Development and application of dot-enzyme-linked immunosorbent (dot-ELISA) assay for detection ofBrucella melitensisand evaluation of the shedding pattern in infected goats

Abstract

Early and accurate diagnosis of Brucella melitensis is essential for the treatment and control of brucellosis both in animals and humans. The thrust for the development of a rapid diagnostic technique to overcome the limitations of conventional microbiological and serological tests brought about this investigation on the development and application of dot-ELISA for antigen and antibody detection in infected goats. Fifteen apparently healthy Boer aged 2-3 years which tested negative for brucellosis using PCR and ELISA, were grouped into A (10 goats infected intraocularly with 10 CFU of B. melitensis) and B (5 goats) as control. Discharges (ocular, nasal, and vaginal) and blood were collected at days 3, 7, 10, 14, weekly until 42 post-infection (pi) for dot-ELISA, PCR, and RBPT. Dot-ELISA detected B. melitensis antigen and antibody in group A at day 3 and 7 pi, respectively with adequate sensitivity and specificity relative to PCR and RBPT. The bacteria shedding detected from discharges at day 3 pi in the nasal and ocular route with dot-ELISA. Group B were consistently negative. Values such as speed, simplicity, field adaptability, high sensitivity, and specificity make dot-ELISA a rapid and adequate technique for diagnosis of brucellosis in B. melitensis infected goats within few hours.

Immunohistochemical detection of <i>Brucella melitensis</i> antigens in lungs tissues of sheep and goats diagnosed with different diseases at necropsy

“…The primary antibody that was used was prepared following the guide lines stated in OIE (OIE Terrestrial Manual, 2009) as described in our earlier study (Onilude et al, 2017).…”

“…This was to promote uniform distribution of antibody on tissue sections. Primary antibody was derived from serum of B. melitensisinfected goat which was tested seropositive by RBPT (Onilude et al, 2017). Sections were incubated with primary antibody at the dilution of 1:50 in BSA-PBS for at least one hour at 37C in an incubator.…”

“…Brucellosis is regarded by World Health Organization (OIE Terrestrial Manual, 2009) as one of the most important zoonosis. Brucellosis in small ruminants, especially goats is caused by Brucella melitensis and the rate of transmission to humans have substantially increased in countries that lack effective control based programmes (Megid et al, 2010;Onilude et al, 2017;Higgins et al, 2017). Infected animals are often the main source of human infection (Tanko et al, 2013;Higgins et al, 2017).…”

“…Currently, diagnosis of Brucellosis is based on serological and microbiological methods which are not without limitations. The limitations for serological tests include cross-reactivity with other antigenically related bacteria and inability of the serological techniques to detect early or latent infection (Nwunuji et al, 2013;Onilude et al, 2017). Bacteriological methods on the other hand are time consuming, cumbersome, relatively expensive and with limited sensitivity (Ahmed et al, 2010).…”