RESUMO: A resistência à meticilina no grupo Staphylococcus intermedius (GSI) tem aumentado na clínica de pequenos animais. Membros GSI resistentes à meticilina (GSIRM) têm sido causas de infecções tanto em animais de companhia e humanos. Cassetes cromossômicos estafilocócicos mec (SCCmec) carregam os genes mecA/C, que codificam a transpeptidase PBP2a (PBP2’) responsável pela resistência aos antibióticos β-lactâmicos em estafilococos. Nosso objetivo foi investigar os elementos SCCmec de GSIRM isolados de diferentes amostras clínicas de cães que exibiram CIM de meticilina ≥ 0,5 μg/mL por meio de um sistema automatizado em um Centro Veterinário de Diagnósticos em São Paulo, Brasil. A sensibilidade à meticilina foi determinada por meio do teste de microdiluição em caldo e fitas Oxoid® M.I.C.Evaluator®. A produção de PBP2a foi detectada usando um ensaio de aglutinação de látex. A tipagem dos elementos SCCmec foi realizada de acordo com as diretrizes do International Working Group on the Classification of Staphylococcal Cassette Chromosome Elements (IWG-SCC). SCCmec tipo II (2A), SCCmec tipo III (3A), SCC compostos de um complexo mec de classe A com múltiplos complexos ccr, e elementos SCCmec não tipáveis foram encontrados nesses isolados GSIRM. Variantes que diferem dos elementos SCCmec reconhecidos até o momento pelo IWG-SCC foram encontradas, indicando novos rearranjos no contexto genético de mecA nesses isolados GSIRM caninos.

ABSTRACT: Methicillin resistance in the Staphylococcus intermedius group (SIG) has emerged in small animal practice. Methicillin-resistant SIG (MRSIG) members have been implicated as causes of infections in both companion animals and humans. Staphylococcal cassette chromosome mec (SCCmec) elements carry the mecA/C genes, which encode for the transpeptidase PBP2a (PBP2’) responsible for β-lactam antibiotic resistance in staphylococci. This study examined the SCCmec types of MRSIG isolates from different clinical specimens of dogs that exhibited methicillin MIC ≥ 0.5 μg/mL by an automated identification and susceptibility system in a Center for Veterinary Diagnostics in São Paulo, Brazil. Susceptibility to methicillin was determined by broth microdilution testing, and Oxoid® M.I.C.Evaluator® strips. PBP2a production was detected using a latex agglutination assay. SCCmec typing was performed according to the International Working Group on the Classification of Staphylococcal Cassette Chromosome Elements (IWG-SCC) guidelines. SCCmec type II (2A), SCCmec type III (3A), composite SCC structures consisting of a class A mec gene complex in addition to multiple ccr gene complexes, and non-typable SCCmec elements were reported in these MRSIG isolates. SCCmec type variants differing from those so far acknowledged by IWG-SCC were found, indicating new rearrangements in the genetic context of mecA in these canine MRSIG isolates.

It is known that Aeromonas spp. possess different chromosomal β-lactamase genes. Presence and phenotypic expression of blaTEM, blaSHV, and blaCTX-M ESBL-encoding genes were investigated in environmental water isolates of Aeromonas hydrophila and Aeromonas jandaei. Presence of blaSHV and blaCTX-M genes was not observed, and blaTEM gene was verified in 91% of the isolates. Sequencing of 10 fragments showed the occurrence of blaTEM-116.

Methicillin-resistant Staphylococcus aureus is an established nosocomial pathogen (HA-MRSA, hospital acquired MRSA), but has recently begun to appear in the community (CA-MRSA, community acquired MRSA). The cause of resistance to methicillin and all other β-lactam antibiotics is the mecA gene, which is situated on a mobile genetic element, the Staphylococcal Cassette Chromosome mec (SCCmec). Seven major variants of SCCmec, type I to VII are distinguished. HA-MRSA disseminated worldwide and causes the majority of S. aureus nosocomial infections with a limited number of clones disseminated including the Brazilian Epidemic Clone (BEC, ST239-MRSA-III). CA-MRSA isolates are susceptible to non-β-lactam antibiotics, usually isolated from healthy individuals which do not possess any unknown risk factors for MRSA infection and are associated with a larger clonal diversity compared with HA-MRSA. However, during recent years distinction between HA-MRSA and CA-MRSA is beginning to fade. Actually, knowledge about MRSA disseminating clones is required to implement any strategies to control the transmission of MRSA either within hospitals or in community. For this reason, rapid identification of strains is an important issue. The rate of HA-MRSA can be reduced substantially through the implementation of interventions strategies, even in settings where MRSA is endemic as in most Brazilian hospitals. However, these policies could be quite complicated in the light of an increasing CA-MRSA prevalence in healthcare facilities, considering that distinction between HA-MRSA and CA-MRSA has started to disappear.

Methicillin-resistant Staphylococcus aureus (MRSA) commonly causes infection in hospitalized patients. Since its appearance in the 1960s, the SCCmec has evolved throughout the years into 5 different types (I-V), each bearing a different set of genes. Infection with MRSA SCCmec types I, II or III is almost exclusively restricted to hospitalised patients. However, recently, community acquired MRSA (CA-MRSA) infections have been reported with increasing frequency, usually caused by a type IV SCCmec MRSA in nosocomial settings. We studied the prevalence of SCCmec types in 50 nosocomial strains collected from 1995 to 1999. The SCCmec complex type and presence of Panton-Valentine leukocidin (PVL) were determined by PCR. Strains had been previously typed by PFGE and were now typed by MLST. We found that 3 of the isolates studied bore a type IVc SCCmec all having different PFGE and MLST profiles (ST3, ST5 and ST88). All strains bearing a type III SCCmec belonged to MLST ST239 (Brazilian/Iberian clone). Only the strain which presented the ST5 profile bore the pvl gene. The type IVc SCCmec strains presented relatively lower levels of resistance to oxacillin in comparison to the type III SCCmec strains. The pattern of dissemination of the type IV SCCmec remains to be elucidated. The finding of strains carrying a type IV SCCmec in the present study among strains isolated at least 7 years ago indicates that clones bearing a type IV SCCmec have been present in Brazil for quite some time, and must have gone by undetected.

Resistance in Mycobacterium tuberculosis to isoniazid (INH) is caused by mutations in the catalase-peroxidase gene (katG) , and within the inhA promoter and/or in structural gene. A small percentage (~ 10%) of INH-resistant strains do not present mutations in both of these loci. Other genes have been associated with INH resistance including the gene encoding for NADH dehydrogenase (ndh) . Here we report the detection of two ndh locus mutations (CGT to TGT change in codon 13 and GTG to GCG change in codon 18) by analyzing 23 INH-resistant and in none of 13 susceptible isolates from Brazilian tuberculosis patients. We also detected two isolates without a mutation in ndh, or any of the other INH resistance-associated loci examined, suggesting the existence of additional, as yet to be described, INH resistance mechanisms.

A habilidade do brometo de dioctadecildimetilamônio (DOBAB), em formar bicamada de lipídio sintético e a demonstração prévia do forte poder solubilizante de anfotericina B (ANB) , incentivou-nos a realizar a avaliação da atividade de DODAB/AMB in vivo contra candidíase sistêmica em modelo de camundongos para verificar a sua sobrevida bem como a recuperação das leveduras de C. albicans dos órgãos colonizados (baço e rins). O AMB foi simplesmente adicionado à DODAB em pó previamente disperso em água e sonicado com auxílio de ponteiras, nas concentrações de <0,1e 10 mg/mL, respectivamente, assegurando-se a completa ausência de solventes orgânicos nesta formulação. O estado de agregação do AMB foi avaliado por meio do espectro de absorção da luz UV-visível e a distribuição de tamanhos das formulações estudadas, determinadas por meio de analisador ZetaPlus Brookhaven Instruments Corporation, Holtsville, NY) equipado com um laser de 570 nm e dynamic light scattering (90ºC) para a medição dos tamanhos (Grabowski & Morrison, 1983). AMB foi estabilizada na bicamada de DODAB em forma monomérica, eliminando-se os grandes agregados de AMB insolúveis em água. Tanto a sobrevida dos animais como os experimentos com recuperação das leveduras dos órgãos colonizados (baço e rins) mostraram eficácia equivalente à demonstrada por Fungizona (DOC/AMB) - a sobrevida foi de 100 e 70% respectivamente nas concentrações de 0.4 mg/kg/dia via i.p. por 10 dias (P>0.05), em relação a eliminação da colonização de C. albicans dos rins e baço. Em resumo, DOD/AMB foi efetivo no tratamento de candidíase sistêmica em modelo animal.

The ability of the versatile dioctadecyldimethylammonium bromide (DODAB), a bilayer-forming synthetic lipid previously shown to solubilize Amphotericin B (AMB), inspired this evaluation of in vivo activity of the DODAB/AMB formulation (DOD/AMB) against systemic candidiasis in a mouse model from survival and tissue burden experiments. AMB was simply added to a DODAB powder dispersion in water previously obtained by sonication with tip at concentrations <0.1 and 10 mg/mL, respectively, organic solvents completely absent. AMB aggregation state was evaluated from UV-visible light absorption and dynamic light scattering for aggregate sizing. AMB was stabilized by the DODAB bilayer fragments in its monomeric form, causing disappearance of large water insoluble drug aggregates. From survival and tissue burden experiments, DOD/AMB efficacy was equivalent to the one exhibited by Fungizone (DOC/AMB) - 100 and 70% survival respectively, at 0.4 mg/kg/day given i.p. for 10 days (P>0.05) -, regarding elimination of Candida colonization in spleen and kidneys. In summary, DOD/AMB, was effective for treating systemic candidiasis in a mouse model.

O objetivo deste trabalho foi o desenvolvimento de imunorreagentes policlonais convenientes para uso em um teste imunoenzimático para detecção rápida de Salmonella em alimentos. Foram produzidos seis anti-soros policlonais anti-Salmonella contendo as aglutininas e, h; 1,6; i; 1,2; f,g,s e m,t. Como antígenos, foram empregadas culturas formolizadas de quatro sorotipos de Salmonella (S. Typhimurium fases 1 e 2, S. Anatum fases 1 e 2, S. Agona monofásica e S. Oranienburg monofásica) cultivadas em caldo tripticase de soja. O teste imunoenzimático utilizado foi o indireto, empregando-se anti-IgG-peroxidase como conjugado e OPD-H2O2 como sistema cromógeno. Os testes imunoenzimáticos foram conduzidos com os anti-soros policlonais individuais absorvidos e não-absorvidos, bem como empregando-se um anti-soro polivalente, correspondente a um "pool" de anti-soros absorvidos e não-absorvidos com culturas puras de Salmonella e outras enterobactérias. A absorção dos soros eliminou as reações cruzadas com todas as enterobactérias testadas nesse estudo, apresentadas tanto por alguns anti-soros individuais quanto pelo polivalente. Entre os seis anti-soros estudados, os que apresentaram melhor desempenho foram o f,g,s não-absorvido e o polivalente absorvido.

This research aimed for the development of convenient policlonal immunoreagents to be used in an enzyme immunoassay for detection of Salmonella. Six anti-Salmonella antisera were prepared, which contained the following agglutinins: e, h. 1.6; i; 1.2; f,g,s and m,t. The antigens consisted of formolized cultures of four Salmonella serotypes (S. Typhimurium phases 1 and 2, S. Anatum phases 1 and 2, S. Agona and S. Oranienburg), grown in tryptic soy broth. An indirect-type immunoassay was used, employing anti-IgG-peroxidase as conjugate and OPD-H2O2 as chromogen. These tests were developed with individual and polivalent sera (pool of the individual sera), absorbed and non-absorbed with cultures of Salmonella and other enterobacteria. Absorption eliminated the cross reactions with enterobacteria, presented by some individual sera and by the polivalent serum. The best performance in the immunoassay was achieved by the non-absorbed f,g,s and by the absorbed polivalent sera.

Cinqüenta felinos hígidos, adultos - vinte e seis machos e vinte e quatro fêmeas - sem definição racial, com distintas idades, reunidos em dois grupos (GRUPO 1- animais domiciliados e GRUPO 2 - animais que rendados) de vinte e cinco animais cada. Após anamnese e prévia sedação, os gatos foram submetidos a exame físico e à otoscopia, comprovando-se a ausência de lesões meatais e timpânicas, com cureta de Buck e wragatoas estéreis, foi colhido material meatal, que foi então submetido a exames bacteriológico e micológico. No Grupo 1, foi evidenciado presença de Otodectes cynotis (4% das amostras), em 72% dos casos de bolores e leveduras (Cladosporium sp 66,6%, Malassezia sp 40%, Penicillium sp 33%, Aspergillus sp 33,3%, Rhodotorula sp 20,0%, Mycelia sp 13,3% e Alternaria sp, Aureobasidium sp, Ryzopus sp, Trichosporon sp, todos com 6,6%) e, finalmente, em 64% da amostragem, bactérias dos gêneros Staphylococcus spp (81,2%), Pseudomonas sp (12,5%), Klebsiella sp (12,5%), Acinetobacter sp, Bacilos difteróides, Enterobacter sp, Lactobacillus spp (todos com 6,2%). No Grupo 2, o Otodectes sp foi identificado em 36% das amostras, em 96% daquelas isolaram-se fungos dos gêneros: Malassezia sp - 54,1%, Aspergillus e Penicillium sp, ambos com 33,3%, Microsporum sp - 29,1%, Cladosporium sp - 16,6%, Trichoderma sp - 12,5%, Alternaria e Phoma sp, ambos com 8,3% e Epicoccum sp, Neurospora sp. Mycelia sp, Rhodotorula sp, todos com 4,1% e, por fim, em 20 das 25 amostras (80%) isolaram-se pelo menos uma cepa bacteriana (Staphylococcus spp 75%, Klebsiella sp 20,8%, Bacilos difteróides 12,5%, Pseudomonas sp, 8,3%) e Acinetobacter sp, Enterobacter sp e Escherichia sp, todos com 4,1% cada um em cultivo monoespecífico ou em associação.

The study aimed to find out the frequency of microrganisms in the external ear canal in healthy cats. Fifty adult male (26) and female (24) short hair cats were utilized for the purpose, and they were divided according to living enviroment (Group l - indoor and Group 2 - outdoor). All animals were submitted to sedation prior to physical and otoscopic examinations in order to confirm to be healthy. Samples were obtained from ear canal by use of Buck curetes and sterile cotton sticks for microbiological investigation. In cats of Group 1, Otodects cynotis were observed in 4%. In 72% of the animals yeasts and molds were found (Cladosporium sp 66.6%, Malassezia sp 40%, Penicillium sp 33%, Aspergillus sp 33.3%, Rhodotorula sp 20%, Mycelia sp 13.3%, and 6.6% for Alternaria sp, Aureobasidium sp, Ryzopus sp and Trichosporon sp), finally, m 64% of the samples, was detected Staphylococcus spp (81.2%), Pseudomonas sp (12.5%), Klebsiella sp (12.5%), and 6,2% for Acinetobacter sp. Bacilos difteroides, Enterobacter sp, Lactobacillus spp. In Group 2, O. cynotis was isolated in 36% of cats, and in 96% of the animals wasfound Malassezia sp (54.1%), Aspergillus sp (33.3%), Penicillium (33.3%), Microsporum sp (29.1%), Cladosporium sp (16.6%), Trichoderma sp (12.5%) Alternaria sp (8.3%), Phoma sp (8.3%), Epicoccum sp (4.1%), Neurospora sp (4.1%), Mycelia sp (4.3%), Rhodotorula sp (4.1%). In 20 out of 25 (80%), at least one bacterial microrganism was isolated (Staphylococcus sp 75%, Klebsiela sp 20.8%, Bacilos difteroides 12.5%, Pseudomonas sp 8.3%, and 4.1% for Acynetobacter sp, Enterobacter and Escherichia sp).