Excretion of Brucella abortus vaccine B19 strain during a reproductive cycle in dairy cows

This paper aimed to determine the excretion period of B19 vaccine strain during a complete reproductive cycle (from estrus synchronization, artificial insemination, pregnancy and until 30 days after parturition) of dairy cows from 3 to 9 years old that were previously vaccinated from 3 to 8 months. Three groups were monitored with monthly milk and urine collection during 12 months: G1 with seven cows from 3 to 4 years old; G2 with three cows from 5 to 6 years old; and G3 with four cows from 7 to 9 years old. Urine and milk samples were submitted to bacteriological culture and urine and PCR reactions for detection of Brucella spp. and PCR-multiplex for B19 strain identification. Ring test (RT) was also performed in the milk samples, and serum samples were tested by buffered acidified plate antigen test (BAPA). All animals were serologically negative at BAPA and Brucella spp. was not isolated from both urine and milk samples. RT revealed 13/210 (6.2%) positive milk samples. PCR reactions detected DNA of Brucella spp. in 86/420 (20.5%) samples. In urine it was found a significantly higher frequency (35.2%; 74/210) than in milk (5.7%; 12/210), more frequently from the estrus to 150 days of pregnancy and after parturition (6.7%; 10/150), and from 150 days of pregnancy to parturition (3.4%; 2/60), and they were all identified as B19 strain. In three groups, intermittent excretion of B19 strain was detected mainly in urine samples, which confirmed its multiplication and persistence in cows for until 9 years.

Validação de nova proposta de espermocultura quantitativa aplicada a sêmen industrializado de touros / Validation of a new proposal of quantitative sperm culture for industrialized bull semen

O Ministério da Agricultura, Pecuária e Abastecimento (MAPA) tem discutido a obrigatoriedade da realização de espermocultura em sêmen industrializado não somente para garantia da biossegurança mas também por falhas na fertilização in vitro pela contaminação dos ovócitos com bactérias ubiquitárias e oportunistas da microbiota prepucial. A técnica quantitativa de Pour Plate, preconizada pela Organização Internacional de Epizootias, é operacionalmente difícil e de custo elevado para a rotina em centrais de inseminação artificial (CIAs). Desse modo, avaliou-se a técnica de contagem de bactérias viáveis por superfície (CBVS) em UFC/mL de sêmen industrializado de touros de CIAs, comparando-a à técnica de Pour Plate, com intuito de validação. Foram empregadas palhetas de sêmen de touros do Projeto Hungria do MAPA. A partir da diluição seriada de 10-1 a 10-5 do sêmem, foram empregadas, paralelamente, as técnicas de Pour Plate e CBVS. A média ou mediana de UFC/mL obtida em cada diluição foi comparada entre as técnicas, e não foram observadas diferenças estatísticas entre as duas técnicas quanto à quantificação de bactérias em UFC/mL, sugerindo a possibilidade de substituição da técnica de Pour Plate pela de CBVS, com vantagens de praticidade e menor custo.

The Brazilian Ministry of Agriculture (MAPA) has discussed the mandatory culture of industrialized semen, both to ensure biosefety, and to prevent in vitro fertilization problems caused by oocyte contamination with ubiquitous and opportunistic bacteria from preputial microbiota. Pour plate, a quantitative technique recommended by the World Organization for Animal Health (OIE), is operationally difficult and costly for routine analysis in Artificial Insemination Centers (AICs). The objectives of this study were to evaluate and validate viable superficial bacteria counts (VSBC), in CFU/mL, compared with pour plate counts, in industrialized bull semen samples from AICs. Semen straws from Projeto Hungria - MAPA bulls were used. VSBC and pour plate were carried out in parallel in serial dilutions of the samples, from 10-1 to 10-5. CFU/mL means or medians recorded in each dilution and technique were compared, and no statistical differences were observed between the two techniques regarding the quantification of bacteria in CFU/mL, suggesting that it may be possible to replace pour plate for CBSV, a cheaper and more practical technique.

Detecção dos genes da toxina citoletal distensiva em estirpes de Campylobacter jejuni isoladas de carcaças de frangos / Detection of cytolethal distending toxin genes in strains of Campylobacter jejuni isolated from broiler carcasses

Foram analisadas 80 amostras de sobrecoxas de frangos de corte resfriados provenientes de feiras livres e hipermercados do município de São Paulo, SP. Treze estirpes de Campylobacter spp. foram isoladas em 10 (12,5 por cento) sobrecoxas, sendo cinco amostras originárias de feiras livres e cinco de hipermercados. Onze estirpes foram identificadas como Campylobacter jejuni e duas como Campylobacter coli. As 11 estirpes foram confirmadas como C. jejuni pela PCR do gene da hipuricase (hip), e destas, quatro (36,4 por cento) apresentaram os três genes (cdtA, cdtB e cdtC) codificantes da toxina citoletal distensiva pela multiplex-PCR, sendo três estirpes provenientes de hipermercados e uma de feira livre. Observou-se a presença de estirpes virulentas de C. jejuni, portadoras do complexo de genes cdt, nas amostras de frango resfriado, não só na linha de abate, mas até o ponto final da cadeia de distribuição, nos dois principais centros de venda a varejo.

Eighty samples of refrigerated broiler thighs purchased in street markets and supermarkets in the city of São Paulo, SP, were analyzed. Thirteen Campylobacter spp. strains were isolated in 10 (12.5 percent) thighs, five of them from street market samples and other five from supermarkets. Eleven strains were identified as Campylobacter jejuni and two of them as Campylobacter coli. The 11 strains were confirmed to be C. jejuni using PCR for hippuricase (hip) gene. From these, multiplex-PCR showed that four (36.4 percent) strains presented the three genes (cdtA, cdtB, and cdtC) encoding cytolethal distending toxin: three strains from supermarket and one from street market samples. These results are important, because they demonstrate the presence of virulent C. jejuni strains in refrigerated broiler thigh samples, not only in the slaughterhouse but in the final point of the distribution chain, at the two most important food retail commercer.

Influência das condições ambientais na transmissão da leptospirose entre criações de ovinos e bovinos da região de Sorocaba, SP / The influence of environment conditions on the transmission of leptospirosis between sheep and cattle herds of Sorocaba region, SP, Brazil

Leptospiras excretadas pela urina podem sobreviver por longos períodos em águas de superfície e solos, na dependência do pH e teor de umidade e de matéria orgânica. Investigou-se a influência do meio ambiente na transmissão da leptospirose em dois rebanhos exclusivos de ovinos (A e C) e dois de ovinos consorciados com bovinos (F e H) da região de Sorocaba, SP, no período de dezembro de 2007 a setembro de 2008. Foram examinadas amostras de soro pela reação de soroaglutinação microscópica; de urina, água e solo pelo cultivo para leptospiras e urina de ovinos pela PCR. Condições edafoclimáticas, pH das águas de superfície e solo, granulometria e permeabilidade do solo foram analisadas. Todos os rebanhos apresentaram pelo menos um animal sororeagente para Leptospira spp. Apenas a PCR de um pool de urina de ovinos (H) foi positiva. Leptospira spp. foi isolada do lago de F. O pH das águas de superfície variou entre 6,0-7,0; e nos solos entre 4,5 e 6,8. Os índices de matéria orgânica em A, C e H variaram de 24 a 35 g/dm3, e 63 g/dm3 em F. A composição do solo de A e F mostrou-se franco-argiloarenosa, C argilosa e H franco-siltosa; como texturas mistas são capazes de manter a umidade, principalmente devido a argila. Diante da presença de animais sororeatores e portanto da circulação de Leptospira spp. nos rebanhos, conclui-se que o ciclo de transmissão é dependente da interação sinérgica e antagônica de muitas variáveis; onde o pastejo num habitat com alto teor de umidade parece ser limitante.

Leptospires excreted by urine are able to survive for long periods in surface water and soil depending on the pH, humidity and organic matter presence. This paper reported the influence of environment conditions on the transmission of leptospirosis in two sheep-only farms (A and C) and two cattle-sheep farms (F and H) from December 2007 to September 2008. Serum samples were examined by microscopic agglutination test; urine, surface water and soil samples were cultured for leptospires, and ovine urine pools were analyzed by PCR. Regional edaphoclimatic conditions, pH of surface water and soil, granulometry and permeability of soil were analyzed. All herds presented at least one reactor to Leptospira spp. Only the PCR of an ovine urine pool of herd H was positive and Leptospira spp. was isolated from the F lake. The pH of water samples ranged from 6.0 to 7.0; while in soil it was around from 4.5 to 6.8. Soil organic matter were 24 to 35 g/dm3 in A, C e H, and 63 g/dm3 in F. Soil samples of A and F showed loamy-clay texture; C had clay soil, and H loamy-silt soil; as mixed compositions are able to maintain the humidity, mainly where clay is present. As the presence of reactors in all herds indicated the contact with Leptospira spp., it was concluded that the cycle of transmission is dependent on the synergistic and antagonistic interaction of many variables; but the close contact of animals grazing in a high humidity habitat seems to be limiting.

Use of pcr-rflp of thefla a gene for detection and subtyping of Campylobacter jejuni strains Potentially related to Guillain-barré syndrome, isolated from humans and animals

The objectives of the present study were the subtyping of Campylobacter jejuni subsp. jejuni strains obtained from humans and different animal species using PCR-RFLP, and the detection, by means of the same technique, of strains related to serotype PEN O19:LIO 7, the main C. jejuni serotype linked to Guillain-Barré Syndrome (GBS). Seventy C. jejuni strains isolated from human feces (n=33), primates (n=15), dogs (n=5), swine (n=2), bovines (n=1), abortion material from goats (n=2) and poultry carcasses (n=12), all collected in the state of São Paulo, were subtyped by means of PCR-RFLP of fla A gene, using restriction endonucleases Hae III, Afa I and Mbo I. Seven subtypes were observed when using the enzyme Hae III; eight when using Mbo I; and seven when using Afa I. The combination of the three endonucleases led to 16 fla-RFLP subtypes, from which ten subtypes shared strains of human and animal origin. From these, seven subtypes were observed in human and broiler strains. In eight subtypes, the other animal species shared patterns with human strains. It was inferred that, besides broilers, swine, goats, dogs and primates may be sources of infection for human in São Paulo. PCR-RFLP is a highly discriminatory technique that may be applied to molecular epidemiology studies of samples from different origins. Besides, the study also enabled the detection of two human strains and two primate strains related to serotype PEN O19: LIO 7.

Fatores de risco para a leptospirose em fêmeas bovinas em idade reprodutiva no Estado de São Paulo / Risk factors to leptospirosis in cows in São Paulo state, Brazil

Risk factors associated to leptospirosis were identified in cows from São Paulo State. The State was divided into seven productive circuits from which 8,216 cows older than 24 months of age from 1,021 herds were sampled. For serological diagnosis of Leptospira spp. infection, the microscopic agglutination test (MAT) was carried out. Of the investigated herds, 718 (71.3%; 95% CI = 68.5% - 74.0%) presented at least one reactant animal at MAT to any serovar. Serovar Hardjo was the most prevalent, with 55.2% (95% CI = 51.4% - 58.9%) of the positive herds. Herd size, animal purchase, share pasture, presence of ovine and swine, and utilization of artificial insemination were identified as risk factors. Utilization of maternity pens was a protective factor against leptospirosis.

Detecção de ácidos nucléicos de Brucella spp., Leptospira spp., herpesvirus bovino e vírus da diarréia viral bovina, em fetos bovinos abortados e em animais mortos no perinatal / Detection of Brucella spp., Leptospira spp., bovine herpesvirus and bovine viral diarrhea virus nucleic acids in aborted fetuses and bovines dead perinatal

Samples of 114 bovine fetuses and 10 calves, which dead in perinatal period, were examined for detection of DNA. The most common detected agent was Brucella spp. in 17 samples (13.7 percent) followed by Leptospira spp. in 4 cases (3.2 percent),bovine herpesvirus (BHV) and bovine viral diarrhea (BVDV) in 3 animals (2.4 percent) each, and 1 for the association of BVDV and BHV. In 77.4 percent (96/124) of the samples it was not possible to detect any agent.

Isolamentos bacterianos de fetos abortados bovinos examinados no Instituto Biológico de Säo Paulo, no período de 1985 a 1992 / Bacterial isolations from bovine aborted fetuses examined in Instituto Biologico of Sao Paulo State, Brazil, from 1985 to 1992

No período de 1985 a 1992, foram analisadas bacteriologicamente 544 amostras de órgäos e anexos fetais, provenientes de 282 fetos bovinos, oriundos de rebanhos, na sua maioria leiteiros, procedentes de vários estados do Brasil. Foram consideradas como possíveis causas de abortamento, as bactérias patogênicas e as culturas puras ou preponderantes de bactérias oportunistas. Excluindo-se os materiais impróprios para exame (25/282), dos 257 restantes, em 37,4 por cento foram diagnosticadas causas bacterianas, tais como: Brucella abortus (6,2 por cento), Leptospira spp (6,2 por cento), Staphylococcus aureus (5,4 por cento), Campylobacter fetus (4,7 por cento) e Streptococcus Beta hemolítico (3,5 por cento). Os focos destes agentes apresentavam-se amplamente distribuídos no Estado de Säo Paulo