RESUMO: A mastite bovina é a doença mais impactante na indústria de laticínios e é caracterizada por uma complexidade de agentes causais, os quais têm revelado uma variação geográfica entre regiões e países. Os patógenos relacionados à mastite têm sido tradicionalmente classificados como contagiosos ou ambientais, com base na fonte de infecção ou origem e nas vias de transmissão. Além disso, a gravidade das infecções mamárias tem sido associada à virulência dos patógenos, e ao estado imunológico e nutricional dos animais. Considerando esse cenário, investigamos os agentes etiológicos, os escores de gravidade clínica e os dados de dias em lactação (DEL) em 4.273 casos clínicos de mastite bovina em dez fazendas de grande porte, quatro localizadas no sudoeste do estado de São Paulo e seis no sul do Estado de Minas Gerais, Brasil. Streptococcus dysgalactiae (283/4.273 = 6,6%), Escherichia coli (190/4.273 = 4,4%), Prototheca spp. (112/4.273 = 2,6%) e Streptococcus uberis (95/4.273 = 2,2%) foram os patógenos predominantes isolados. Entre os 4.273 casos clínicos, a pontuação de gravidade clínica estava disponível em 43,8% (1.871/4.273) dos animais. Desses, 69,8% (1.306/1.871), 27,3% (510/1.871) e 2,9% (55/1.871) foram classificados como leves, moderados e graves, respectivamente. Os patógenos foram observados principalmente em amostras obtidas durante os primeiros 100 dias em lactação, e a gravidade clínica foi principalmente classificada como leve. Nossos resultados contribuem para a identificação etiológica, a pontuação de gravidade clínica e os aspectos de ordenha de vacas com mastite em fazendas leiteiras com histórico de infecções mamárias clínicas. RESUMO causais países ambientais transmissão disso animais cenário etiológicos DEL (DEL 4273 4 273 4.27 porte Gerais Brasil 283/4.273 2834273 283 (283/4.27 6,6%, 66 6,6% , 6 6,6%) 190/4.273 1904273 190 (190/4.27 4,4%, 44 4,4% 4,4%) spp 112/4.273 1124273 112 (112/4.27 2,6% 26 2 95/4.273 954273 95 (95/4.27 2,2% 22 isolados 438 43 8 43,8 1.871/4.273 18714273 1 871 (1.871/4.273 Desses 698 69 69,8 1.306/1.871, 13061871 1.306/1.871 306 (1.306/1.871) 27 3 27,3 510/1.871 5101871 510 (510/1.871 29 9 2,9 55/1.871 551871 55 (55/1.871 leves graves respectivamente 10 leve etiológica clínicas 427 4.2 283/4.27 283427 28 (283/4.2 6,6 190/4.27 190427 19 (190/4.2 4,4 112/4.27 112427 11 (112/4.2 2,6 95/4.27 95427 (95/4.2 2,2 43, 1.871/4.27 1871427 87 (1.871/4.27 69, 1306187 1.306/1.87 30 (1.306/1.871 27, 510/1.87 510187 51 (510/1.87 2, 55/1.87 55187 5 (55/1.87 42 4. 283/4.2 28342 (283/4. 6, 190/4.2 19042 (190/4. 4, 112/4.2 11242 (112/4. 95/4.2 9542 (95/4. 1.871/4.2 187142 (1.871/4.2 130618 1.306/1.8 (1.306/1.87 510/1.8 51018 (510/1.8 55/1.8 5518 (55/1.8 283/4. 2834 (283/4 190/4. 1904 (190/4 112/4. 1124 (112/4 95/4. 954 (95/4 1.871/4. 18714 (1.871/4. 13061 1.306/1. (1.306/1.8 510/1. 5101 (510/1. 55/1. 551 (55/1. 283/4 (283/ 190/4 (190/ 112/4 (112/ 95/4 (95/ 1.871/4 1871 (1.871/4 1306 1.306/1 (1.306/1. 510/1 (510/1 55/1 (55/1 283/ (283 190/ (190 112/ (112 95/ (95 1.871/ 187 (1.871/ 130 1.306/ (1.306/1 510/ (510/ 55/ (55/ (28 (19 (11 (9 1.871 18 (1.871 13 1.306 (1.306/ (510 (55 (2 (1 ( 1.87 (1.87 1.30 (1.306 (51 (5 1.8 (1.8 1.3 (1.30 1. (1. (1.3

ABSTRACT: Bovine mastitis is the most impacting disease of dairy industry, and it is characterized by a complexity of causal agents, which have revealed a geographical variation among regions and countries. The mastitis-related pathogens have been traditionally classified as contagious or environmental, based on habits of the microorganisms and transmission routes. Besides, the severity of mammary infections has been associated with the virulence of the pathogens, and immune and nutritional status of the hosts. Considering this scenario, we investigated the etiological pattern, clinical severity scores, and days in milk (DIM) data in 4,273 clinical cases of bovine mastitis among ten large-dairy farms located in Brazil. Streptococcus dysgalactiae (283/4,273 = 6.6%), Escherichia coli (190/4,273 = 4.4%), Prototheca spp. (112/4,273 = 2.6%), and Streptococcus uberis (95/4,273 = 2.2%) were the predominant pathogens isolated. Among 4,273 clinical cases, clinical severity score was available in 43.8% (1,871/4,273) animals. From these, 69.8% (1,306/1,871), 27.3% (510/1,871), and 2.9% (55/1,871) were scored as mild, moderate, and severe, respectively. Pathogens were observed mainly in samples obtained during the first 100 days in milk, and clinical severity scored mainly as mild. Our results contributed to the etiological identification, clinical severity scoring, and milking aspects of mastitic cows in dairy farms with a history of clinical mammary infections. ABSTRACT industry agents countries mastitisrelated related environmental routes Besides hosts scenario pattern scores DIM (DIM 4273 4 273 4,27 largedairy large Brazil 283/4,273 2834273 283 (283/4,27 6.6%, 66 6.6% , 6 6.6%) 190/4,273 1904273 190 (190/4,27 4.4%, 44 4.4% 4.4%) spp 112/4,273 1124273 112 (112/4,27 2.6%, 26 2.6% 2 2.6%) 95/4,273 954273 95 (95/4,27 2.2% 22 isolated 438 43 8 43.8 1,871/4,273 18714273 1 871 (1,871/4,273 animals these 698 69 69.8 1,306/1,871, 13061871 1,306/1,871 306 (1,306/1,871) 27 3 27.3 510/1,871, 5101871 510/1,871 510 (510/1,871) 29 9 2.9 55/1,871 551871 55 (55/1,871 mild moderate severe respectively 10 identification scoring 427 4,2 283/4,27 283427 28 (283/4,2 6.6 190/4,27 190427 19 (190/4,2 4.4 112/4,27 112427 11 (112/4,2 2.6 95/4,27 95427 (95/4,2 2.2 43. 1,871/4,27 1871427 87 (1,871/4,27 69. 1306187 1,306/1,87 30 (1,306/1,871 27. 510187 510/1,87 51 (510/1,871 2. 55/1,87 55187 5 (55/1,87 42 4, 283/4,2 28342 (283/4, 6. 190/4,2 19042 (190/4, 4. 112/4,2 11242 (112/4, 95/4,2 9542 (95/4, 1,871/4,2 187142 (1,871/4,2 130618 1,306/1,8 (1,306/1,87 51018 510/1,8 (510/1,87 55/1,8 5518 (55/1,8 283/4, 2834 (283/4 190/4, 1904 (190/4 112/4, 1124 (112/4 95/4, 954 (95/4 1,871/4, 18714 (1,871/4, 13061 1,306/1, (1,306/1,8 5101 510/1, (510/1,8 55/1, 551 (55/1, 283/4 (283/ 190/4 (190/ 112/4 (112/ 95/4 (95/ 1,871/4 1871 (1,871/4 1306 1,306/1 (1,306/1, 510/1 (510/1, 55/1 (55/1 283/ (283 190/ (190 112/ (112 95/ (95 1,871/ 187 (1,871/ 130 1,306/ (1,306/1 510/ (510/1 55/ (55/ (28 (19 (11 (9 1,871 18 (1,871 13 1,306 (1,306/ (510/ (55 (2 (1 ( 1,87 (1,87 1,30 (1,306 (510 (5 1,8 (1,8 1,3 (1,30 (51 1, (1, (1,3

Abstract Background: The early symptoms of leptospirosis and dengue fever are difficult to distinguish and can cause diagnostic confusion. Due to the large dengue epidemics that has occurred in Brazil in recent years, it is possible that cases of leptospirosis were unreported. Therefore, we performed a retrospective study to detect leptospirosis in patients who were tested for dengue, but whose laboratory diagnoses were negative. Methods: Sera samples from 2,017 patients from 48 cities located in the central region of São Paulo state, Brazil, were studied. All samples were subjected to the microscopic agglutination test (MAT), 305 of which were taken from patients five days or less since the onset of symptoms, and were additionally subjected to real-time polymerase chain reaction (PCR). Results: The overall prevalence of leptospirosis cases was 21 (1.04%), with 20 through MAT (18 for Icterohaemorrhagiae and two for the Cynopteri serogroup) and one through PCR (amplicon sequencing compatible with Leptospira interrogans). According to previously established criteria, eight cases of leptospirosis were classified as “confirmed” and 13 as “probable”. The Brazilian notification system for health surveillance had no records for 16 patients positive for leptospirosis and, thus, they were considered unreported cases. Statistical analyses revealed that the prevalence of leptospirosis was higher in men (1.56%) than in women (0.56%), and the mean age was higher in positive patients (43.7 years) than in negative ones (32.3 years). Conclusion: The results indicated that patients suspected of dengue fever had evidence of leptospirosis or Leptospira infection, and most of these cases were unreported in the Brazilian notification system. The high burden of dengue may contribute to the misdiagnosis of leptospirosis, and health professionals should increase their awareness of leptospirosis as an important differential diagnosis of patients with suspicion of dengue.

Abstract INTRODUCTION: Norway rats (Rattus norvegicus) are zoonotic reservoirs for Leptospira spp. and Toxoplasma gondii, and influence diseases in urban areas. METHODS: Free-ranging and laboratory-raised rats from two zoos in southern Brazil were tested for Leptospira spp. and T. gondii using microscopic agglutination and modified agglutination tests, respectively. RESULTS: Overall, 25.6% and 4.6% free-ranging rats tested positive for Leptospira spp. and T. gondii, respectively, with co-seropositivity occurring in two animals. For laboratory-raised rats, 20% tested positive for Leptospira spp. Also, Leptospira biflexa serovar Patoc and Leptospira noguchii serovar Panama were found. CONCLUSIONS: Serosurveys can show the environmental prevalence of zoonotic pathogens.

RESUMO: A mastite é considerada o maior problema dos animais destinados à produção de leite. Altera a sua composição e aumenta a contagem de células somáticas (CCS). Os micro-organismos envolvidos na doença podem ser origem infecciosa, como Staphylococcus aureus, ou ambiental, tal como Escherichia coli. A cultura bacteriana é uma ferramenta de diagnóstico e auxilia na detecção do patógeno causador da mastite. No entanto, fatores como fagocitose podem desencadear um resultado negativo. Quando estabelecido um programa de controle de mastite, o diagnóstico precoce e o início do tratamento adequado dos casos clínicos são fundamentais para se atingir os objetivos e seu sucesso, está relacionado com o patógeno envolvido. A indicação do tratamento de longa duração, ou terapia estendida, tem melhorado a resposta ao tratamento em casos de mastite por S. aureus, no entanto, com 30-50% de cura. Do ponto de vista do manejo dos animais, devido a alta contagiosidade deste patógeno, sua persistência no rebanho e custo em função ao tratamento, muitas vezes, o descarte do animal tem sido priorizado a fim de controlar os casos de mastite em propriedades. As medidas de controle são muito importantes para contribuir com a redução de casos de mastite por este patógeno. A indicação do tratamento intramamário associada com sistêmico tem poder efetivo em casos de mastite por E. coli, cujos casos agudos apresentam-se com sepse e toxemia. São abordados ainda aspectos de tratamentos alternativos das mastites, utilizados principalmente no processo orgânico de produção leiteira.

ABSTRACT: Mastites are considered a major problem on animals for milk production. Changes the milk composition and increases the somatic cell count (SCC). The microorganism involved in the disease may be infectious origin such as Staphylococcus aureus or environmental such as Escherichia coli. Bacterial culture is a diagnostic tool and aids in the detection of pathogenic causing masitis. However factor such as phagocytosis may result a negative result. When established a mastitis control program early diagnose and the initiation of appropriate treatment of clinical cases are fundamental for achieving the goals and success is related to the pathogen involved. The indication of treatment of long duration, or extended therapy has improved the response to the treatment in cases of matitis by S. aureus, however with 30-50% of cure. From the view point of handling of animals, given the high infectiousness of this pathogen, its persistence in the herd and cost-effective as a function of response to treatment often has prioritized the animal’s discard in order to control the mastitis cases in properties. Control measures are very important to contribute the reduction of cases of mastitis symptoms by this pathogen. The indication of intramammary treatment associated with systemic has power effective in cases of mastitis by E. coli, whose acute cases present with sepsis and toxemia. Also address aspects of alternative treatments of mastitis, mainly used in the organic process of milk production.

RESUMO: Além de Staphylococcus aureus atualmente outros estafilococos coagulase positiva (SCP) e estafilococos coagulase-negativos (SCN), anteriormente considerados de menor relevância, são reconhecidos como importantes patógenos para humanos e animais. O envolvimento desses micro-organismos na etiologia da mastite bovina e a possibilidade da sua transmissão através do leite aos humanos justifica a utilização de métodos confiáveis para a identificação das espécies mais frequentes. O objetivo deste estudo foi comparar as técnicas fenotípicas com o método genotípico realizada por sequenciamento do gene rpoB na identificação de espécies do gênero Staphylococcus spp. isolados de mastite bovina. Um total de 300 estafilococos isolados de casos de mastite bovina em diferentes rebanhos leiteiros brasileiros foram estudados por técnicas fenotípicas e genotípicas, respectivamente: 150 linhagens de SCP e 150 linhagens de SCN. Foram identificados um total de 18 espécies de SCN e 4 espécies SCP. Entre os SCN as seguintes espécies identificadas: 48 (32%) Staphylococcus warneri, 22 (15%) S. epidermidis, 20 (13%) S. hyicus, 10 (7%) S. xylosus, 7 (5%) S. haemolyticus, 6 (4%) S. simulans, 6 (4%) S. schleiferi subsp schleiferi, 6 (4%) S. hominis, 5 (3%) S. pasteuri, 4 (2,7%) S. cohnii, 3 (2%) S. saprophyticus subsp. saprophyticus, 3 (2%) S. chromogenes, 3 (2%) S. sciuri, 2 (1%) S. saccharolyticus, 2 (1%) S. lugdunensi, 1 (0,7%) S. auricularis, 1 (70 %) S. saprophyticus subsp. bovis, 1 (0,7%) S. capitis. E entre as 150 SCP foram identificados, 105 (70%) S. aureus, 21 (14%), S. hyicus, 19 (13%) S. intermedius e 5 (3%) S. schleiferi subsp coagulans. Considerando-se os 150 SCN isolados, as identificações realizadas por testes fenotípicos e genotípicos apresentaram 96,7% de concordância, coeficiente de concordância kappa = 0,933, SE (erro padrão) de kappa = 0,021 (95% intervalo de confiança: 0,893-0,974), coeficiente de correlação de Pearson (r) = 0,9977, (intervalo de confiança de 95%: 0,9938 a 0,9992) e em relação a 150 SCP isolados foi observado uma concordância de 98,7%, kappa = 0,960, sE de kappa = 0,016, (95% de intervalo de confiança: 0,929 a 0,992) coeficiente de correlação de Pearson (r) = 0,9994 (95% intervalo de confiança: 0,9681-1,0000). A correlação entre os métodos de identificação pode ser considerada como excelente. Esses resultados demonstraram que de acordo com os recursos disponíveis no laboratório, poderia ser utilizada qualquer uma das metodologias.

ABSTRACT: In addition to Staphylococcus aureus nowadays other coagulase-positive staphylococci (CoPS) and coagulase-negative staphylococci (CoNS), earlier considered of minor importance, are now accepted as relevant pathogens for humans and animals. The involvement of these microorganisms in bovine mastitis etiology and the possibility their transmission through milk to humans justify the requirement of developing reliable methods for identification of the most frequent species among them. The purpose of this study was to compare the phenotypic techniques with the genotypic method carried out by sequencing of the rpoB gene in identification of several species of the genus Staphylococcus isolated from bovine mastitis. A total of 300 staphylococci isolates of bovine mastitis cases from several Brazilian dairy herds were studied by phenotypic and genotypic techniques, respectively: 150 CoPS and 150 CoNS strains. A total of 18 CoNS different species and 4 CoPS species were identified. Among the CoNS the following species were recognized: 48 (32%) Staphylococcus warneri, 22(15%) S. epidermidis, 20(13%) S. hyicus, 10(7%) S. xylosus, 7(5%) S. haemolyticus, 6(4%) S. simulans, 6(4%) S. schleiferi subsp schleiferi, 6(4%) S. hominis, 5(3%) S. pasteuri, 4(2.7%) S. cohnii, 3(2%) S. saprophyticus subsp. saprophyticus 3(2%) S. chromogenes 3(2%) S. sciuri, 2(1%) S. saccharolyticus, 2(1%) S. lugdunensi, 1(0,7%) S. auricularis, 1(70%) S. saprophyticus subsp. bovis, 1(0.7%) S. capitis. And among the 150 CoPS were identified respectively: 105 (70%) S. aureus, 21(14%), S. hyicus, 19(13%) S. intermedius e 5(3%) S. schleiferi subsp coagulans. Considering the 150 CoNS isolates, the identifications performed by phenotypic and genotypic tests presented 96.7% of concordance, kappa coefficient of agreement = 0.933, SE (standard error) of kappa=0.021 (95% confidence interval: 0.893 to 0.974), Pearson’s correlation coefficient (r) = 0.9977, (confidence interval 95%: 0.9938 a 0.9992) and in relation to 150 CPS isolates it was detected an agreement of 98.7%, kappa = 0.960, SE of kappa = 0.016, (95% confidence interval: 0.929 to 0.992) Pearson’s correlation coefficient (r) = 0.9994 (95% confidence interval: 0.9681 to 1.0000). The verified agreement strength between the identification methods can be considered as excellent. These results assure that according to laboratory resources any of them will be suitable to perform the staphylococci identification.

Resumo: A mastite é a principal afecção do gado destinado à produção leiteira, que impacta significativamente a cadeia produtiva do leite, com reflexos ainda para a saúde pública. Estudou-se aspectos relacionados à etiologia, celularidade e de contagem bacteriana em 10 propriedades leiteiras, localizadas no Estado de São Paulo. Foram examinadas 1148 vacas em lactação, totalizando 4584 glândulas mamárias. Foram considerados os casos, em que houve isolamento de estafilococos coagulase positiva (SCP) e estafilococos coagulase negativa (SCN). Os resultados revelaram microbiota com vários patógenos e diferentes espécies de SCN (128 casos) e SCP (45), Staphylococcus aureus(90), Streptococcus agalactiae(70), Streptococcus dysgalactiae (69), Streptococcus uberis(29), Corynebacteriumspp. (230), Klebsiella pneumoniae(28), Klebsiella oxytoca(2), Escherichia coli(15), Enterobactersp. (3). Os resultados de contagem de células somáticas (CCS) relacionados aos SCP e SCN não mostraram diferenças entre as propriedades avaliadas, entretanto com diferenças significantes ao se avaliar a CCS entre os dois grupos de estafilococos, como pode ser evidenciado ao comparar SCN Discreto e SCP exuberante (P<0,01), SCP Discreto e SCP exuberante (P<0,001) e SCN moderado e SCP exuberante (P<0,01). A avaliação da CCS relacionada à intensidade da infecção, considerando-se como crescimento discreto o isolamento de até nove colônias, moderado de dez a 29 colônias e exuberante, com 30 ou mais colônias, revelou para ambos os grupos de estafilococos que quanto maior o número de unidades formadoras de colônias (UFC), a CCS é mais elevada, sendo sempre maior nos casos de SCP. Conclui-se que quando há maior número de UFC, há concomitantemente maior CCS/mL de leite, no caso dos SCP e SCN, o que mostra relação direta da intensidade do processo infeccioso com a resposta da celularidade do leite, bem como pela relevância desses na etiologia das mastites e dos aspectos negativos tanto para a produção, quanto na qualidade do leite produzido nas propriedades.

Abstract: Mastitis is the main affection of cattle intended for dairy production, which significantly impacts the milk production chain, with consequences yet to public health. It was studied aspects related to etiology, cellularity and bacterial count in 10 dairy farms, in the State of São Paulo. There were examined 1148 milking cows, totaling 4584 mammary glands. Cases in which there was isolation coagulase positive staphylococci (CPS) and coagulase-negative staphylococci (CNS) were considered. The results showed microbiota with various different pathogens and CNS (128 cases) and CPS (45), Staphylococcus aureus (90), Streptococcus agalactiae (70), Streptococcus dysgalactiae (69), Streptococcus uberis (29), Corynebacterium spp . (230), Klebsiella pneumoniae (28), Klebsiella oxytoca (2), Escherichia coli (15), Enterobacter spp. (3). The somatic cell count (SCC) related to CPS and CNS showed no differences among the evaluated properties, although with significant differences when evaluating the SCC between both groups of staphylococci, as evidenced by comparing discreet CNS and exuberant CPS (P <0.01), discreet CPS and exuberant CPS (P <0.001) and moderate SCN and exuberant SCP (P <0.01). The evaluation of SCC was related to the intensity of infection, considering how slight growth isolation of up to nine colonies, moderate ten to 29 colonies and lush, with 30 or more colonies, revealed for both groups of staphylococci that the higher the number of colony forming units (CFU), SCC is higher being larger in cases of CPS. The results indicate that the importance of both CPS and CNS when considering the high occurrence of the evaluated properties and the fact that it raises the SCC, which compromises the quality of milk. We conclude that when there is a larger number of CFU, is concomitantly higher SCC/ml of milk, in the case of CPS and CNS, which shows a direct relationship of the intensity of the infectious process with the response of milk cellularity, as well as the relevance of these in the etiology of mastitis and negatives aspects to both the production and the quality of milk produced in properties.