Evaluation of a multi-species Protein A-ELISA assay for plague serologic diagnosis in humans and other mammal hosts

Abstract

Background The Hemagglutination assay (HA) is widely used in plague diagnosis, however, it has a subjective interpretation and demands high amounts of antigen and other immunobiological supplies. On the other hand, the conventional Anti-IgG ELISA is limited by the need of specific conjugates for multiple plague hosts, which leaves a gap for new diagnostic methods able to cover both the diagnosis of human cases and the epidemiological surveillance of multiple sentinel species. Methods We developed an ELISA Protein A-peroxidase method to detect anti-F1 antibodies across several species, including humans. To determine the cut-off and performance rates, HA results from 288 samples (81 rabbits, 64 humans, 66 rodents and 77 dogs) were used as reference. Next, we evaluated the agreement between Protein A-ELISA and Anti-IgG ELISA in an expanded sample set (n = 487). Results Optimal conditions were found with 250ng/well of F1 and 1:500 serum dilution. Protein A-ELISA showed high repeatability and reproducibility. We observed good correlation rates between the Protein A and IgG ELISAs optical densities and a higher positive/negative OD ratio for the Protein A-ELISA method. The overall sensitivity, specificity and area under the curve for Protein A-ELISA were 94%, 99% and 0.99, respectively. Similar results were observed for each species separately. In the analysis of the expanded sample set, there was a strong agreement between Protein A and IgG assays (kappa = 0.97). Furthermore, there was no cross-reaction with other common infectious diseases, such as dengue, Zika, Chagas disease, tuberculosis (humans) and ehrlichiosis, anaplasmosis and leishmaniasis (dogs). Conclusions Altogether, the Protein A-ELISA showed high performance when compared both to HA and Anti-IgG ELISA, with a polyvalent single protocol that requires reduced amounts of antigen and can be employed to any plague hosts.

Refinement of the rKLi8.3-Based Serodiagnostic ELISA Allows Detection of Canine Leishmaniosis in Dogs with Low Antibody Titers

“…As an alternative to enzyme-conjugated antibodies, the S. aureus protein A (PA) and streptococcal protein G (PG) have been applied in the diagnosis of many infectious diseases with the advantage of having an affinity for immunoglobulins from various animal species. The evidence for a higher binding affinity of both PA and PG to IgG, promoting the detection of lower amounts of antibodies than anti-IgG conjugates, has been demonstrated in the diagnosis of diverse infectious diseases [ 29 , 30 ].…”

Lessons from a multilaboratorial task force for diagnosis of a fatal toxoplasmosis outbreak in captive primates in Brazil

“…For confirmatory purposes, two distinct multi-specie approaches were used: hemagglutination and ELISA-Protein A. The serology tests were performed by the Plague National Reference Service at the Instituto Aggeu Magalhães (IAM) -Fiocruz and the detailed protocol is described elsewhere [42].…”

Lessons from a Multilaboratorial Task Force for Diagnosis of a Fatal Toxoplasmosis Outbreak in Captive Primates in Brazil

“…For confirmatory purposes, two distinct multi-specie approaches were used: hemagglutination and ELISA-Protein A. The serology tests were performed by the Plague National Reference Service at the Instituto Aggeu Magalhães (IAM)-Fiocruz, and the detailed protocol is described elsewhere [40].…”