Infectious bursal disease virus tissue tropism and pathogenesis of the infection in chickens by application of in situ PCR, immunoperoxase and HE staining

Hussein, Elawad A.; Hair-Bejo, M.; Liew, Pit Sze; Adamu, Lawan; Omar, Abdul Rahman; Arshad, Siti Suri; Mohammed, M.H.; Ideris, Aini

doi:10.1016/j.micpath.2019.01.049

Cited by 8 publications

(2 citation statements)

References 13 publications

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“…PCR has been widely used in the epidemiological investigation of various poultry pathogens and antibiotic resistance genes (Okamatsu et al 2016 ; Stoute et al 2019 ; Hussein et al 2019 ; de Wit et al 2018 ; Kaltenboeck et al 2005 ; Luan et al 2016 ; Zhang et al 2017 ; Räty et al 2005 ). To ensure the accuracy of PCR results, it is essential that appropriate and reliable EICs are available which can be used to monitor the entire process; sample collection, transport and storage, DNA extraction, and PCR.…”

Section: Discussionmentioning

confidence: 99%

“…Due to its exquisite sensitivity and specificity, polymerase chain reaction (PCR) has become one of the most valuable, powerful and widely-used tools for rapid detection of infectious agents in poultry (Okamatsu et al 2016 ; Stoute et al 2019 ; Hussein et al 2019 ; de Wit et al 2018 ; Kaltenboeck et al 2005 ; Luan et al 2016 ). However, the accuracy of PCR results is highly dependent on the quantity and quality of the DNA in the test sample.…”

Section: Introductionmentioning

confidence: 99%

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Hydroxymethylbilane synthase (HMBS) gene-based endogenous internal control for avian species

et al. 2020

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With PCR becoming one of the most important and widely-used diagnostic tools for infectious diseases of poultry, an urgent need has developed for an endogenous internal control (EIC) that monitors the quality and quantity of poultry DNA in test samples. In this study we developed a SYBR-qPCR to target the poultry homolog of the hydroxymethylbilane synthase (HMBS) gene as an EIC for avian species. The avian HMBS-based qPCR was very sensitive, detecting one HMBS gene copy in a 20 µL reaction, and is highly specific for avian species. It amplified DNA from 11 organs and tissues of chickens showing it can be used as an EIC on a large variety of samples. The application of the established EIC on clinically and experimentally infected samples demonstrated that false negativity and result variations could result from samples being collected using different operators, techniques, preservatives, and storage times. The high sensitivity and specificity of the avian HMBS-based qPCR, its ability to quantify DNAs extracted from a wide range of tissues and poultry species along with its usefulness in reducing false negativity in PCR results associated with inadequate sampling and storage degradation makes it an ideal EIC for poultry DNA and RNA PCR diagnostics. The study also highlights the importance of appropriate sampling and storage of samples in ensuring accuracy of molecular diagnostic testing.

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