Optimization of the polymerase chain reaction (PCR) method for the detection of Oryctes rhinoceros virus

Ramle Moslim, Idris Abd. Ghani, Mohd Basri Wahid, Travis R. Glare, Trevor A. Jackson

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

Optimization of the polymerase chain reaction (PCR) met hod for the rapid detection of Oryctes rhinoceros virus (OrV) was studied. The virus DNA was ext racted from the gut tissues by a robust method. Using a pair of specific primers, Primer 15a and 15b, infection was confirmed when the PCR product produced a single 945 bp DNA band. The optimized concentrations of the PCR components were at 2.0 mM MgC12, 1.0 mM lOX PCR buffer, 0.2 mM Primer 15a and 15b, 0.5 U Taq-DNA polymerase and 0.4 mg bovine serum albumin (BSA). All tested virus DNA concentrations at 0.085, 0.170 and 0.255 g p0 were suitable for virus detection. Addition of BSA (20 mg ml) at 0.4 mg in the reaction increased the PCR sensitivity. The method is capable of detecting OrV infection from DNA diluted one million times or equivalent to a virus DNA concentration as low as 2.23 pg p0. The PCR detected 83.2% adult beetles from pheromone traps as being infected by OrV, 13.6% higher (P<0.05) than the results based on observations on the gut morphological appearance (69.6%). A typical OrV infection symptom is a swollen gut filled with milky fluid. Of the 839 guts with this symptom, 97.6% were diagnosed to be infected which was not significantly different (P>0.05) compared to the method based on gut morphological appearance. The PCR was also capable in detecting virus at an early infection stage and in dead adults with decayed tissues. Of the 307 adults that appeared to be healthy, 36.1% of them were found to be infected. As much as 61.6% of dead adults with decayed tissues (N = 428) were diagnosed to be infected by the OrV The method can he used in further research studies relating to OrV for the management of the rhinoceros beetle.

Original languageEnglish
Pages (from-to)736-749
Number of pages14
JournalJournal of Oil Palm Research
Volume22
Issue numberAPRIL
Publication statusPublished - Apr 2010

Fingerprint

Oryctes rhinoceros nudivirus
Baculoviridae
Polymerase chain reaction
Viruses
polymerase chain reaction
Polymerase Chain Reaction
DNA viruses
DNA Viruses
DNA
Beetles
bovine serum albumin
Bovine Serum Albumin
methodology
digestive system
Tissue
Taq Polymerase
infection
Coleoptera
Rhinoceros
viruses

Keywords

  • Oil palm pests
  • Oryctes rhinoceros virus
  • PCR
  • Rhinoceros beetle

ASJC Scopus subject areas

  • Agronomy and Crop Science
  • Food Science
  • Renewable Energy, Sustainability and the Environment
  • Biomaterials

Cite this

Optimization of the polymerase chain reaction (PCR) method for the detection of Oryctes rhinoceros virus. / Moslim, Ramle; Abd. Ghani, Idris; Wahid, Mohd Basri; Glare, Travis R.; Jackson, Trevor A.

In: Journal of Oil Palm Research, Vol. 22, No. APRIL, 04.2010, p. 736-749.

Research output: Contribution to journalArticle

Moslim, Ramle ; Abd. Ghani, Idris ; Wahid, Mohd Basri ; Glare, Travis R. ; Jackson, Trevor A. / Optimization of the polymerase chain reaction (PCR) method for the detection of Oryctes rhinoceros virus. In: Journal of Oil Palm Research. 2010 ; Vol. 22, No. APRIL. pp. 736-749.
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