Enzyme-linked immunosorbent assay for immunoglobulin M specific antibody for the diagnosis of melioidosis

Abstract

Indirect hemagglutination (IHA) is commonly used for serodiagnosis of melioidosis. However, in endemic areas, high background titers in normal populations and occasional low titers in patients with septicemic melioidosis prompted a search for a more sensitive and more specific method of serodiagnosis. An indirect fluorescent-antibody test for immunoglobulin M (IgM) specific antibody to Pseudomonas pseudomallei was more sensitive and more specific, but fluorescence microscopes are rarely available in the endemic areas. An enzyme-linked immunosorbent assay (ELISA) for IgM antibody is an attractive alternative. An indirect ELISA for IgM antibody (IgM ELISA) and an IgM antibody capture ELISA for melioidosis were developed. Both tests, together with IHA, were evaluated for 153 serum specimens from blood donors and 16 serum specimens from 16 melioidosis patients. It was found that IHA, the IgM ELISA, and the IgM antibody capture ELISA had sensitivities of 88, 88, and 75%, respectively, with specificities of 97.4, 92.2, and 91.5%, respectively. When IHA was combined with IgM ELISA, a sensitivity of 100% and a specificity of 95.4% were obtained. The IgM ELISA and IHA should be used in combination for serodiagnosis of melioidosis.

Gold blot for detection of immunoglobulin M (IgM)- and IgG-specific antibodies for rapid serodiagnosis of melioidosis

“…As shown in our control from an endemic area (Table 2), subclinical infection forced the increase of the cutoff value with a subsequent decrease in sensitivity. To compensate for the loss of sensitivity and to cover both chronic and acute cases, parallel combination of the tests have been proposed (6). IHA which measured total antibody showed better agreement with the protein A gold blot which reacted preferably with IgG (3).…”

Enzyme-linked immunosorbent assay (ELISA) using a glycolipid antigen for the serodiagnosis of melioidosis

“…The IHA test is widely used for the serodiagnosis of melioidosis because of its simplicity and low cost. The usefulness of IHA test was compared with that of IgM-ELISA [2,6,15], IgM gold blot [16] and IFA [4] tests. These studies concluded that IHA should not be used alone in the diagnosis of melioidosis, especially in the endelmic areas of melioidosis.…”

“…A partially purified protein antigen (19.5 kDa) was reported to be highly sensitive and specific for the diagnosis of melioidosis [3], however, due to its protein nature, the stability has not been guaranteed and satisfactory techniques for the preparation of this antigen make it difficult for use in most endemic areas. In comparison with IHA, the ELISA generally gives higher sensitivity and specificity, however, in some melioidosis patients, ELISA shows negative results in IHA-positive sera [2]. The combined results of IHA and ELISA tests, therefore, may be more reliable in the diagnosis of melioidosis than either test alone.…”

“…cannot be based solely on clinical symptoms but requires bacteriological examinations including culture confirmation. Unfortunately, the latter is usually time-consuming (minimum 2 days) [2] and the result is often too late to be useful for appropriate early treatment [3]. Moreover, the prognosis of acute septicaemic melioidosis depends on early diagnosis and appropriate treatment [I].…”

“…The serodiagnosis of melioidosis often gives some undesirable results, for example the indirect haemagglutination (IHA) test is hampered by a high background antibody titre in endemic areas resulting in low specificity [2,4] and may be negative in some patients with septicaemic melioidosis [2,5]. An enzyme-linked immunosorbent assay (ELISA) with a protein as the antigen had excellent sensitivity but false positive results occurred with P. aeruginosa and B. cepaciu infections [6].…”