Application of loop-mediated isothermal amplification (LAMP) assays for the detection of bovine herpesvirus 1

Abstract

Bovine herpesvirus-1 (BoHV-1), a causative agent of Infectious Bovine Rhinotracheitis (IBR), is responsible for high economic losses in cattle farming industry. The use of testing methods that allow early detection of BoHV-1-infected animals is a key element of each program of IBR eradication. The aim of the study was to design and evaluate two variants of LAMP isothermal tests with SYBR Green fluorescence probes, specific to the genes encoding gD and gE glycoproteins of BoHV-1. LAMP gE BoHV-1 assay was able to distinguish between gE-and gE+ strains of the virus. Both LAMP gD and gE assays were specific to BoHV-1 and did not react with other related to BoHV-1 alphaherpesviruses. Sensitivity of LAMP gD was 2x10 4 copies of the viral genome whereas for LAMP gE it was 2x10 5 . Diagnostic sensitivity calculated for LAMP gD was 64.7% whereas for LAMP gE it was 80%. Diagnostic specificity for LAMP gD and LAMP gE was 78.9% and 89.3%, respectively. LAMP assay can be a rapid and simple method of diagnosis of acute BoHV-1 infections and discrimination of gE -strains.However, relatively low diagnostic sensitivity of the method can limit its use in routine diagnostics.

Establishment and application of isothermal multiple-self-matching-initiated amplification (IMSA) in detecting Type II heat-labile enterotoxin of Escherichia coli

“…Since 2000, LAMP has been broadly employed to detect a variety of pathogenic microorganisms. Subsequently, CPA, another isothermal amplification method, was firstly used to detect Mycobacterium tuberculosis in sputum specimens [9, 21]. Similar to LAMP, CPA is based on amplification using many cross-linked primers.…”

Diagnosis of Ruminant Viral Diseases with Loop-Mediated Isothermal Amplification

“…In clinical nasal swab samples, differentiating LAMP assays have lower diagnostic sensitivity and specificity. Neither assay developed by Socha and colleagues showed any cross-reactivity with other alpha herpesviruses, and LAMP glycoprotein-E successfully differentiated gE-deleted vaccine strains from the wild-type strains [ 21 ]. LAMP assays’ target region in the viral genome and assay conditions are summarized in Table 1 .…”

Diagnostic techniques for infectious bovine rhinotracheitis: a review

“…Bovine herpesvirus-1 LAMP assays were composed of 6 µL of Isothermal Master Mix (Optigen ISO-001) and 2.5 µL of DNA sample (El-Kholy et al, 2014;Xu et al, 2016). Reactions were run in the thermoblock for 60 minutes at 66 o C. Results were visualized by the addition of 1 µL of 1000x concentrated SYBR Green per sample (Socha et al, 2017). Positive samples turned green, while negative samples remained orange, as indicated in Figure 3 below.…”