Salmonella Brandenburg in New Zealand sheep : the development of a serological diagnostic test and a case control study : a thesis presented in partial fulfilment of the requirements for the degree of Master of Veterinary Studies at Massey University

Abstract

Salmonella Brandenburg causes acute diarrhoea and severe illness in a variety of animals and was first isolated in New Zealand in 1986. Since 1996 Salmonella Brandenburg has been associated with an emerging epidemic of abortions and deaths in sheep in the southern regions of the South Island. Little is known about the specific epidemiology of Salmonella Brandenburg in sheep and as a result control to date has been largely based on anecdotal evidence and general principles. This study focused on the following aims: • To develop a serological test for use in epidemiological studies and for monitoring future control efforts targeting Salmonella Brandenburg in New Zealand sheep. • To identify factors associated with the occurrence and severity of Salmonella Brandenburg outbreaks in New Zealand sheep. Traditionally Salmonella diagnosis has depended on bacteriological culture. Such tests are time consuming, labour and equipment intensive, and may lack sensitivity. ELISA (Enzyme-Linked Immunosorbent Assay) methodologies offer an alternative for the diagnosis of Salmonella infection. Therefore the development of an ELISA test for detecting antibodies to Salmonella Brandenburg organism in sheep plasma was undertaken. Expression of common antigens has resulted in a high level of antibody cross-reactivity between different serovars in serological tests. Lipopolysaccharides (LPS) (O Antigens) are the primary cause of these cross-reactions. Cross-reactivity with two common sheep serovars (Salmonella Typhimurium and Salmonella Brandenburg) was of major concern for the development of a Salmonella Brandenburg ELISA. This was overcome by preparing an antigen mainly composed of flagella and fimbria proteins (LPS free). The antigen preparation was of a relatively crude and non-characterised nature and could only produce a reasonable optical density response at a high concentration. Unfortunately, while the ELISA was responsive, the specificity of the ELISA for Salmonella Brandenburg antibodies remained poor. Further investigation of the specificity of the antigen preparation, through the use of different sera, or through the development of a more pure and specific antigen, is needed for the successful development of a sensitive and specific serological test for determining Salmonella Brandenburg exposure in New Zealand sheep. A case control study was performed as part of a large-scale ongoing investigation aimed at identifying factors associated with Salmonella Brandenburg disease in New Zealand sheep. Details of disease prevalence and farm management methods were collected from two affected regions in southern New Zealand. Associations between possible risk factors and Salmonella Brandenburg were evaluated using odds ratios, with analyses being performed at two different levels: • farm level analysis to compare affected vs. unaffected farms using a case-control approach. • within farm analysis restricted only to affected farms to evaluate risk factors associated with severity of reported disease on affected farms. Data were collected from 405 farms containing a total of 1, 170,737 ewes. Of the 175 case farms, 97% had diseased mixed age (MA) ewes, 45% had diseased two-tooth (TT) ewes, and 5% had diseased hogget (H) ewes. Salmonella Brandenburg appeared to occur in better performing flocks, which are often associated with intensive farming methods. At the farm level, factors such as increased total number of ewes, feeding of hay, and controlled winter grazing appeared to increase the risk of disease. Farming methods such as controlled winter grazing may result in higher stress levels and increase the shedding of Salmonella Brandenburg organisms. This may create a higher risk of exposure in sheep yards and on pasture, resulting in a higher risk of disease. Feeding crop and having hilly terrain decreased the risk of a farm having disease. A protective effect of hilly terrain could be due to less intensive farm management, with a subsequent reduction in stress associated disease risk. Within affected farms, disease appeared to be more severe with the removal of rams after July, feeding of hay, and the practices of strop grazing. Shearing after July, increasing the total number of pre-lamb yardings, and vaccinating for Salmonella appeared to be protective. Therefore reducing stress and vaccinating ewes appear to reduce the risk of a Salmonella Brandenburg outbreak.