Relationship between Biofilm Production and High Somatic Cell Count in Streptococcus agalactiae Isolated from Milk of Cows with Subclinical Mastitis.

Streptococcus agalactiae (S. agalactiae) is one of the main agents that causes mastitis in dairy cows, mainly inducing the subclinical form, which is characterized by a high somatic cell count (SCC). The aim of this study was to correlate the increase in SCC caused by S. agalactiae in cows with subclinical mastitis to the presence of genes related to adhesion and invasion in bovine mammary epithelial cells (BMEC) and biofilm formation. Considering the 145 isolates tested, 57.2% presented the capsular type Ia and 42.8% presented type III. We identified the virulence genes among the isolates and determined nine genetic profiles. The most common profile was identified in 69 isolates (47.5%): Ia, fbsA+, fbsB-, pI1-, pI2a-, pI2b+, and hylb+. All isolates produced biofilm, with 58.6% classified as strong producers, 29% as moderate producers and 12.4% as weak producers. No statistical correlation was found between the presence of virulence genes and increased SCC or biofilm production. However, biological evidence was observed between increased SCC and biofilm production. One isolate from each profile was randomly subjected to adhesion and invasion assays, and all of them adhered to BEMC, but none were able to invade. Our results showed that different genetic profiles do not provide advantages for bacteria to invade BMEC in vitro. In addition, biofilm production appears to be related to high SCC.

Efficacy of cefquinome and a combination of cloxacillin and ampicillin for treatment of dairy cows with Streptococcus agalactiae subclinical mastitis.

A randomized clinical trial was conducted to assess efficacy of intramammary cloxacillin and ampicillin (CLOXIMM), intramammary cefquinome (CEFIMM), and intramuscular cefquinome (CEFIM) to treat Streptococcus agalactiae intramammary infections (Trial 1). Subsequently, two treatment groups were extended to assess whether CLOXIMM was not inferior to CEFIMM (Trial 2). Nine farms were included in the study. Milk samples were collected from all quarters of all lactating cows for microbiological identification of S. agalactiae. Positive cows were randomly allocated into four groups: CLOXIMM, CEFIMM, CEFIM, or negative control (CONTROL). Study outcomes were bacteriological cure at 14 (CURE14), 21 (CURE21), and 14 and 21 (CURE1421) days after treatment onset, and somatic cell count. Logistic regression was used to estimate the odds of cure between each treatment and CONTROL. Non-inferiority analysis was performed considering a one-sided 95% confidence interval (CI) and non-inferiority margins (Δ) of 0.10, 0.15, 0.20, and 0.25. Adjusted S. agalactiae bacteriological cure for CLOXIMM, CEFIMM, CEFIM, and CONTROL was 86, 98, 55, and 25% at day 14; 82, 93, 52, and 0% at day 21; and 82, 92, 40, and 0% at days 14 and 21, respectively. Treatment with CLOXIMM and CEFIMM resulted in greater bacteriological cure rates, as compared with CEFIM or CONTROL, which does not justify the use of CEFIM in S. agalactiae eradication programs. The CURE14 difference between CEFIMM and CLOXIMM was of 12.1 percentage points (95% CI: 0.056-0.184). CLOXIMM was considered not inferior to CEFIMM for Δ = 0.20 or 0.25 and inconclusive for Δ = 0.10 or 0.15. Thus, it should be pondered by veterinarians whether an expected 12.1 (5.6-18.4) percentage points increase in cure rate would justify the use of a fourth-generation cephalosporin, as opposed to a combination of traditional IMM drugs (cloxacillin and ampicillin) to treat S. agalactiae subclinical mastitis.

Hygienic-sanitary quality of Minas fresh cheese sold in the city of Botucatu, São Paulo / Avaliação higiênico-sanitária de queijos Minas frescal comercializados na cidade de Botucatu, São Paulo

The Minas fresh cheese is a fresh and moist cheese and, therefore, has a short shelf-life. The use of raw milk as the main feedstock, the absence of ripening and the facility of contamination are factors that can compromise the product's microbiological quality. The aim of this study was to describe the hygienic-sanitary quality of 50 Minas fresh cheeses marketed in Botucatu city, São Paulo, Brazil, close to their production date, and another 50 units from the same lot exactly on the expiration date, according to the Brazilian Sanitary Surveillance Agency (ANVISA). We also searched for enterotoxins genes in Staphylococcus aureus and its in vitro expression. In 36% of the first analyzed samples, the count of coliforms at 45ºC was above the limit and, as for the second analysis, 44%. Regarding coagulase positive staphylococci, 10 (20%) samples showed concentration above the permitted by law next to the production date and 14% on the expiration date. Salmonella was only observed in one sample analyzed near the date of production, while L. monocytogenes only in one sample analyzed on the expiration day. We isolated three enterotoxigenic strains of S. aureus that produced Staphylococcal Enterotoxin B (SEB) and Staphylococcal Enterotoxin C (SEC) in vitro, highlighting the importance of proper storage of this product due to its potential to cause intoxication. Overall, the quality of Minas fresh cheese is still unsatisfactory, leading to risks to consumers' health.(AU)

O queijo Minas frescal é muito apreciado no Brasil. Por ser fresco e úmido, seu tempo de prateleira é curto. A utilização de leite cru como matéria prima, a não maturação do queijo e a facilidade de contaminação durante seu processamento são fatores que comprometem a qualidade microbiológica do produto. Este trabalho verificou a qualidade higiênico-sanitária de 50 queijos Minas frescal comercializados na cidade de Botucatu, São Paulo, Brasil, em datas próximas a sua fabricação e na data de expiração da validade, utilizando os parâmetros propostos pela RDC nº 12 da Agência Nacional de Vigilância Sanitária (ANVISA). Cepas de S. aureus foram submetidas à pesquisa de genes codificadores de produção de enterotoxinas clássicas e, também, sua produção in vitro. Encontraram-se coliformes a 45ºC, acima do permitido por legislação, em 36% das amostras durante a primeira análise e, na segunda, em 44%. Quanto à contagem de estafilococos coagulase positiva, 10% não atendiam ao padrão exigido por lei na data de produção, e 14%, na data de expiração da validade. Salmonella foi apenas observada em uma amostra próxima à data de produção, enquanto L. monocytogenes somente em uma amostra na data de expiração da validade. Observou-se produção de Staphylococcal Enterotoxin B (SEB)e Staphylococcal Enterotoxin C (SEC) in vitro por três cepas de S. aureus. Ressalta-se, em decorrência disso, a importância de armazenar corretamente esse alimento, por seu potencial de causar intoxicação. Concluiu-se que a qualidade higiênico-sanitária desses queijos Minas frescais foi insatisfatória, implicando riscos para a saúde do consumidor.(AU)