Rapid detection of the common avian leukosis virus subgroups by real-time loop-mediated isothermal amplification

Abstract

BackgroundSubgroups A, B, E and J are the major subgroups of avian leukosis virus (ALV) infecting chickens. ALV infection has become endemic in China and has a significant negative effect on the poultry industry. Consequently, there is an urgent need for a specific, sensitive and rapid method for diagnosis and eradication of ALV. Therefore, we developed a simple and rapid real-time loop-mediated isothermal amplification (LAMP) reaction for the timely detection of the common ALV subgroups, whereby the amplification can be obtained in 35 min under isothermal conditions at 63 °C, ability to specific, sensitive and rapid detect all the common ALV subgroups.MethodsA set of four specific primers was designed to target the sequences of the pol gene of ALV, and the loop-mediated isothermal amplification (LAMP) assay were developed and compared with PCR and virus isolation methods.ResultsThe results from specificity of the LAMP assay showed that only target ALVs DNA was amplified. The LAMP assay demonstrated a sensitivity of 20 copies/reaction of ALV DNA, which was 10 times higher than the conventional PCR measurement. To further evaluate the reliability of the method, the assay was evaluated with ALV DNA from a panel of 81 clinical samples suspected of ALV infection. The results verify that the LAMP method was more sensitive than the conventional PCR and virus isolation method.ConclusionIn conclusion, the developed LAMP assay was a simple, inexpensive, sensitive method for the rapid detection of the most common subgroups of ALV, and it provided a useful and practical tool in the eradication program for ALV in the poultry industry.

Characterization of Marek’s disease virus and phylogenetic analyses of meq gene from an outbreak in poultry in Meghalaya of Northeast India

“…Though the MD vaccines are capable of preventing lymphoma formation and clinical disease, do not prevent superinfection by pathogenic MDV strains [15]. Therefore, detection and elimination of infected birds form the basis for diagnosis and eradication programmes for oncogenic viruses [16]. In the present study, we investigated the samples from an outbreak, the lesions suspected of oncogenic viruses in poultry by histopathology of organs involved and molecular characterization of virus to understand the strain of virus present in the poultry flock in the region.…”

Biotic concerns in generating molecular diagnosis matrixes for 4 avian viruses with emphasis on Marek’s disease virus

“…Many LAMP assays have been developed and validated for important epizootic diseases, including 18 viruses thought notifiable of ruminants, swine and poultry by the World Organization for Animal Health (OIE) (Mansour et al, 2015). The LAMP assay was also developed for detection of the Tembusu virus (Tang et al, 2016), four immunosuppressive viruses (Song et al, 2018), avian leucosis virus (Peng et al, 2015), avian reovirus (Kumar et al, 2017), and found to be useful also in the detection of MDV in feathers and internal organs of infected chickens (Woźniakowski and Samorek-Salamonowicz, 2014). Direct detection of MDV DNA in poultry dust has been conducted without DNA extraction.…”

GLAPD: Whole Genome Based LAMP Primer Design for a Set of Target Genomes

“…A group-specific primer for white spot syndrome virus should only amplify the 16 strains but no other viruses. Traditionally, the group-specific primers are designed based on conserved genes (Peng et al, 2015), genome regions (Yao et al, 2016) or the multiple sequence alignment (MSA) of these genes or genomic regions (Kurosaki et al, 2010). But this method is limited by the small number of suitable genes and the difficulty to generate MSA for a large number of sequences (Chen and Tompa, 2010).…”