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Fontes ambientais de infeção por Cryptosporidium parvum em vitelos leiteiros recém nascidos
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Advisor(s)
Abstract(s)
Cryptosporidium parvum é um protozoário parasita responsável pela infeção intestinal de humanos e animais, causando alta morbilidade e morte em hospedeiros imunocomprometidos, e é considerado uma das principais causas de diarreia em vitelos. A principal via de transmissão dos oocistos é fecal-oral. Este parasita representa um problema relevante tanto para a saúde e produção animal como para a saúde pública, uma vez que alguns genótipos são zoonóticos. O presente trabalho visou avaliar possíveis fontes de contaminação por C. parvum em vitelos numa vacaria leiteira, o que pode ajudar a desenvolver estratégias mais eficazes para prevenir e controlar a disseminação do parasita, protegendo a saúde e o bem-estar tanto de animais como de humanos. Amostras fecais de vacas adultas foram recolhidas nos períodos pré-parto (n=6) e pós parto (n=7), assim como ao parto (n=7). Foram colhidas diariamente amostras de fezes dos respetivos 7 vitelos desde o nascimento até ao 21º dia de vida. As amostras ambientais incluíram luvas, cama da maternidade, placas de Petri para recolha de aerossóis da limpeza com água a alta pressão e zaragatoas de diversos materiais como botas, jaulas, baldes, tetinas, comedouros e sondas esofágicas. A metodologia baseou-se na identificação de oocistos de Cryptosporidium spp. pela coloração de Ziehl-Neelsen e imunofluorescência direta. Foi extraído DNA das amostras ambientais e das amostras fecais dos vitelos ao 10º dia e analisado por PCR 18S rRNA para confirmar a presença de Cryptosporidium spp. Algumas jaulas pré-entrada (4/6), baldes de água (2/5), tetinas (1/5), comedouros (4/5), placas de Petri (2/7), cama da maternidade (1/1), todos os bezerros (7/7), jaulas de saída (7/7), luvas (2/2) e botas (2/2) testaram positivos. A presença de Cryptosporidium parvum foi confirmada por RFLP. A análise da sequência do gene gp60 confirmou o genótipo como IIaA15G2R1. Dado que as amostras das 7 vacas revelaram-se negativas, podem ser excluídas como uma potencial fonte de contaminação para os vitelos. No dia 6, 4 de 7 vitelos estavam positivos e ao 7º dia todos testaram positivo, sugerindo que a infeção ocorre entre os dias 0 e 4, considerando um período pré-patente de 3 a 6 dias. Cryptosporidium spp. foi detetado em todos os tipos de amostras ambientais (exceto nas sondas esofágicas), suportando a hipótese de que o problema está nas estratégias de maneio e limpeza. Nestas condições, os vitelos recém-nascidos são expostos ao parasita desde o dia 0, o que pode justificar a alta prevalência da infeção e doença em vacarias leiteiras
ABSTRACT - Cryptosporidium parvum, calves, neonatal diarrhea, environment, molecular characterization - Cryptosporidium parvum is a protozoan parasite responsible for intestinal infection of humans and animals causing high morbidity and death in immunocompromised hosts and it is considered a major cause of diarrhea in calves. The main transmission route of the oocysts is fecal-oral. This parasite represents a serious problem both in animal health and production and public health since some genotypes are zoonotic. This project aims to assess possible sources of C. parvum contamination for calves in a dairy farm, which could help develop more effective strategies to prevent and control the spread of this parasite, safeguarding the health and well-being of both animals and humans. Fecal samples from adult cows were collected in prepartum (n=6) and postpartum (n=7) periods as well as at birth (n=7). Fecal materials from the 7 calves were collected daily from birth until day 21. Environmental samples such as gloves, calving pen, Petri dishes collecting aerosols from high pressure water cleaning, swabs from various equipment such as boots, housing, buckets, teats, feed troughs and esophageal probe, were collected. The methodology was based on the identification of Cryptosporidum spp. oocysts by Ziehl-Neelsen stain and a direct immunofluorescence assay. DNA from environmental samples and calves' fecal samples from the 10th day were extracted and analyzed by PCR of the 18S rRNA gene to confirm the presence of Cryptosporidium spp. Some pre-entrance boxes (4/6), water buckets (2/5), teats (1/5), feeders (4/5), Petri dishes (2/7), bedding from the calving pen and all calves (7/7), exit boxes (7/7), gloves (2/2) and boots (2/2) tested positive. RFLP confirmed the presence of Cryptosporidium parvum. Sequence analysis of the gp60 gene confirmed the genotype as IIaA15G2R1. Given that the samples of the 7 cows tested negative at calving, they can be excluded as a potential source of infection to the calves. By day 6, 4 out of 7 calves were positive and by day 7 all tested positive, suggesting that the infection occurs between days 0 and 4, considering a pre-patent period of 3 to 6 days. Cryptosporidium spp. was detected in all types of environmental samples (except esophageal tube). This supports the hypothesis that the problem is in the handling and cleaning strategies. In these conditions, newborn calves are exposed to the agent since day 0, what may justify the high prevalence of the infection and disease on dairy farms
ABSTRACT - Cryptosporidium parvum, calves, neonatal diarrhea, environment, molecular characterization - Cryptosporidium parvum is a protozoan parasite responsible for intestinal infection of humans and animals causing high morbidity and death in immunocompromised hosts and it is considered a major cause of diarrhea in calves. The main transmission route of the oocysts is fecal-oral. This parasite represents a serious problem both in animal health and production and public health since some genotypes are zoonotic. This project aims to assess possible sources of C. parvum contamination for calves in a dairy farm, which could help develop more effective strategies to prevent and control the spread of this parasite, safeguarding the health and well-being of both animals and humans. Fecal samples from adult cows were collected in prepartum (n=6) and postpartum (n=7) periods as well as at birth (n=7). Fecal materials from the 7 calves were collected daily from birth until day 21. Environmental samples such as gloves, calving pen, Petri dishes collecting aerosols from high pressure water cleaning, swabs from various equipment such as boots, housing, buckets, teats, feed troughs and esophageal probe, were collected. The methodology was based on the identification of Cryptosporidum spp. oocysts by Ziehl-Neelsen stain and a direct immunofluorescence assay. DNA from environmental samples and calves' fecal samples from the 10th day were extracted and analyzed by PCR of the 18S rRNA gene to confirm the presence of Cryptosporidium spp. Some pre-entrance boxes (4/6), water buckets (2/5), teats (1/5), feeders (4/5), Petri dishes (2/7), bedding from the calving pen and all calves (7/7), exit boxes (7/7), gloves (2/2) and boots (2/2) tested positive. RFLP confirmed the presence of Cryptosporidium parvum. Sequence analysis of the gp60 gene confirmed the genotype as IIaA15G2R1. Given that the samples of the 7 cows tested negative at calving, they can be excluded as a potential source of infection to the calves. By day 6, 4 out of 7 calves were positive and by day 7 all tested positive, suggesting that the infection occurs between days 0 and 4, considering a pre-patent period of 3 to 6 days. Cryptosporidium spp. was detected in all types of environmental samples (except esophageal tube). This supports the hypothesis that the problem is in the handling and cleaning strategies. In these conditions, newborn calves are exposed to the agent since day 0, what may justify the high prevalence of the infection and disease on dairy farms
Description
Dissertação de Mestrado Integrado em Medicina Veterinária, área científica de Clínica
Keywords
Cryptosporidium parvum Vitelos Diarreia neonatal Ambiente Caracterização molecular Cryptosporidium parvum Calves Neonatal diarrhea Environment Molecular characterization
Pedagogical Context
Citation
Rodrigues MLFRF. 2024. Fontes ambientais de infeção por Cryptosporidium parvum em vitelos leiteiros recém nascidos [dissertação de mestrado]. Lisboa: FMV-Universidade de Lisboa
Publisher
Universidade de Lisboa, Faculdade de Medicina Veterinária