibiogram and molecular testing such as: DNA extraction by Alkaline Lysis, molecular identification for the amplification of genes both for identification of Staphylococcus aureus (nucA and femB), and for antibiotic resistance (blah, mega and vanA) by PCR, the disassociation of the amplicons was preforming by Agarose gel electrophoresis and results were shown in a UV translluminator. Results. From the 200 samples taken, two showed positive (1%) for Staphylococcus aureus, with 100% penicillin-resistant and sensitive to all other antibiotics tested. Conclusions. Nowadays identification of the bacteria and its resistance is carried out mostly through molecular testing, ruling out phenotypic identification, which in this case it determined important results such as the positive D-test. Resistance to beta-lactam drugs is considered a serious health issue, therefore it requires a safer epidemiological vigilance, an increase in sensitivity testing and the accuracy of results. It is recommended to carry out the D-test in laboratories to identify resistance mechanisms and to guide health personnel to select the best option regarding specific and effective treatment for patients affected with MRSA.

Um dos microrganismos mais importantes em Infecções Associadas à Saúde (HAIs) é o Staphylococcus aureus, uma bactéria aeróbica gram-positiva resistente a diferentes condições ambientais. Objetivo. Identificar Staphylococcus aureus e sua resistência aos principais antibióticos beta-lactam, isolados em áreas inertes. Materiais e métodos. Análise fenotípica, antibiograma e métodos moleculares foram realizados tais como: extração de DNA por Alkaline Lysis, identificação molecular para a amplificação dos genes tanto da identificação da bactéria (nucA e femB), e resistência a antibióticos (blaZ, mecA e vanA) pelo ponto final da PCR, a separação dos amplicons foi realizada por eletrofose em gel de Agarose, os produtos PCR foram revelados usando transiluminador UV. Resultados. De 200 amostras colhidas, duas amostras positivas (1%) foram obtidas para o Staphylococcus aureus, com 100% de resistência à penicilina e sensível a todos os outros antibióticos testados. Conclusões. A identificação da bactéria e sua resistência hoje é realizada principalmente por métodos moleculares, o que não exclui a identificação fenotípica que, neste caso, determinou resultados importantes como o teste D positivo. A resistência aos medicamentos beta-lactam é considerada um grave problema de saúde, portanto, é necessária uma vigilância epidemiológica constante.

Colonización nasal por Staphylococcus aureus y Staphylococcus aureusresistente a meticilina (SARM) en auxiliares de enfermería / Nasal carriage of Staphylococcus aureus and Methicillin-resistant Staphylococcusaureus (MRSA) among nursing assistants

Introducción: Staphylococcus aureus, es la principal causa de bacteriemia infecciosa y endocarditis, así como de infecciones osteoarti-culares, de piel y tejidos blandos, su reservorio principal es la mucosa nasal. Los trabajadores de la salud son una fuente importante de transmisión de S. aureus y S. aureus resistente a la meticilina (SARM). Objetivo: determinar la presencia de Staphylococcus aureus y SARM en la fosa nasal de auxiliares de enfermería en la ciudad de Bogotá. Materiales y métodos: estudio descriptivo de corte transversal, en auxiliares de enfermería de diferentes instituciones hospitalarias y clínicas en la ciudad de Bogotá, Colombia. Se realizó un muestreo aleatorio. El tamaño de la muestra fue de 491 hisopados de la fosa nasal derecha de igual número de auxiliares de enfermería que al momento del estudio se encontraban laborando a nivel clínico. Se tomó un intervalo de confianza del 95% y error máximo admisible del 5%, se consideró el valor de p= 0,5. Se realizó un estudio de frecuencias y determinación de prevalencias mediante un análisis univariado. Resultados: la presente investigación encontró queel 28,5% de los participantes fueron portadores del Staphylococcus aureus y el 6,1% fueron SARM. Conclusiones: la colonización por Staphylococcus aureus y SARM es frecuente en auxiliares de enfermería.

Introduction: Staphylococcus aureus is a human pathogen of clinical severe relevance, it is the leading cause of infectious bacteremia and endocarditis, as well as osteoarticular, skin, and soft tissue infections; its, main reservoir is the nasal mucosa. Healthcare workers are a significant source of transmission of S. aureus and methicillin-resistant S. aureus (MRSA). Objective: To determine the presence of Staphylococcus aureus and MRSA in the nostrils of nursing assistants in Bogotá's city. Materials and methods: a descriptive cross-sec-tional study in nursing assistants from different hospitals and clinical institutions in Bogotá's city, Colombia. Random sampling was carried out. The sample size was 491 swabs from the right nostril from the same number of nursing assistants working at the clinical level at the time of the study. A confidence interval of 95% and maximum permissible error of 5% were taken, the value of p = 0.5 was considered. A study of frequencies and determination of prevalence was carried out through univariate analysis. Results: the present investigation found that 28.5% of the participants were carriers of Staphylococcus aureus, and 6.1% were methicillin-resistant S. aureus(MRSA). Conclusions: colonization by Staphylococcus aureus and methicillin-resistant S. aureus (MRSA) is common in nursing assistants.

mecA positive Staphylococcus spp in bovine mastitis, milkers, milking environment, and the circulation of different MRSA clones at dairy cows farms in the Northeast region of Brazil / Staphylococcus spp mecA positivo em mastite bovina, ordenhadores, ambiente de ordenha e circulação de diferentes clones de MRSA em fazendas de vacas leiteiras na região Nordeste do Brasil

This study detected the presence and distribution of mecA in Staphylococcus spp. in the dairy production environment at farm level in Brazil. We analyzed 335 samples of mastitis cow milk, 15 samples of nostrils and hand swabs from milkers, 14 teat cup swabs, and 9 milking buckets swabs. Initially, the samples were subjected to microbiological analysis to detect Staphylococcus spp. and then S. aureus and mecA positive isolates were identified by PCR. All S. aureus isolates carrying the mecA genes were subjected to DNA macro-restriction analysis by Pulsed-Field Gel Electrophoresis (PFGE). The mecA gene was detected in 6/335 (1.78%) of mastitis cow milk, 5/15 (33.3%), and 5/15 (33.3%) of nostrils and hand swab, and 4/14 (28.5%) of the teat cup isolates. MRSA genotyping was performed by PFGE, a total of seven pulsotypes were grouped in two clusters. This study identified the occurrence and spread of MRSA at dairy environment of farms, and also the existence of distinct genetic profiles between isolates.

Este estudo teve como objetivo detectar a presença e distribuição de mecA em Staphylococcus spp. no ambiente de produção leiteira em fazendas no Brasil. Foram analisadas 335 amostras de leite de vaca com mastite, 15 amostras de swabs de narinas e mãos de ordenhadores, 14 swabs de teteiras e nove swabs de baldes de ordenha. Inicialmente, as amostras foram submetidas a análises microbiológicas para detecção de Staphylococcus spp. e os isolados positivos foram identificados por PCR para S. aureus e mecA. Todos os isolados de S. aureus portadores do gene mecA foram submetidos à análise de macrorrestrição do DNA por Pulsed-Field Gel Electrophoresis (PFGE). O gene mecA foi detectado em 6/335 (1,78%) de leite de vaca com mastite, 5/15 (33,3%) e 5/15 (33,3%) de swab de narinas e de mãos, e 4/14 (28,5%) de teteiras. A genotipagem de MRSA realizada por PFGE identificou um total de sete pulsotipos, que foram agrupados em dois clusters. Este estudo identificou a ocorrência e disseminação de MRSA no ambiente das fazendas leiteiras, e também a existência de perfis genéticos distintos entre os isolados.

Antimicrobial resistance and mecA gene in Staphylococcus spp. isolates from horses used in family farming / Resistência antimicrobiana e gene mecA em Staphylococcus spp. isolados de cavalos usados na agricultura familiar

Horses can contribute to the spread of bacterial diseases, which can be caused mainly by, Staphylococcus spp., which are part of the animals' commensal microbiota, but it is also considered a pathogenic microorganism capable of causing serious infections. vancomycin, when it is resistant to methicillin. Antimicrobial resistance is considered a major health problem by the World Health Organization and the emergence of the mecA gene, responsible for resistance to the class of beta-lactam antibiotics. Thus, the aim of this work was to investigate the antimicrobial resistance profile and the presence of the mecA gene in Staphylococcus spp. isolated from the nasal, oral and auricular microbiota of horses used as animal traction on small family farms. Nasal, oral and auricular swabs were collected from 38 horses, with 29 (76.3%) isolated in nasal swab, 15 (39.4%) in auricular swab and 9 (23.6%) in oral swab, totaling 53 Staphylococcus spp. and 50 (94.33%) samples were resistant to the 11 antimicrobials tested, none of which were positive for molecular tests to identify the mecA gene. The results demonstrate the presence of Staphylococcus spp. associated with high (94.33%) bacterial resistance, indicating that horses can be considered reservoirs of multi-resistant microorganisms.

Os equinos podem contribuir para a disseminação de doenças bacterianas, podendo ser causadas principalmente pelo, Staphylococcus spp., que fazem parte da microbiota comensal dos animais, mas também é considerado microrganismo patogênico capaz de causar infecções graves, em seu tratamento o medicamento mais utilizado é a vancomicina, quando há resistência a meticilina. A resistência antimicrobiana é considerada um dos principais problemas de saúde pela Organização Mundial de Saúde e o surgimento do gene mecA, responsável pela resistência à classe dos antibióticos beta-lactâmicos. Deste modo, o objetivo deste trabalho foi investigar o perfil de resistência antimicrobiana e a presença do gene mecA em Staphylococcus spp. isoladas da microbiota nasal, oral e auricular de cavalos usados como tração animal em pequenas propriedades familiares. Foram coletados swabs nasal, oral e auricular de 38 cavalos, sendo identificados 29 (76,3%) isolados em swab nasal, 15 (39,4%) em swab auricular e 9 (23,6%) em swab oral, totalizando 53 Staphylococcus spp. e 50 (94,33%) amostras foram resistentes aos 11 antimicrobianos testados, nenhuma amostra foi positiva aos testes moleculares para identificação do gene mecA. Os resultados demonstram a presença de Staphylococcus spp. associada à alta (94,33%) resistência bacteriana, indicando que os cavalos podem ser considerados reservatórios de microrganismos multirresistentes.

A3 receptor agonist modulates IL-1ß hippocampus levels in a rat model of neuropathic pain

Introduction: Considering the lack of specific treatments for neuropathic pain, this study aimed to evaluate the effect of a single dose of adenosine A3 receptor IB-MECA on inflammatory and neurotrophic parameters in rats subjected to a neuropathic pain model. Methods: 64 adult male Wistar rats were used. Neuropathic pain was induced by chronic constriction injury (CCI) of the sciatic nerve and the treatment consisted of a 0.5 µmol/kg dose of IB-MECA, a selective A3 adenosine receptor agonist, dissolved in 3% DMSO; vehicle groups received DMSO 3% in saline solution, and morphine groups received 5 mg/kg. Cerebral cortex and hippocampus IL-1ß, BDNF, and NGF levels were determined by Enzyme-Linked Immunosorbent assay. Results: The main outcome was that a single dose of IB-MECA was able to modulate the IL-1ß hippocampal levels in neuropathic pain induced by CCI and the DMSO increased IL-1ß and NGF hippocampal levels in sham-operated rats. However, we did not observe this effect when the DMSO was used as vehicle for IB-MECA, indicating that IB-MECA was able to prevent the effect of DMSO. Conclusions: Considering that the IL-1ß role in neuropathic pain and the contributions of the hippocampus are well explored, our result corroborates the relationship between the A3 receptor and the process of chronic pain maintenance.

Portación nasal, antibiotipo y genotipo de cepas de Staphylococcus aureus aisladas en estudiantes de Medicina y de Enfermería Campus San Felipe, Universidad de Valparaíso, Chile, durante el año 2017 / Nasal carriage, antibiotype and genotype of isolated Staphylococcus aureus from Medicine and Nursing students of Campus San Felipe, University of Valparaiso, Chile, during 2017

INTRODUCCIÓN. Staphylococcus aureus es parte de la microbiota nasal en 20-30% de la población general, colonización que constituye un reservorio para su transmisión, lo que es preocupante en cepas resistentes a meticilina (SARM). OBJETIVO: Determinar la prevalencia de S. aureus en estudiantes de Medicina y Enfermería del Campus San Felipe y caracterizar sus aislamientos. MATERIAL Y MÉTODOS: El 2017 se midió la portación nasal a 225 estudiantes, a las cepas aisladas se le analizó su antibiotipo por difusión en agar, la relación clonal por electroforesis de campo pulsado y MLST. En SARM se determinó el cassette SCCmec y gen de la leucocidina de Panton-Valentine. RESULTADOS: 61 estudiantes portaron S. aureus (27,1%) incluyendo dos cepas SARM (0,9%). Staphylococcus aureus mostró resistencia a penicilina (75%), eritromicina (14%) y clindamicina (10%), cloranfenicol (1,6%) y levofloxacina, oxacilina, cefoxitina (3,3%). Se diferenciaron diecinueve pulsotipos y el secuenciotipo coincidió con complejos clonales descritos a nivel mundial en portadores de S. aureus: CC30, CC8, CC97, CC15, CC22 y CC1. Las dos cepas SARM correspondieron con los clones chileno/cordobés y USA100NY/J, ambas del CC5. CONCLUSIÓN: La portación nasal de S. aureus y SARM en los estudiantes coincidió con la portación en la población general y las cepas sensibles a meticilina mostraron diversidad clonal y alta susceptibilidad antimicrobiana, exceptuando a penicilina.

BACKGROUND: Staphylococcus aureus is part of the nasal microbiota in 20-30% of the population. This colonization is also a reservoir for its dissemination, which is worrying in the case of strains with resistance to methicillin (MRSA). AIM: To determine S. aureus nasal carriage in nursing and medical students of San Felipe Campus and characterize theirs isolates. METHODS: During 2017, nasal swabs were taken from 225 students and seeded in salt manitol agar. Antibiotypes were determined by agar diffusion and the genetic clonality was assessed by PFGE and MLST in isolated S. aureus. SCCmec cassette and Panton-Valentine leukocidin gene (pvl) presence were determined in the MRSA isolates. RESULTS: 61 students carried S. aureus (27.1%) including two MRSA strains (0.9%). S. aureus showed resistance to penicillin (75%), erythromycin (14%) and clindamycin (10%), chloramphenicol (1.6%) and levofloxacin, oxacillin, cefoxitin (3.3%). Nineteen PFGE-types were differentiated, and their sequence-types coincided with main clonal complexes described in S. aureus carriers from different places worldwide: CC30, CC8, CC97, CC15, CC22 and CC1. MRSA strains belonged to CC5 and they corresponded to the Chilean/Cordobes and USA100NY/J clones. CONCLUSION: Nasal carriage of S. aureus and MRSA in students, coincided with the general population and sensitive-methicillin strains showed clonal diversity and high antimicrobial susceptibility except for penicillin.

Incidence and characteristics of methicillin-resistant coagulase-negative Staphylococcus aureus in peritoneal dialysis-associated peritonitis in a single center using molecular methods.

PURPOSE: Peritonitis is a serious complication of peritoneal dialysis and coagulase-negative Staphylococcus (CNS) is the most frequent cause of peritoneal dialysis (PD)-infections in many centers. This study aimed to investigate the molecular epidemiology of CNS isolated from PD-peritonitis in a Brazilian single center, focusing on the genetic determinants conferring methicillin resistance. METHODS: Bacterial strains were isolated from peritoneal fluid of patients presenting PD-peritonitis, identified by phenotypic and molecular methods, and those identified as CNS were submitted to mecA detection, SCCmec, pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). RESULTS: Over the 18-year period of this study (1995-2011), a total of 878 peritonitis episodes were diagnosed in this unit, 115 were caused by coagulase-negative staphylococci of which 72 by Staphylococcus epidermidis. mecA gene was detected in 55 CNS (47.8%), more frequently on the more recent years. SCCmec type III was the most frequent cassette, followed by SCCmec type IV and SCCmec type II. A diverstity of pulsotypes was observed among the S. epidermidis isolates, but five clusters (based on the 80% cutoff) were identified. Diversified sequence types (ST02, ST05, ST06, ST09, ST23, ST59 and ST371) were detected. CONCLUSIONS: Detection of SCCmec type III among coagulase-negative Staphylococcus underscores the role of hospital environments as potential source of methicillin-resistant Staphylococcus causing peritonitis in PD patients.

Molecular Detection and Characterization of the mecA and nuc Genes From Staphylococcus Species (S. aureus, S. pseudintermedius, and S. schleiferi) Isolated From Dogs Suffering Superficial Pyoderma and Their Antimicrobial Resistance Profiles.

Canine superficial pyoderma (CSP) is a bacterial infection secondary to several skin diseases of the dog. Staphylococcus pseudintermedius, which is a commensal bacterium of the dog's skin, is the leading agent found in dogs affected by CSP, which can progress to deep pyoderma. It is also of clinical significance because S. pseudintermedius strains carry antimicrobial resistance genes, mainly the mecA gene. In this descriptive longitudinal study, molecular characterization of bacterial isolates from dogs affected by CSP was performed in addition to phenotyping, antimicrobial profiling, and assessment of resistance carriage status. Fifty dogs (24 females and 26 males) attending the CES University Veterinary Teaching Hospital were included in the study. CSP was confirmed according to clinical signs and cytological examination. Swabs were taken from active skin lesions for bacterial culture, and phenotyping and antimicrobial resistance profiles were assessed using API-Staph phenotyping and the Kirby-Bauer method, respectively. We also performed molecular detection and characterization of the mecA and nuc encoding gene of coagulase-positive Staphylococci. The mecA gene frequency was established by qPCR amplification of a 131bp gene fragment. Data were evaluated by descriptive statistics. Erythema, peeling, pruritus, and alopecia were the predominant symptoms (72, 56, and 46%, respectively). We isolated bacteria compatible with Staphylococcus species from all samples tested. API phenotyping showed 83.1 to 97.8% compatibility with S. pseudintermedius. PCR-genotyping resulted in 15, 3, and 1 isolates positive for S. pseudintermedius, S. aureus, and S. schleiferi, respectively. Isolated strains showed high susceptibility to Imipenem, Ampicillin/Sulbactam, and Rifampicin (100, 94, and 92%, respectively). The highest resistance was against Vancomycin and Trimethoprim/Sulfamethoxazole (98 and 74%, respectively). S. pseudintermedius, S. aureus, and S. schleiferi isolates were cloned and shared 96% sequence homology. Finally, we found 62% carriage status of the mecA gene in isolates of CSP patients, although only 36% of the isolates were methicillin-resistant. Identification of three Staphylococcus species causing CSP, high-level resistance against conventional antimicrobials, and carriage of the mecA gene highlight the importance of performing molecular characterization of bacteria causing dermatological conditions in dogs.

Tipificación del cassette cromosómico estafilocócico de Staphylococcus aureus resistentes al meticilino en el estado de Aragua, Venezuela / Typification of the staphylococcal chromosome cassette of methicillin-resistant Staphylococcus aureus in the state of Aragua, Venezuela

RESUMEN Objetivos: Tipificar el casette SCCmec en cepas de Staphylococcus aureus resistentes a meticilino (SARM) en aislados clínicos de centros de salud del Estado Aragua-Venezuela y comparar la presencia de los genotipos SCCmec entre los centros de salud del estado y según el tipo de infección. Materiales y métodos: Durante enero y agosto de 2015 se estudiaron 81 cepas SARM de cuatro centros de salud del estado de Aragua en Venezuela. La resistencia al meticilino se midió con el método de Kirby-Bauer con discos de oxacilina (1 µgr) y cefoxitina (30 µgr). El gen mecA y el SCCmec se analizaron por la técnica de reacción en cadena de polimerasa múltiple. Resultados: 55 aislados (67,9%) amplificaron el gen mecA, y 24 cepas (43,6%) amplificaron el SCCmec. El SCCmec I fue el más frecuente, seguido de SCCmecIV y SCCmec III, representaron el 62,5%, 25% y 12,5%, respectivamente. El SCCmec I fue predominante en el centro de salud A (80%), mientras que el SCCmec IV se encontró en el centro de salud B (60%) y C (100%). En el centro de salud D, 50% resultó ser SCCmec I y 50% SCCmec IVd. Se encontró relación entre el SCCmec y el centro de salud con significancia estadística. En infecciones de piel y tejidos blandos y en las respiratorias predominó el SCCmec I con 63,2% y 50% respectivamente. Conclusiones: La frecuencia de SCCmec I y IV permitirá establecer nuevas medidas en el uso y control de la resistencia a los antibióticos.

ABSTRACT Objective: Typify the SCCmec cassette in methicillin-resistant strains of Staphylococcus aureus in clinical isolates from health centers in the State of Aragua-Venezuela and compare the presence of SCCmec genotypes among the state health centers and according to the type of infection. Materials and methods: 81 MRSA strains from four health centers of the Aragua-Venezuela State were studied. Methicillin resistance was performed with the Kirby-Bauer method with oxacillin (1 µg) and cefoxitin (30 µg) disks. The mecA gene and SCCmec were analyzed by the multiple PCR technique. Results: Only 55 isolates (67.9%) amplified the mecA gene, and 24 strains (43.6%) amplified SCCmec. SCCmec type I was the most frequency, followed by SCCmec IV and SCCmec III, representing 62.5%, 25% and 12.5%, respectively. SCCmec I was predominant in health center A (80%), while in B and C 60% and 100% respectively were SCCmec IV. At health center D, 50% turned out to be SCCmec I and 50% SCCmec IVd. A relationship was found between the SCCmec and the health center with statistical significance. SCCmec I predominated in skin and soft tissue and respiratory infections with 63.2% and 50%, respectively. There was no association between genotype and type of infection with a p value greater than 0.05. Conclusions: The prevalence of SCCmec I and IV will allow establishing new measures in the use of antibiotics and epidemiological control.