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Development of a lateral flow test for the rapid detection of Avibacterium paragallinarum in chickens suspected of having infectious coryza

BMC Vet Res.. 2018-12;

Sandra Morales Ruiz, Jorge Bendezu, Ricardo Choque Guevara, Ricardo Montesinos, David Requena, Luz Choque Moreau, Ángela Montalván Ávalos and Manolo Fernández-Díaz

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Polyclonal Antibody Services	...Then, peptides were predicted on the unflagged regions using Bebipred 2.0 and Optimum Antigen (GenScript Laboratories, Piscataway, NJ, USA)...For monoclonal antibody production, the peptide (Ma-4) was synthesized and used to immunize a BalbC mouse which was provided by GenScript Laboratories... The peptide synthesis, hybridoma production, clone selection, isotyping and antibody titers were performed by GenScript Laboratories...The immune reaction was performed with peroxidase-labelled goat IgG anti-mouse antibody (GenScript Laboratories) using radiance as a substrate...	Get A Quote
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摘要

Background: Infectious coryza (IC) is an acute respiratory disease of growing chickens and layers caused by Avibacterium paragallinarum. The development of tools that allow rapid pathogen detection is necessary in order to avoid disease dissemination and economic losses in poultry. An Av. paragallinarum-specific Ma-4 epitope of the TonB-dependent transporter (TBDT) was selected using bioinformatic tools in order to immunize a BalbC mouse and to produce monoclonal antibodies to be used in a lateral flow test (LFT) developed for Av. paragallinarum detection in chicken nasal mucus samples. Results: The 1G7G8 monoclonal antibody was able to detect TBDT in Av. paragallinarum cultures (serogroups: A, B and C) by Western blot and indirect ELISA assay. Consequently, we developed a self-pairing prototype LFT. The limit of detection of the prototype LFT using Av. paragallinarum cultures was 1 × 104 colony-forming units (CFU)/mL. Thirty-five nasal mucus samples from chickens suspected of having infectious coryza were evaluated for the LFT detection capacity and compared with bacterial isolation (B.I) and polymerase chain reaction (PCR). Comparative indicators such as sensitivity (Se), specificity (Sp), positive predictive value (PPV), negative predictive values (NPV) and the kappa index (K) were obtained. The values were 100.0% Se, 50% Sp, 65.4% PPV, 100% NPV, and 0.49 K and 83.9% Se, 100% Sp, 100% PPV, 44.4% NPV, and 0.54 K for the comparison of the LFT with B.I and PCR, respectively. Additionally, the LFT allowed the detection of Av. paragallinarum from coinfection cases of Av. paragallinarum with Gallibacterium anatis. Conclusions: The results indicate that the self-pairing prototype LFT is suitable for the detection of TBDT in Av. paragallinarum cultures as well as in field samples such as nasal mucus from Av. paragallinarum-infected chickens. Therefore, this prototype LFT could be considered a rapid and promising tool to be used in farm conditions for Av. paragallinarum diagnosis.