Antimicrobial resistance of Staphylococcus spp. from small ruminant mastitis in Brazil

The study aimed to determine the antimicrobial resistance patterns and to identify molecular resistance markers in Staphylococcus spp. (n=210) isolated from small ruminant mastitis in Brazil. The antimicrobial resistance patterns were evaluated by the disk diffusion test and by detection of the presence of mecA, blaZ, ermA, ermB, ermC and msrA genes by PCR. The efflux pump test was performed using ethidium bromide and biofilm production was determined by Congo red agar test along with PCR for detection of the icaD gene. The isolates were most resistant to amoxicillin (50.0%), streptomycin (42.8%), tetracycline (40.4%), lincomycin (39.0%) and erythromycin (33.8%). Pan-susceptibility to all tested drugs was observed in 71 (33.8%) isolates and 41 Staphylococcus isolates were positive for the efflux pump. Although phenotypic resistance to oxacillin was observed in 12.8% of the isolates, none harbored the mecA gene. However, 45.7% of the isolates harbored blaZ indicating that beta-lactamase production was the main mechanism associated with staphylococci resistance to beta-lactams in the present study. The other determinants of resistance to antimicrobial agents ermA, ermB, ermC, and msrA were observed in 1.4%, 10.4%, 16.2%, and 0.9% of the isolates, respectively. In addition, the icaD gen was detected in 32.9% of the isolates. Seventy three isolates (54 from goats and 19 from sheep) were negative for all resistance genes tested and 69 isolates presented two or more resistance genes. Association among blaZ, ermA, ermB, ermC and efflux pump were observed in 17 isolates, 14 of which originated from goats and three from sheep. The data obtained in this study show the resistance of the isolates to beta-lactamics, which may be associated with the use of antimicrobial drugs without veterinary control.

Etiology, antimicrobial susceptibility profile of Staphylococcus spp. and risk factors associated with bovine mastitis in the states of Bahia and Pernambuco

The purpose of this paper was to study the etiology of mastitis, determine the antimicrobial susceptibility profile of Staphylococcus spp. and to identify the risk factors associated with infection in dairy cows in the states of Bahia and Pernambuco, Brazil. From the 2,064 milk samples analyzed, 2.6% were associated with cases of clinical mastitis and 28.2% with subclinical mastitis. In the microbiological culture, Staphylococcus spp. (49.1%) and Corynebacterium spp. (35.3%) were the main agents found, followed by Prototheca spp. (4.6%) and Gram negative bacilli (3.6%). In the antimicrobial susceptibility testing, all 218 Staphylococcus spp. were susceptible to rifampicin and the least effective drug was amoxicillin (32.6%). Multidrug resistance to three or more drugs was observed in 65.6% of Staphylococcus spp. The risk factors identified for mastitis were the extensive production system, not providing feed supplements, teat drying process, not disinfecting the teats before and after milking, and inadequate hygiene habits of the milking workers. The presence of multiresistant isolates in bovine milk demonstrates the importance of the choice and appropriate use of antimicrobial agents. Prophylactic and control measures, including teat antisepsis and best practices for achieving hygienic milking should be established in order to prevent new cases of the disease in herds.

Effects of environmental modification on mastitis occurrence and hormonal changes in Holstein cows

The purpose of this research was to evaluate the effects of evaporative cooling in freestall on mastitis occurrence, milk production, and composition, as well as cortisol, T3 (triiodothyronine), and T4 (thyroxin) levels in lactating dairy cows. Twenty-eight multiparous cows averaging 70 ± 10 day postpartum were used in four treatments from January to March 2003. The treatments were: Day (cooling from 7:00 a.m. to 7:00 p.m.); Night (cooling from 7:00 p.m. to 7:00 a.m.); 24-hour (cooling 24-hour); and Control (no cooling). Wired cup test was used for clinical mastitis diagnosis, and the California Mastitis Test (CMT) was used to identify subclinical mastitis. Blood and milk samples were taken weekly for microbiological and hormonal analyses. The cortisol levels were higher than normal values in all treatment groups, suggesting stress conditions, but T3 and T4 levels remained normal in all groups. The occurrence of subclinical mastitis was lower in Day and Night groups than in Control and 24-hour groups. Regarding the microbiological analyses, in all groups the isolation of Corynebacterium sp. from milk samples increased while negative coagulase staphylococci (CNS) declined as etiological agents of subclinical mastitis. However, in Day and 24-hour groups, coagulase positive staphylococci (CPS) increased mainly Staphylococcus aureus (49.8% and 47.7% respectively). The Night group showed a decrease in subclinical mastitis occurrences. Our data indicate that all animals subjected to treatments presented high levels of cortisol, indicating a stress condition. The Night treatment presented a reduction in microbial isolation, suggesting a reduced susceptibility to mastitis.

NaOCl effect on biofilm produced by Staphylococcus aureus isolated from the milking environment and mastitis infected cows

Biofilms constitute a physical barrier, protecting the encased bacteria from detergents and sanitizers. The objective of this work was to analyze the effectiveness of sodium hypochlorite (NaOCl) against strains of Staphylococcus aureus isolated from raw milk of cows with subclinical mastitis and Staphylococcus aureus isolated from the milking environment (blowers and milk conducting tubes). The results revealed that, in the presence of NaOCl (150ppm), the number of adhered cells of the twelve S. aureus strains was significantly reduced. When the same strains were evaluated in biofilm condition, different results were obtained. It was found that, after a contact period of five minutes with NaOCl (150ppm), four strains (two strains from milk , one from the blowers and one from a conductive rubber) were still able to grow. Although with the increasing contact time between the bacteria and the NaOCl (150ppm), no growth was detected for any of the strains. Concerning the efficiency of NaOCl on total biofilm biomass formation by each S. aureus strain, a decrease was observed when these strains were in contact with 150 ppm NaOCl for a total period of 10 minutes. This study highlights the importance of a correct sanitation protocol of all the milk processing units which can indeed significantly reduce the presence of microorganisms, leading to a decrease of cow´s mastitis and milk contamination.

Beta-lactamase detection in Staphylococcus aureus and coagulase-negative Staphylococcus isolated from bovine mastitis

The objectives of the study were to evaluate the presence/production of beta-lactamases by both phenotypic and genotypic methods, verify whether results are dependent of bacteria type (Staphylococcus aureus versus coagulase-negative Staphylococcus - CNS) and verify the agreement between tests. A total of 200 bacteria samples from 21 different herds were enrolled, being 100 CNS and 100 S. aureus. Beta-lactamase presence/detection was performed by different tests (PCR, clover leaf test - CLT, Nitrocefin disk, and in vitro resistance to penicillin). Results of all tests were not dependent of bacteria type (CNS or S. aureus). Several S. aureus beta-lactamase producing isolates were from the same herd. Phenotypic tests excluding in vitro resistance to penicillin showed a strong association measured by the kappa coefficient for both bacteria species. Nitrocefin and CLT are more reliable tests for detecting beta-lactamase production in staphylococci.

Occurrence of enterotoxin-encoding genes in Staphylococcus aureus causing mastitis in lactating goats

Staphylococcal enterotoxins are the leading cause of human food poisoning worldwide. Staphylococcus spp. are the main mastitis-causing agents in goats and frequently found in high counts in goat milk. This study aimed to investigate the occurrence of enterotoxin-encoding genes in Staphylococcus aureus associated with mastitis in lactating goats in Paraiba State, Brazil. Milk samples (n=2024) were collected from 393 farms. Staphylococcus aureus was isolated in 55 milk samples. Classical (sea, seb, sec, sed, see) and novel (seg, seh, sei) enterotoxin-encoding genes were investigated by means of polymerase chain reaction (PCR). From thirty-six tested isolates, enterotoxin-encoding genes were detected in 7 (19.5%) S. aureus. The gene encoding enterotoxin C (seC) was identified in six isolates, while seiwas observed in only one isolate. The genes sea, seb, sed, see, seg and seh were not observed amongst the S. aureus investigated in this study. In summary, S. aureus causing mastitis in goats can harbor enterotoxin-encoding genes and seC was the most frequent gene observed amongst the investigated isolates. This finding is important for surveillance purposes, since enterotoxin C should be investigated in human staphylococcal food poisoning outbreaks caused by consumption of goat milk and dairy products.

Risk factors associated with the antimicrobial resistance of Staphylococcus aureus isolated from bovine mastitis

The objective of this study was to evaluate herd management practices and mastitis treatment procedures as risk factors associated with Staphylococcus aureus antimicrobial resistance. For this study, 13 herds were selected to participate in the study to evaluate the association between their management practices and mastitis treatment procedures and in vitro antimicrobial susceptibility. A total of 1069 composite milk samples were collected aseptically from the selected cows in four different periods over two years. The samples were used for microbiological culturing of S. aureus isolates and evaluation of their antimicrobial susceptibility. A total of 756 samples (70.7%) were culture-positive, and S. aureus comprised 27.77% (n=210) of the isolates. The S. aureus isolates were tested using the disk-diffusion susceptibility assay with the following antimicrobials: ampicillin 10mg; clindamycin 2μg; penicillin 1mg; ceftiofur 30μg; gentamicin 10mg; sulfa-trimethoprim 25μg; enrofloxacin 5μg; sulfonamide 300μg; tetracycline 30μg; oxacillin 1mg; cephalothin 30μg and erythromycin 5μg. The variables that were significantly associated with S. aureus resistance were as follows: the treatment of clinical mastitis for ampicillin (OR=2.18), dry cow treatment for enrofloxacin (OR=2.11) and not sending milk samples for microbiological culture and susceptibility tests, for ampicillin (OR=2.57) and penicillin (OR=4.69). In conclusion, the identification of risk factors for S. aureus resistance against various mastitis antimicrobials is an important information that may help in practical recommendations for prudent use of antimicrobial in milk production.

Tratamiento de la mastitis crónica recurrente

Tesis (Esp. Ginecología y Obstetricia) U.A.N.L.

Comparison of 3M Petrifilm™ Staph Express, 3M Petrifilm™ Rapid Coliform and 3M Petrifilm™ Aerobic count plates with standard bacteriology of bovine milk

Mémoire numérisé par la Division de la gestion de documents et des archives de l'Université de Montréal

Evidencia actual sobre características demográficas en mastitis granulomatosa idiopática

Introducción: La mastitis granulomatosa idiopática es una enfermedad crónica benigna, rara y de etiología desconocida; tiende a confundirse con otras enfermedades debido a síntomas similares. Este estudio pretende identificar y cuantificar las características demográficas, los antecedentes ginecoobstétricos relevantes y las manifestaciones clínicas prediagnósticas de esta enfermedad Metodología: Se realizó una revisión sistemática con análisis agrupado de datos tipo meta análisis. Se utilizó una estrategia de búsqueda en PubMed. Todos los estudios relacionados con la definición, manifestaciones clínicas, diagnóstico, tratamiento y pronóstico de la mastitis granulomatosa idiopática fueron elegibles. Las variables de interés fueron edad, país, antecedente de contracepción hormonal, tiempo de evolución, tiempo desde el último embarazo, diagnóstico inicial, y manifestaciones clínicas previas a la consulta. No hubo restricción en fechas de publicación. Resultados: Fueron incluidas 641 mujeres con diagnóstico de MGI reportadas en 68 publicaciones que cumplieron los criterios de selección. La edad media fue 35.9 años, 14.1% de ellas estaba embarazada o lactando, el antecedente de consumo de anticonceptivos hormonales fue 21% y el tiempo promedio desde el último parto fue de 3.9 años. La afectación ocurre principalmente en mama izquierda y en cuadrante superoexterno. El cáncer de mama y el absceso mamario son diagnósticos diferenciales en la consulta. Discusión: El diagnóstico de MGI es un reto para el ginecólogo desde la consulta inicial. Debido a que sus manifestaciones clínicas no son específicas, su diagnóstico parece apuntar a la necesidad de un proceso de descarte de otras patologías más frecuentes e incluso de peor pronóstico. Palabras clave mastitis granulomatosa idiopática

The effects of different knowledge-dissemination interventions on the mastitis knowledge of Tanzanian smallholder dairy farmers

We developed three different knowledge-dissemination methods for educating Tanzanian smallholder farmers about mastitis in their dairy cattle. The effectiveness of these methods (and their combinations) was evaluated and quantified using a randomised controlled trial and multilevel statistical modelling. To our knowledge, this is the first study that has used such techniques to evaluate the effectiveness of different knowledge-dissemination interventions for adult learning in developing countries. Five different combinations of knowledge-dissemination method were compared: 'diagrammatic handout' ('HO'), 'village meeting' ('VM'), 'village meeting and video' ('VM + V), 'village meeting and diagrammatic handout' ('VM + HO') and 'village meeting, video and diagrammatic handout' ('VM + V + HO'). Smallholder dairy farmers were exposed to only one of these interventions, and the effectiveness of each was compared to a control ('C') group, who received no intervention. The mastitis knowledge of each farmer (n = 256) was evaluated by questionnaire both pre- and post-dissemination. Generalised linear mixed models were used to evaluate the effectiveness of the different interventions. The outcome variable considered was the probability of volunteering correct responses to mastitis questions post-dissemination, with 'village' and 'farmer' considered as random effects in the model. Results showed that all five interventions, 'HO' (odds ratio (OR) = 3.50, 95% confidence intervals (CI) = 3.10, 3.96), 'VM + V + HO' (OR = 3.34, 95% CI = 2.94, 3.78), 'VM + HO, (OR=3.28, 95% CI=2.90, 3.71), WM+V (OR=3.22, 95% CI=2.84, 3.64) and 'VM' (OR = 2.61, 95% CI = 2.31, 2.95), were significantly (p < 0.0001) more effective at disseminating mastitis knowledge than no intervention. In addition, the 'VM' method was less effective at disseminating mastitis knowledge than other interventions. Combinations of methods showed no advantage over the diagrammatic handout alone. Other explanatory variables with significant positive associations on mastitis knowledge included education to secondary school level or higher, and having previously learned about mastitis by reading pamphlets or attendance at an animal-health course. (c) 2005 Elsevier B.V. All rights reserved.

Clinical and subclinical mastitis in smallholder dairy farms in Tanzania: Risk, intervention and knowledge transfer

In a cross-sectional study of 400 randomly selected smallholder dairy farms in the Tanga and Iringa regions of Tanzania, 14.2% (95% confidence interval (CI) = 11.6-17.3) of cows had developed clinical mastitis during the previous year. The point prevalence of subclinical mastitis, defined as a quarter positive by the California Mastitis Test (CMT) or by bacteriological culture, was 46.2% (95% Cl = 43.6-48.8) and 24.3% (95% Cl = 22.2-26.6), respectively. In a longitudinal disease study in Iringa, the incidence of clinical mastitis was 31.7 cases per 100 cow-years. A randomised intervention trial indicated that intramammary antibiotics significantly reduced the proportion of bacteriologically positive quarters in the short-term (14 days post-infusion) but teat dipping had no detectable effect on bacteriological infection and CMT positive quarters. Other risk and protective factors were identified from both the cross-sectional and longitudinal included animals with Boran breeding (odds ratio (OR) = 3,40, 95% CI = 1.00-11.57, P < 0.05 for clinical mastitis, and OR = 3.51, 95% CI = 1.299.55, P < 0.01 for a CMT positive quarter), while the practice of residual calf suckling was protective for a bacteriologically positive quarter (OR = 0.63, 95% Cl = 0.48-0.81, P <= 0.001) and for a CMT positive quarter (OR = 0.69, 95% Cl = 0.63-0.75, P < 0.001). A mastitis training course for farmers and extension officers was held, and the knowledge gained and use of different methods of dissemination were assessed over time. In a subsequent randomised controlled trial, there were strong associations between knowledge gained and both the individual question asked and the combination of dissemination methods (village meeting, video and handout) used. This study demonstrated that both clinical and subclinical mastitis is common in smallholder dairying in Tanzania, and that some of the risk and protective factors for mastitis can be addressed by practical management of dairy cows following effective knowledge transfer. (c) 2006 Elsevier B.V. All rights reserved.

Clinical and subclinical mastitis in smallholder dairy farms in Tanzania: Risk, intervention and knowledge transfer

In a cross-sectional study of 400 randomly selected smallholder dairy farms in the Tanga and Iringa regions of Tanzania, 14.2% (95% confidence interval (CI) = 11.6-17.3) of cows had developed clinical mastitis during the previous year. The point prevalence of subclinical mastitis, defined as a quarter positive by the California Mastitis Test (CMT) or by bacteriological culture, was 46.2% (95% Cl = 43.6-48.8) and 24.3% (95% Cl = 22.2-26.6), respectively. In a longitudinal disease study in Iringa, the incidence of clinical mastitis was 31.7 cases per 100 cow-years. A randomised intervention trial indicated that intramammary antibiotics significantly reduced the proportion of bacteriologically positive quarters in the short-term (14 days post-infusion) but teat dipping had no detectable effect on bacteriological infection and CMT positive quarters. Other risk and protective factors were identified from both the cross-sectional and longitudinal included animals with Boran breeding (odds ratio (OR) = 3,40, 95% CI = 1.00-11.57, P < 0.05 for clinical mastitis, and OR = 3.51, 95% CI = 1.299.55, P < 0.01 for a CMT positive quarter), while the practice of residual calf suckling was protective for a bacteriologically positive quarter (OR = 0.63, 95% Cl = 0.48-0.81, P <= 0.001) and for a CMT positive quarter (OR = 0.69, 95% Cl = 0.63-0.75, P < 0.001). A mastitis training course for farmers and extension officers was held, and the knowledge gained and use of different methods of dissemination were assessed over time. In a subsequent randomised controlled trial, there were strong associations between knowledge gained and both the individual question asked and the combination of dissemination methods (village meeting, video and handout) used. This study demonstrated that both clinical and subclinical mastitis is common in smallholder dairying in Tanzania, and that some of the risk and protective factors for mastitis can be addressed by practical management of dairy cows following effective knowledge transfer. (c) 2006 Elsevier B.V. All rights reserved.

Prevalence of subclinical mastitis and associated risk factors in smallholder dairy cows in Tanzania

A cross-sectional study was carried out on 200 randomly selected farms in each of the Iringa and Tanga regions of Tanzania to estimate the prevalence and risk factors for subclinical mastitis in dairy cows kept by smallholders. Subclinical mastitis was assessed using the California mastitis test (CMT), and by the bacteriological culture of 1500 milk samples collected from 434 clinically normal cows. The percentages of the cows (and quarters) with subclinical mastitis were 75.9 per cent (46.2 per cent) when assessed by the CMT and 43.8 per cent (24.3 per cent) when assessed by culture. Factors significantly associated with an increased risk of a CMT-positive quarter were Boran breed (odds radio [OR]=3.51), a brought-in cow (rather than homebred) (OR=2.39), peak milk yield, and age. The stripping method of hand milking was associated with a significantly lower prevalence Of CMT-positive quarters (OR=0.51). The CMT-positive cows were more likely to be culture positive (OR=4.51), as were brought-in (OR=2.10) and older cows.

Streptococcus equi subsp ruminatorum subsp nov., isolated from mastitis in small ruminants

Six isolates of an unknown Gram-positive, catalase-negative, chain-forming, coccus-shaped organism isolated from ovine and caprine mastitis were characterized by phenotypic and molecular taxonomic methods. On the basis of cellular morphology and the results of biochemical tests, the organism was tentatively identified as a streptococcal species. Comparative 16S rRNA gene sequencing studies confirmed that the organism is a member of the genus Streptococcus, with Streptococcus equi as its closest phylogenetic relative (98(.)8% similarity). DNA-DNA pairing studies showed that the unidentified organism displayed more than 70% relatedness to the type strains of S. equi subsp. equi and subsp. zooepidemicus. Despite the relatively high DNA-DNA reassociation values, biotyping and ribotyping allowed clear differentiation of the unknown bacterium from the two recognized subspecies of S. equi. On the basis of phenotypic and molecular genetic evidence, it is proposed that the unknown Streptococcus isolates from ovine and caprine mastitis be classified as a novel subspecies, Streptococcus equi subsp. ruminatorum subsp. nov. The type strain is CECT 5772(T) (=CCUG 47520(T) = Mt 167(T)).