On-Site Test for Equine Glanders

Editorial note: This paper is a follow-up to a report on glanders that appeared in the October 2003 Lloyds Equine Disease Quarterly.

Diagnostic testing is evaluated according to how precise the test is in identifying a truly positive animal versus animals that are truly negative.

Sensitivity is the proportion of true positives (bacterial culture positive) that are identified by the test as being positive and is a measure of how accurately the test can identify animals with the disease.

Specificity is the proportion of true negatives (in this study, Pakistan Army horses) identified by the test as being negative. This is a measure of how accurately the test can identify animals without the disease.

Positive predictive values reflect the proportion of animals with a positive test result that are truly positive for the disease; negative predictive values reflect the proportion of animals with a negative test result that are truly negative for the disease.

A test with high sensitivity and specificity would be ideal; however, no test exists that is 100% sensitive and 100% specific. Higher sensitivity tests are important to identifying diseased animals. Higher specificity tests are useful for identifying disease-free animals.

Glanders is an important infectious equine disease of imposing antiquity. It is endemic in Brazil, Turkey, United Arab Emirates, Iran, Iraq, India, Pakistan, and some other countries. Equine glanders has been eradicated from North America and Europe, but its potential as a weapon of bio-sabotage still haunts professionals as well as many others.

Researchers in developing countries are still working on diagnosis, treatment, and vaccine development. Since a confirmatory laboratory diagnosis isn’t available in many areas, diagnosis often has been made based on clinical signs and the mallein test. The mallein test is a cornerstone for the field diagnosis of glanders, but its limitation in terms of sensitivity, specificity, and accuracy are well recognized. These pitfalls of the mallein test prompted Russian scientists to develop a new on-site screening test for glanders, the Rose Bengal plate agglutination test (RBT). This test has not as yet been fully validated in terms of sensitivity, specificity, and accuracy under field conditions.

To this end, 70 positive (culture positive; gold standard) sera were collected from two separate outbreaks of glanders that occurred among horses of the Faisalabad Metropolis Mounted Police and Lahore Polo Club in Pakistan. Ninety-six sera from clinically normal but potentially exposed equines (in contact animals/ cohort at the Lahore Polo Club) were also included. One-hundred and ten sera collected from the Pakistan Army establishment (Mona Depot) served as negative controls.

Antigen for RBT was prepared from three different strains of Burkholderia mallei as per the Office International des Epizooties (OIE) (2003) method. Indirect Haemagglutination (IHA), Complement Fixation (CF), and modified Counter immunoelectrophoresis (mCIE) tests were run in tandem. The results of these tests are summarized in Figure 1.

Figure 1. Results for Five Glanders Test Methods

Both mallein and mCIE tests were significantly (P<0.05) less sensitive (81.4 and 75.7%) than IHA, CF, and RBT tests. The low sensitivity of the mallein test in this study concurs with that previously documented by this institution (Muhammad et al., 1998; J. Equine Sci. (Japan), 9(3):93–96). All tests but mCIE gave 100% specificity when testing sera from animals belonging to the glanders-free group (Pakistan Army horses).

In the present study the mallein test was least sensitive (75.5%) for clinical and bacteriologically positive equines. The RBT test in comparison with the mallein test was more sensitive and gave higher proportion of agreement to CF and IHA tests.

In the light of technically acceptable sensitivity, specificity, and negative and positive predictive values, the Rose Bengal test seems to be a good on-site field diagnostic tool for equine glanders. In comparison with other serological tests (IHA, CF, and mCIE) evaluated in this study, RBT is economical, rapid, and compatible with the technical capabilities of practicing veterinarians.

Authors: Dr. Abeera Naureen, Muhammad Saqib, Muhammad Nadeem Asi, Muhammad Hammad Hussain, Ghulam Muhammad
Department of Clinical Medicine and Surgery, Faculty of Veterinary Science - University of Agriculture, Faisalabad, Pakistan