Analysis of the Association between ELISA and Nested PCR on Blood and Milk for Mycobacterium avium subsp. paratuberculosis Detection in Holstein Cows

Abstract

Paratuberculosis is a chronic, infectious disease of ruminants characterized by progressive weight loss and profuse diarrhea. It is caused by the acid-fast bacillus Mycobacterium avium subspecies paratuberculosis (MAP), has a worldwide distribution and is categorized by the OIE as a List B disease that has serious economic impact or is a public health concern. Disease diagnosis is hampered by a lack of sensitive tests. Available methods fail to identify all infected animals, and many produce substantial numbers of false positives and false negatives. This is of particular importance, as they relate to detection of the organism in subclinically infected animals. Several commercial, enzyme-linked immunosorbent assay (ELISA) tests are available, but it is generally accepted that their sensitivity in detecting infected animals is only about 50%. In fact, because of its low sensitivity, the ELISA test is rarely positive in animals under two years of age. Polymerase chain reaction (PCR) tests based on the insertion element IS900 have been the most widely used for MAP identification. However, detection of the etiologic agent is limited by frequency and number of organisms present in the fluid or tissue. A combination of serologic tests, such as ELISA, and agent detection through nested PCR could be a useful strategy to improve the sensitivity of paratuberculosis diagnosis, especially when detectable target levels for each test follow different temporal patterns.

The objective of this study was to compare the performance of ELISA testing of sera, and nested PCR in milk and blood, for diagnosis of paratuberculosis in Holstein cows.