Development of Rose Bengal test against mallein test for rapid diagnosis of equine glanders

Karimi, Amin; Mosavari, Nader

doi:10.1007/s11250-019-01890-6

Cited by 7 publications

(7 citation statements)

References 10 publications

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“…Although this test is believed to have a high specificity, a considerable number of false-positive results were observed when different antigens and assay protocols were applied (10,11); complexity, anti-complementary reactions and poor standardization are other shortcomings of the CFT (12). The Rose Bengal plate agglutination test is a rapid agglutination assay using a heat-inactivated bacterial suspension colored with Rose Bengal; it is fast and simple but requires antigens purified from B. mallei cultures (9,13).…”

Section: Complement Fixation Test (Cft) Is the Most Accurate And Relimentioning

confidence: 99%

A glycoengineered antigen exploiting a conserved proteinO-glycosylation pathway in theBurkholderiagenus for diagnosis of glanders infections

Wang

Glaser

Scott

et al. 2020

Preprint

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We recently described a protein O-glycosylation pathway conserved in all species of the Burkholderia genus that results in synthesis and incorporation of a trisaccharide glycan to membrane-exported proteins. Here, we exploited this system to construct and evaluate a diagnostic tool for glanders. Burkholderia mallei, the causative agent of glanders, is a highly infectious and fatal zoonotic pathogen that mainly infects horses, mules, donkeys and occasionally humans. A highly sensitive and specific diagnostic tool is crucial for the control, elimination and eradication of B. mallei infections. We constructed plasmids carrying synthetic genes encoding a modified, previously unannotated Burkholderia glycoprotein containing three glycosylation sequons fused to the cholera toxin B-subunit. The resulting proteins were glycosylated in the B. cenocepacia K56-2 parental strain, but not in glycosylation-deficient mutants, as determined by SDS-PAGE and fluorescent lectin blots. One of these glycoproteins was used as an antigen in ELISA and western blots to screen a panel of serum samples collected from glanders-infected and healthy horses previously investigated by complement fixation test and indirect ELISA based on a semi-purified fraction of B. mallei. We show that ELISA and western blot assays based on our glycoprotein antigen provide 100 % specificity, with a sensitivity greater than 88%. The glycoprotein antigen was recognized by serum samples collected from patients infected with B. pseudomallei, B. mallei, B. multivorans and B. cenocepacia. Our results indicate that protein O-glycosylation in Burkholderia can be exploited as a biomarker for diagnosis of Burkholderia-associated infections.IMPORTANCEGlanders is a severe zoonotic disease caused by the Gram-negative bacterium Burkholderia mallei, which affects horses, mules and donkeys, as well as humans. B. mallei is also considered a category B biothreat agent. Due to insufficient pathognomonic symptoms in the early stages of glanders, diagnosis can be difficult. Complement fixation is the most accurate and reliable serological test prescribed by the World Organization for Animal Health; however, this test has a considerable number of false-positive results. We have recently described a conserved protein O-glycosylation pathway present in all species of the Burkholderia genus; we also demonstrated that Burkholderia-infected humans develop anti-glycan antibodies. Here, we exploited this system to construct and evaluate a synthetic glycoengineered protein antigen as a diagnostic tool for glanders. Our results show 100 % specificity in the detection of antibodies from infected horses, indicating that protein O-glycosylation in Burkholderia can be exploited as a biomarker for diagnosis of Burkholderia-associated infections.

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Section: Complement Fixation Test (Cft) Is the Most Accurate And Relimentioning

confidence: 99%

A glycoengineered antigen exploiting a conserved proteinO-glycosylation pathway in theBurkholderiagenus for diagnosis of glanders infections

Wang

Glaser

Scott

et al. 2020

Preprint

View full text Add to dashboard Cite

show abstract

“…Although this test is believed to have a high specificity, a considerable number of false-positive results were observed when different antigens and assay protocols were applied [10,11]; complexity, anti-complementary reactions and poor standardization are other shortcomings of the CFT [12]. The Rose Bengal plate agglutination test is a rapid agglutination assay using a heatinactivated bacterial suspension colored with Rose Bengal; it is fast and simple but requires antigens purified from B. mallei cultures [9,13].…”

Section: Introductionmentioning

confidence: 99%

A glycoengineered antigen exploiting a conserved protein O-glycosylation pathway in the Burkholderia genus for detection of glanders infections

et al. 2021

View full text Add to dashboard Cite

We recently described a protein O-glycosylation pathway conserved in all species of the Burkholderia genus that results in the synthesis and incorporation of a trisaccharide glycan to membrane-exported proteins. Here, we exploited this system to construct and evaluate a diagnostic tool for glanders. Burkholderia mallei, the causative agent of glanders, is a highly infectious and fatal zoonotic pathogen that infects horses, mules, donkeys, and occasionally humans. A highly sensitive and specific diagnostic tool is crucial for the control, elimination, and eradication of B. mallei infections. We constructed plasmids carrying synthetic genes encoding a modified, previously unannotated Burkholderia glycoprotein containing three glycosylation sequons fused to the cholera toxin B-subunit. The resulting proteins were glycosylated in the B. cenocepacia K56-2 parental strain, but not in glycosylationdeficient mutants, as determined by SDS-PAGE and fluorescent lectin blots. One of these glycoproteins was used as an antigen in ELISA and western blots to screen a panel of serum samples collected from glanders-infected and healthy horses, which were previously investigated by complement fixation test and indirect ELISA based on a semi-purified fraction of B. mallei. We show that ELISA and western blot assays based on our glycoprotein antigen provide 100% specificity, with a sensitivity greater than 88%. The glycoprotein antigen was recognized by serum samples collected from patients infected with B. pseudomallei, B. mallei, B. multivorans, and B. cenocepacia. Our results indicate that protein O-glycosylation in Burkholderia can be exploited as a biomarker for diagnosis of Burkholderiaassociated infections.

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“…The diagnosis of glanders in horses is problematic, especially during the early stages of infection, in which infected horses do not show outward symptoms [9]. The mallein skin test [2,3,8,24] (allergic hypersensitivity test) has been widely used in conventional field tests. However, due to animal welfare concerns, this test is not currently recommended [26].…”

Section: Introductionmentioning

confidence: 99%

“…CFT is mandated by the World Organization for Animal Health (OIE) as a confirmatory test for international trade [26]. Rose Bengal plate agglutination test [8,25] (RBT) is approved in Russia but, due to its relatively low sensitivity, it is not commonly used in other countries. Other serological tests, including immunoblot assay [28,33] and enzyme-linked immunosorbent assay (ELISA) [4,15,16,29], have also been reported as useful diagnostic methods for glanders; however, these assays have not been From to 1968, a project named "Veterinary expeditions of Central and Eastern European countries against brucellosis, tuberculosis, and glanders in Mongolia" successfully eliminated the disease in this country [13,14].…”

Section: Introductionmentioning

confidence: 99%

Seroprevalence of equine glanders in horses in the central and eastern parts of Mongolia

et al. 2020

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Glanders is a contagious and fatal equine disease caused by the gram-negative bacterium Burkholderia mallei. B. mallei is prevalent among horse populations in Asia, the Middle East, and South America. More than four million horses have been registered in Mongolia in 2020. However, the resent prevalence of glanders has not been well investigated. In this study, we aimed to investigate the seropositivity of B. mallei in horse populations in Mongolia using the complement fixation test (CFT) and Rose Bengal plate agglutination test (RBT). We randomly collected blood samples from horses in central and eastern Mongolia between 2018 and 2019. Of 337 horses, 26 (7.7%) and 28 (8.3%) were seropositive using RBT and CFT, respectively. Interestingly, seropositivity in horses resulting from crossbreeding of Mongolian native horses with thoroughbred horses was higher than that in Mongolian native horses. Our observations suggest that equine glanders are still endemic to Mongolia.

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Development of Rose Bengal test against mallein test for rapid diagnosis of equine glanders

Cited by 7 publications

References 10 publications

A glycoengineered antigen exploiting a conserved proteinO-glycosylation pathway in theBurkholderiagenus for diagnosis of glanders infections

A glycoengineered antigen exploiting a conserved proteinO-glycosylation pathway in theBurkholderiagenus for diagnosis of glanders infections

A glycoengineered antigen exploiting a conserved protein O-glycosylation pathway in the Burkholderia genus for detection of glanders infections

Seroprevalence of equine glanders in horses in the central and eastern parts of Mongolia

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