This Regional Research Communication describes the characterisation of ampicillin, penicillin and tetracycline resistance inStaphylococcus aureusisolated from bovine subclinical mastitis in Minas Gerais State, Brazil. NinetyS. aureusisolates from bovine mastitis exhibiting phenotypic resistance to ampicillin, penicillin and/or tetracycline were selected for this study. The minimum inhibitory concentration (MIC) of each antibiotic was determined using the E-Test^®and the production of beta-lactamase was determined by cefinase disks. The resistance genesblaZ, tet(K),tet(L),tet(M), andtet(O) were investigated by PCR in all of the isolates. The MIC results classified 77, 83 and 71% of the isolates as resistant to ampicillin, penicillin and tetracycline, respectively. The MIC₅₀and MIC₉₀were, respectively, 1 and 2 µg/ml for ampicillin, 0·5 and 1 µg/ml for penicillin and 32 and 64 µg/ml for tetracycline. Eighty-six per cent of beta-lactamase producing isolates were detected. Of the 90 isolates investigated, 97% amplifiedblaZ, 84% amplifiedtet(K), 9% amplifiedtet(L), 2% amplifiedtet(M) and 1% amplifiedtet(O). Seventy-nine isolates (88%) showedblaZtogether with at least onetetgene.S. aureusisolates showed high MIC₅₀and MIC₉₀values for the three antimicrobials. TheblaZ andtet(K) genes were widespread in the herds studied, and most of the isolates harbouredblaZ andtet(K) concomitantly.

Streptococcus pyogenes (group A Streptococcus-GAS) is an important pathogen for humans. GAS has been associated with severe and invasive diseases. Despite the fact that these bacteria remain universally susceptible to penicillin, therapeutic failures have been reported in some GAS infections. Many hypotheses have been proposed to explain these antibiotic-unresponsive infections; however, none of them have fully elucidated this phenomenon. In this study, we show that GAS strains have the ability to form antimicrobial persisters when inoculated on abiotic surfaces to form a film of bacterial agglomerates (biofilm-like environment). Our data suggest that efflux pumps were possibly involved in this phenomenon. In fact, gene expression assays by real-time qRT-PCR showed upregulation of some genes associated with efflux pumps in persisters arising in the presence of penicillin. Phenotypic reversion assay and whole-genome sequencing indicated that this event was due to non-inherited resistance mechanisms. The persister cells showed downregulation of genes associated with protein biosynthesis and cell growth, as demonstrated by gene expression assays. Moreover, the proteomic analysis revealed that susceptible cells express higher levels of ribosome proteins. It is remarkable that previous studies have reported the recovery of S. pyogenes viable cells from tissue biopsies of patients presented with GAS invasive infections and submitted to therapy with antibiotics. The persistence phenomenon described herein brings new insights into the origin of therapeutic failures in S. pyogenes infections. Multifactorial mechanisms involving protein synthesis inhibition, cell growth impairment and efflux pumps seem to play roles in the formation of antimicrobial persisters in S. pyogenes.

The serrano artisan cheese produced from raw milk of dairy cattle is a typical product of high-altitude farms in the states of Santa Catarina and Rio Grande do Sul. However, marketing of the cheeses occurs illegally because they lack the minimum maturation period required for cheese produced from raw milk. The production of artisan cheeses is required to follow strict hygiene standards. This study aimed to test the quality and safety of cheeses that were produced in 31 farms of the Serrana region in Santa Catarina after 14 and 28 days of maturation. Coliform count was measured at 35 °C, and presence of other microorganisms such as Escherichia coli, Staphylococcus, Listeria spp., and Salmonella spp. were also tested. Fat and protein percentages, acidity, salt content, and humidity were also evaluated. Data were subjected to statistical analyses using the SAS® software. After 14 and 28 days of maturation, 74.19% (23/31) and 64.52% (20/31) of samples, respectively, showed higher numbers of coliforms at 35 °C than those permissible by law. Higher than permissible numbers of E. coli were observed in 45.16% (14/31) and 48.39% (15/31) of the samples analyzed after 14 and 28 days of maturation, respectively. Coagulase-positive staphylococci values above 103 CFU/g were observed in 54.84% (17/31) and 51.61% (16/31) of cheese samples after 14 and 28 days of maturation, respectively. Contamination with Salmonella spp. was not detected. However, Listeria monocytogenes serovar 4b was isolated in 3.23% (1/31) and 6.45% (2/31) of samples after 14 and 28 days of maturation, respectively. The results of humidity tests classified the cheese samples into three categories: low, medium, and high humidity. Semi fat cheeses were predominant in both maturation periods, although the samples were classified in thin, semi fat, and fat cheeses. The main variations in the compositions of analyzed samples occurred for salt and acidity levels. The maturation process has not proven to be effective in reducing microbiological contamination to compliance levels. Considering the heterogeneity of the analyzed cheese samples, the frequency of non-conformities with respect to microorganisms and pathogens present in the samples, this study indicates the necessity to improve the Serrano artisan cheese production system through adoption of good manufacturing practice measures.

Introduction. In some species, the population structure of pathogenic bacteria is clonal. However, the mechanisms that determine the predominance and persistence of specific bacterial lineages of group C Streptococcus remain poorly understood. In Brazil, a previous study revealed the predominance of two main lineages of Streptococcus dysgalactiae subsp. equisimilis (SDSE). Aim. The aim of this study was to assess the virulence and fitness advantages that might explain the predominance of these SDSE lineages for a long period of time. Methodology. emm typing was determined by DNA sequencing. Adhesion and invasion tests were performed using human bronchial epithelial cells (16HBE14o-). Biofilm formation was tested on glass surfaces and the presence of virulence genes was assessed by PCR. Additionally, virulence was studied using Caenorhabditis elegans models and competitive fitness was analysed in murine models. Results. The predominant lineages A and B were mostly typed as emm stC839 and stC6979, respectively. Notably, these lineages exhibited a superior ability to adhere and invade airway cells. Furthermore, the dominant lineages were more prone to induce aversive olfactory learning and more likely to kill C. elegans. In the competitive fitness assays, they also showed increased adaptability. Consistent with the increased virulence observed in the ex vivo and in vivo models, the predominant lineages A and B showed a higher number of virulence-associated genes and a superior ability to accumulate biofilm. Conclusion. These results suggest strongly that this predominance did not occur randomly but rather was due to adaptive mechanisms that culminated in increased colonization and other bacterial properties that might confer increased bacteria–host adaptability to cause disease.

Multidrug resistance is commonly acquired by transferring DNA from one bacterium to another. However, the mechanisms that enhance the acquisitions of foreign genes are poorly understood, as well as the dynamics of their transmission between hosts in different environments. Here, genomic approaches were applied to evaluate the enrichment of the S. aureus chromosome with resistance traits in groups of genomes with or without anti-restriction genes and to analyze some evolutionary aspects of these acquisitions. Furthermore, the role played by an anti-restriction gene in improving multiresistance in MRSA was investigated by molecular cloning. A strong association was observed between the presence of anti-restriction gene homologs and patterns of multidrug resistance. Human isolates, mainly ST239-SCCmecIII, carry ardA-H1, and from animal sources, mainly CC398, carry ardA-H2. Increased DNA transfer was observed for clones that express the ardA-H1 allele, corroborating its role in promoting gene transfer. In addition, ardA-H1 was expressed in the dsDNA format in the BMB9393 strain. The evolution of successful multidrug-resistant MRSA lineages of the ST239 and ST398 was initiated not only by the entry of the mec cassette but also by the acquisition of anti-restriction gene homologs. Understanding the mechanisms that affect DNA transfer may provide new tools to control the spread of drug resistance.

Peritoneopericardial diaphragmatic hernia (PPDH) is a communication between the abdomen and the pericardial sac generated by congenital anomalies triggered during diaphragmatic and pericardial development. This report aimed to present the case of an adult, mixed-breed cat, affected by PPDH, focusing on the period from diagnosis to successful surgical correction. The patient had a capricious appetite and weight loss for about four months and started, at the end of this period, a state of apathy. On abdominal ultrasound, the gallbladder (GB) was close to the heart, suggesting diaphragmatic discontinuity. On thoracic radiography, there were changes suggestive of PPDH, pericardial efusion or cardiomegaly with probable dilated cardiomyopathy. Based on these findings, an echocardiogram was performed, highlighting the hepatic lobe and GB internally to the pericardium, causing cardiac compression, although without severe cardiac changes. During surgery, a diaphragmatic defect of 4 cm in diameter was observed with the congested right medial hepatic lobe and hyperemic GB in the pericardial sac. The defect was sutured using the sultan pattern in separate stitches and polyamide threads. The feline returned to feeding with greater interest soon after the surgery, and after 15 days it was fed with dry food and had normal behavior. PPDH can be diagnosed in healthy adult cats, even if there are no apparent respiratory, gastrointestinal, or cardiac signs. The echocardiogram is relevant in the definitive diagnosis, in addition to excluding differential diagnoses, and simple surgical treatment with polyamide thread and sultan suture is successful. A hérnia periotônio-pericárdica diafragmática (HPPD) comunica o abdome e o saco pericárdico, é gerada por anomalias congênitas deflagradas no desenvolvimento diafragmático e pericárdico. O objetivo deste relato é apresentar o caso de uma gata adulta, mestiça, acometida por HPPD, destacando do diagnóstico à correção cirúrgica bem-sucedida. A paciente exibia apetite caprichoso e emagrecimento há cerca de 4 meses, iniciando ao final deste período, apatia. Na ultrassonografia abdominal a vesícula biliar (VB) estava próxima ao coração sugerindo ruptura diafragmática. Na radiografia torácica evidenciou-se alterações sugestivas de HPPD ou cardiomegalia com provável miocardiopatia dilatada. Devido tais achados realizou-se ecocardiograma destacando o lobo hepático e a VB no pericárdio comprimindo o coração sem comprometer sua função. Na cirurgia observou-se defeito diafragmático (4 cm), passagem do lobo hepático medial direito e da VB para o pericárdio. Suturou-se o defeito com padrão sultan e fio poliamida 3.0. A felina voltou a se alimentar com maior interesse logo após a cirurgia. A HPPD pode ser diagnosticada em felinos adultos saudáveis, mesmo que não haja sinais respiratórios, gastrointestinais ou cardíacos aparentes. O ecocardiograma é relevante no diagnóstico definitivo, além de excluir diagnósticos diferenciais, sendo o simples tratamento cirúrgico com fio poliamida e sutura sultan bem-sucedido.