Frozen bovine preputial mucus as a suitable sample for the direct molecular diagnosis of Campylobacter fetus subsp. venerealis

Carli, Sílvia De; Saggin, Bianca Fagundes; Dias, Maria Eduarda; Lopes, Cassiane Elisabete; Dalto, André Gustavo Cabrera; Rolim, Verônica Machado; Zucco, Renato; Pastre, Giovani Batista; Borges, João Baptista; Siqueira, Franciele Maboni

doi:10.1016/j.mimet.2020.106101

Cited by 5 publications

(9 citation statements)

References 13 publications

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“…venerealis because they are independent of bacterial viability. 9,16 Moreover, the frequency of C. fetus subsp. venerealis that we observed is higher than previous reports from herds with an infertility history in other South American countries, such as Uruguay and Argentina.…”

Section: Discussionmentioning

confidence: 99%

“…A soft brush coupled to a pipette was inserted into the animal’s prepuce, and mucus was collected by scraping the preputial and penile mucosa, as described previously. 9 After collection, the mucus and brush were placed in 1.5-mL tubes with 1 mL of buffered PBS, pH 7.0. The samples were stored in a cool box with ice and sent to the laboratory within 48 h. Samples collected in farms far away from the laboratory were frozen at −20°C for up to 10 d, and then sent to the laboratory as standardized previously by our group.…”

Section: Methodsmentioning

confidence: 99%

“…The samples were stored in a cool box with ice and sent to the laboratory within 48 h. Samples collected in farms far away from the laboratory were frozen at −20°C for up to 10 d, and then sent to the laboratory as standardized previously by our group. 9 DNA extraction was performed directly from the collected mucus (Biopur mini spin plus; Biometrix Diagnóstica), according to the manufacturer’s instructions. DNA was stored at −20°C until use.…”

Section: Methodsmentioning

confidence: 99%

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Survey of beef bulls in Brazil to assess their role as source of infectious agents related to cow infertility

et al. 2021

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Poor reproductive performance in beef cattle caused by infectious agents results in major financial losses as a result of reduced pregnancy rates and extended calving intervals. Bulls can be subclinical chronic carriers of bacterial and protozoal agents involved in cow infertility, such as Campylobacter fetus subsp. venerealis, Ureaplasma diversum, Mycoplasma bovigenitalium, Mycoplasma bovis, and Tritrichomonas foetus. Bulls harbor these microorganisms in their preputial crypts and transmit the agents to cows during natural mating. To obtain an overview of the etiologic agents in the preputial mucus of bulls, we aimed to identify, by PCR assay, C. fetus subsp. venerealis, M. bovis, U. diversum, M. bovigenitalium, and T. foetus in Brazilian bulls from farms with high infertility rates. We collected preputial mucus from 210 bulls on 18 beef cattle farms in Brazil between 2019 and 2020. We found at least one of the infectious agents that we were studying in bulls on 16 of the 18 beef cattle farms tested. We detected at least one infectious agent from 159 of 210 (76%) bulls tested, namely C. fetus subsp. venerealis, M. bovis, U. diversum, M. bovigenitalium, and T. foetus in 87 (55%), 84 (53%), 45 (28%), 28 (18%), and 1 (0.6%) animal, respectively. We found 95 bulls (60%) positive for only 1 etiologic agent (single infection) and 64 bulls (40%) carried multiple agents. Our results demonstrate the occurrence of bacterial and protozoal infectious agents that may be related to infertility in Brazilian beef cattle herds.

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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Survey of beef bulls in Brazil to assess their role as source of infectious agents related to cow infertility

et al. 2021

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“…Positive samples were further tested for subspecies identification, using primer sets for Cff and Cfv (Table 1). PCR assays were performed in 12 µL reactions, using 1 U Taq DNA polymerase 5 , Taq DNA Buffer 10x, 50 mM of MgCl 2 , 1 mM of dNTP, 10 pmol of each primer, and 50 ng of DNA. Amplification conditions comprised: an initial denaturation at 95°C for 3 min; followed by 35 cycles of 95°C for 20 s, melting temperature as indicated in Table 1 for 20 s, and 72°C for 2 min; and a final exten-sion step at 72°C for 10 min.…”

Section: Dna Isolation From Abomasum Content Andmentioning

confidence: 99%

Campylobacter fetus in Abomasal Fluid from Spontaneously Aborted Bovine and Ovine Fetuses

Breyer

Dias²,

Henker³

et al. 2021

Acta Scientiae. Vet.

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Background: Pregnancy losses are a major concern in livestock industry due to their economic impact on producers. Campylobacter fetus subspecies fetus (Cff) and C. fetus subspecies venerealis (Cfv) are directly related to reproductive failures in ruminants. Cff colonizes the gastrointestinal tract of a wide range of hosts leading to abortion, while Cfv is restricted to genital tract being generally associated to infertility in bovine. Considering the great economic losses related to campylobacteriosis in cattle and ovine herds, this study aims to investigate the occurrence of C. fetus, considering Cff and Cfv subspecies, in bovine and ovine spontaneously aborted fetuses in state of Rio Grande do Sul, Brazil.Materials, Methods & Results: In this study, samples of abomasal fluid collected from 30 spontaneously aborted bovine (n = 18) and ovine (n = 12) fetuses were investigated for the detection of Campylobacter fetus throughout conventional PCR. Positive fetuses for C. fetus presence were further analyzed by molecular assays for Cff and Cfv detection, in order to determine subspecies identification. When available, samples of the main organs of the thoracic and abdominal cavities, as well as the brain, skeletal muscle, eyelid, skin, and placenta were collected for further histopathological analyses and bacterial culture, aiming to assess the presence of infection lesions and pathogens in those sites, respectively. Additionally, RT-qPCR assays were also performed for the detection of ruminant pestivirus, in order to detect bovine viral diarrhea cases. Throughout the present methodology, C. fetus was detected in the abomasal fluid samples of 2 bovine fetuses, being both identified as Cfv subspecies by PCR. Histopathological analyses demonstrated that macroscopic and microscopic changes found in the Cfv-positive animals were not either specific or directly related to Campylobacter infections. Moreover, no significant bacterial growth was observed in microbiological culture from the collected tissues, and both fetuses were negative for ruminant pestivirus. Differently, there was no detection of C. fetus in any of the analyzed ovine fetuses.Discussion: Considering that abortion diagnosis rates reported in cattle and sheep industry are highly variable among the published studies, and that abortion diagnoses are commonly inconclusive due to difficulties in sampling methodology and inadequate identification of the pathogen involved, it is important to investigate the etiological causes of abortion the herds for better understanding the causes of pregnancy issues and monitoring their occurrence. In addition, the absence of pathognomonic lesions in the tissues investigated in the histopathological analyses observed in this study strongly suggests that well-known etiological agents commonly associated to abortion, such as Leptospira spp., Toxoplasma spp., Chlamydia spp. and Neospora caninum, are unlikely to be the cause of infection of the analyzed fetuses. Taking this into account, the presence of C. fetus in the abomasal fluid samples from two bovine fetuses demonstrated in the present study suggests the possible association of Cfv not only with infertility, but also with cases of bovine abortion, highlighting the importance of investigating unusual causal agents of abortions in sheep and cattle. Overall, an adequate diagnosis is essential for establishing better prevention strategies to avoid the circulation of abortion-related infectious agents in the herds.Keywords: campylobacteriosis, molecular diagnosis, venereal disease, abortion, reproductive disease.

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“…These tests include culture, direct examination, direct immunofluorescence, agglutination of cervical mucus, matrix-assisted laser desorption/ionization-time of flight mass spectrometry, and polymerase chain reaction (PCR) (Sahin et al, 2017). In recent years, PCR has been increasingly used as a diagnostic tool for the direct detection of microorganisms in samples (Van der Graaf-Van Bloois et al, 2013;Carli et al, 2020). PCR offers high sensitivity and specificity (Groff et al, 2010) and does not require extensive precautions during sample transportation, as the DNA remains intact even if the pathogen is non-viable (Botelho et al, 2018).…”

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confidence: 99%

Investigação de Campilobacteriose e Tricomonose genital em touros de propriedades rurais do Pantanal Mato-Grossense, Brasil

Soares¹,

Carvalho²,

Santana³

et al. 2023

Sem. Ci. Agr.

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Campilobacteriose genital bovina (CGB) e Tricomonose bovina (TB) são doenças infectocontagiosas de transmissão venérea, assintomáticas nos touros, sendo consideradas como importantes enfermidades causadoras de falha reprodutiva, morte embrionária ou abortamento, ocasionando perdas econômicas significativas em rebanhos bovinos infectados. CGB é causada pela bactéria Campylobacter fetus subsp. venerealis e Campylobacter fetus subsp. fetus, e TB pelo protozoário Tritrichomonas foetus. O estado de Mato Grosso é detentor do maior rebanho bovino do Brasil, envolve a região do Pantanal Mato-Grossense que possui grandes extensões de terra, com ciclo anual de enchentes e a reprodução dos animais realizada predominantemente por monta natural, condições estas, favoráveis a presença de CGB e TB no rebanho. Considerando a carência de informações recentes sobre a ocorrência dessas enfermidades no estado de Mato Grosso, o objetivo deste estudo foi investigar a presença de Campylobacter spp. e Tritrichomonas foetus em 100 touros provenientes dos municípios de Poconé, Santo Antônio de Leverger e Nossa Senhora do Livramento, localizados na região pantaneira do estado de Mato Grosso. Amostras de esmegma prepucial foram coletadas por meio de escarificação via swab prepucial e armazenadas em solução salina a -80ºC. Para a detecção de Campylobacter fetus subsp. venerealis, Campylobacter fetus subsp. fetus, e Tritrichomonas foetus, foi realizada a reação em cadeia pela polimerase (PCR). Apesar do questionário aplicado nas propriedades revelar condições epidemiológicas que favorecem a manutenção e disseminação desses patógenos, este estudo não identificou a presença dos referidos agentes em touros avaliados nas propriedades rurais do pantanal Mato-Grossense.

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Frozen bovine preputial mucus as a suitable sample for the direct molecular diagnosis of Campylobacter fetus subsp. venerealis

Cited by 5 publications

References 13 publications

Survey of beef bulls in Brazil to assess their role as source of infectious agents related to cow infertility

Survey of beef bulls in Brazil to assess their role as source of infectious agents related to cow infertility

Campylobacter fetus in Abomasal Fluid from Spontaneously Aborted Bovine and Ovine Fetuses

Investigação de Campilobacteriose e Tricomonose genital em touros de propriedades rurais do Pantanal Mato-Grossense, Brasil

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