Date of Award

2009

Document Type

Master Thesis

Degree Name

Masters of Science (Research)

Department

Biological Sciences

First Advisor

Dr. Helen O'Shea

Second Advisor

Dr. Brigid Lucey

Abstract

The issue of food safety is a growing concern to Irish agriculture and to the food industry. In the last twenty years, many novel pathogens, which can cause concern to the food industry, have been identified. Organisms such as Verocytotoxigenic Escherichia coli (VTEC) cause hemorrhagic colitis and Haemolytic Uremic Syndrome (HUS), which is the major cause of acute renal failure in children in many countries, and it’s presence in food is a cause of growing concern. Other pathogens causing concern are those implicated as causative agents of Acute Gastroenteritis (AGE) and these can be bacterial, viral and parasitic organisms.

Consumption of contaminated water has been implicated as a major source of Cryptosporidium infection in various outbreak investigations and case control studies. Recent outbreaks of Cryptosporidiosis, caused by Cryptosporidium parvum in the United States and other countries, as well as the emergence of Cryptosporidiosis as a frequent cause of morbidity and mortality in immunodeflcient individuals, have raised the interest of the research community in this parasite. Cryptosporidium infection is a zoonotic infection and this disease is on the rise and is extremely common in both calves and humans.

Rotavirus and Norovirus are also causative agents of AGE. These pathogens are implicated in the cause of severe disease in both humans and animals and there is growing awareness regarding their spread into the human food chain via water, unpasteurised milk, milk and meat products.

Samples for this project were collected from ten farms in the Cork harbour area on 2 occasions, over a two year study. Faecal samples were taken from twenty calves on each farm and two litre water samples were collected from selected water points on the farm following analysis of a questionnaire supplied to the farm manager. The samples analysed consisted of 400 faecal samples and 50 water samples.

The diagnostic techniques employed to detect and characterize these pathogens included Auramine staining and the modified Ziehl-Neelson staining method, to detect and confirm the presence of Cryptosporidium parvum. Following diagnosis, DNA was extracted from these samples and they were subjected to PCR. Restriction Fragment Length Polymorphism (RFLP) analysis was employed to determine the species of Cryptosporidium present in the faecal samples. From there, a correlation was made between farm animals and contamination of water supplies. Detection of Rotavirus and Norovirus was achieved through the use of an Enzyme Linked Immunosorbent Assay (ELISA) and RNA was extracted using a phenol chloroform method. These viruses were characterized using a Reverse Transcriptase Polymerase Chain Reaction (RT-PCR). The water samples were also tested for the presence of C. parvum, rotavirus and norovirus. The determination of total plate counts at 22®C and 37“C of total coliforms and faecal conforms was also performed on all of the water samples.

From the findings, C. parvum was found at a prevalence rate of greater than 50% in the faeces of the calves analysed over the two year study. These results were consistent for both years of the study. The study showed that there was a higher prevalence of C. parvum infection in the female population, with 69% of positive samples found in female bovines. The prevalence of rotavirus in the bovine population was very low, with 5% of samples determined to be positive. Again, the female bovine population had a higher incidence rate at 3%. The water samples were tested for norovirus, however, no isolates were detected. Results from the MPN and total coliform counts showed that house and milk parlour water supplies on farms were clean but that cow troughs and cow shed supplies had a very high bacterial count. The results of this study are important from a public health point of view, as regards investigating the quality of the water supplies to milk production holdings and determining if a range of pathogens are present in the water and in determining the parasite prevalence in cohort herds. It may also assist in the design of prevention and control strategies and further our understanding of the epidemiology of these important zoonotic pathogens.

Access Level

info:eu-repo/semantics/openAccess

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