First Detection of Honey Bee Viruses in Jordan by RT-PCR

Applied Entomology and Zoology, 2010

Using PCR in the direct diagnosis of bee virus infections has been shown to be an appropriate tool to overcome the difficulties of diagnosing bee virus infections. The occurrence of Israel acute paralysis virus (IAPV) and Kashmir bee virus (KBV) was investigated in honeybee colonies collected from ten different regions throughout Jordan by employing reverse transcription-PCR. The colonies were suffering from symptoms of depopulation, sudden collapse, paralysis, or dark coloring. The two viruses were identified in some of the collected samples: IAPV virus was present in 13% of samples, while KBV was present in 12%. The distribution of IAPV and KBV varied in the different geographic regions of investigation. The RT-PCR product for IAPV was 470-473 nucleotides, and for KBV was 408-409 nucleotides. Sequence analysis indicated that IAPV is most homologous to KBV.

Received on 8/3/2011 and Accepted for Publication on 8/8/2012. ABSTRACT The Reverse Transcription-polymerase Chain Reaction (RT-PCR) is an excellent technique for the detection of honeybee viruses. In this study, the presence of Sac Brood Virus (SBV) and Black Queen Cell Virus (BQCV) was demonstrated in 100 samples, collected from Jordanian honeybee colonies by employing RT-PCR. The collected samples represented infected (depopulation, paralysis, or dark coloring), dead, and apparently healthy honeybees of different developmental stages (adult and larvae). SBV was detected in 37% of the samples, whereas BQCV was detected in 5%. Nucleotide sequences of the PCR products from each virus was determined and found to be 433 and 309 nucleotides in SBV and BQCV, respectively. The identities to the Gene Bank were 95% for SBV and 91% for BQCV. This is the first record of BQCV in Jordan.

Jordan Journal of Agricultural Sciences, 2013

The Reverse Transcription-polymerase Chain Reaction (RT-PCR) is an excellent technique for the detection of honeybee viruses. In this study, the presence of Sac Brood Virus (SBV) and Black Queen Cell Virus (BQCV) was demonstrated in 100 samples, collected from Jordanian honeybee colonies by employing RT-PCR. The collected samples represented infected (depopulation, paralysis, or dark coloring), dead, and apparently healthy honeybees of different developmental stages (adult and larvae). SBV was detected in 37% of the samples, whereas BQCV was detected in 5%. Nucleotide sequences of the PCR products from each virus was determined and found to be 433 and 309 nucleotides in SBV and BQCV, respectively. The identities to the Gene Bank were 95% for SBV and 91% for BQCV. This is the first record of BQCV in Jordan.

Journal of the Institute of Science and Technology

Viruses infecting honey bees are diseases that cause infection and economic damage in every developmental stage of honey bees all over the world. In this study, a survey was conducted to determine the presence and prevalence of two worldwide common honeybee viruses (deformed wing virus [DWV] and chronic bee paralysis virus [CBPV]). 128 apiaries from 9 different localities in the province of Bingöl were visited to determine viral RNA. Collected 384 honey bee samples were tested molecularly (reverse-transcriptase polymerase chain reaction, RTPCR) using genome-specific primers specific to each virus. In molecular tests, 28 of 128 apiaries (21.87%) gave a positive reaction for DWV, but no CBPV pathogen was detected in any apiary. The 711 bp nucleotide sequence revealed by cloning one of the randomly selected positive samples was registered in the gene bank with the accession number MZ357973. According to the BLASTn analysis in the NCBI database, the nucleotide sequence of the DWV agent ...

Journal of Invertebrate Pathology, 2007

A single-step multiple-target (multiplex) reverse transcription-PCR (RT-PCR) was developed for the simultaneous detection and diVerentiation of three economically important viruses of the honeybee Apis mellifera L.: Acute bee paralysis virus (ABPV), Black queen cell virus (BQCV) and Sacbrood virus (SBV). Three compatible sets of primers, speciWc for each virus, were designed in conserved regions of the viral genomes for use in a one-step (single tube) RT-PCR assay. The individual RT-PCR assays and the combined multiplex assay were optimized for highest sensitivity and speciWcity. The multiplex RT-PCR assay was tested on Weld samples collected from Austrian honeybee colonies. All three viruses were detected, and their identity was conWrmed by sequencing of the PCR products. The described multiplex RT-PCR proved to be an accurate tool for rapid simultaneous detection of ABPV, BQCV and SBV directly in honeybee specimens.

Applied and Environmental Microbiology, 2001

A reverse transcriptase PCR (RT-PCR) assay was developed for the detection of acute bee paralysis virus (ABPV) and black queen cell virus (BQCV), two honeybee viruses. Complete genome sequences were used to design unique PCR primers within a 1-kb region from the 3′ end of both genomes to amplify a fragment of 900 bp from ABPV and 700 bp from BQCV. The combined guanidinium thiocyanate and silica membrane method was used to extract total RNA from samples of healthy and laboratory-infected bee pupae. In a blind test, RT-PCR successfully identified the samples containing ABPV and BQCV. Sensitivities were approximately 1,600 genome equivalents of purified ABPV and 130 genome equivalents of BQCV.

African Journal of Biotechnology, 2005

A single multiplex reverse transcriptase (RT) polymerase chain reaction (PCR) assay was developed for the simultaneous detection of three honeybee viruses: acute bee paralysis virus (ABPV), sacbrood virus (SBV) and black queen cell virus (BQCV). Unique PCR primers were designed from the complete genome sequence to amplify fragments of 900 bp from ABPV, 434 bp from SBV and 316 bp from BQCV. Individual bee pupae homogenates or total RNA extracted from these crude extracts were used in the RT-PCR amplification. Sequence analysis of the fragments amplified revealed nucleotide sequence identities between 97 and 98% for each virus against its reference strain. In a blind test, samples containing various combinations of ABPV, SBV and BQCV were successfully identified. Furthermore, field samples of honeybee pupae were screened for viral infections, and evidence of virus inapparent infection as well as virus co-infection were found.

Apidologie, 2002

A two years survey was undertaken to determine the occurrence of Acute Bee Paralysis Virus (ABPV) in field samples of adult bees and the parasitic mite Varroa destructor. To detect the viral nucleic acid we used polymerase chain reaction following reverse transcription (RT-PCR). We demonstrated the presence of ABPV RNA in 14 from 114 seemingly healthy colony samples collected from Hungarian honey bees. The investigation revealed that two third of the apiaries were infected with ABPV at a 12.2% infection rate. In seven other apiaries out of eight investigated (87.5%) the presence of the virus was detected from colonies following a sudden collapse; these colonies were simultaneously infected with Nosema apis or infested with Varroa destructor. Virus specific nucleic acid was also identified in the mites collected from two apiaries falling into the latter category. The amplicon of RT-PCR was sequenced and the nucleic acid sequence was aligned to the complete ABPV sequence deposited in the GeneBank database revealing a 93% homology.

Journal of Virological Methods, 2011

Viruses

In recent years, there has been growing evidence that certain types of honeybee viruses could be transmitted between different pollinators. Within a voluntary monitoring programme, 180 honeybee samples (Apis mellifera carnica) were collected from affected apiaries between 2007 and 2018. Also from August 2017 to August 2018, a total 148 samples of healthy bumblebees (Bombus lapidarius, B. pascuorum, B. terrestris, B. lucorum, B. hortorum, B. sylvarum, B. humilis) were collected at four different locations in Slovenia, and all samples were tested by using RT-PCR methods for six honeybee viruses. Direct sequencing of a total 158 positive samples (acute bee paralysis virus (ABPV n = 33), black queen cell virus (BQCV n = 75), sacbrood bee virus (SBV n = 25) and Lake Sinai virus (LSV n = 25)) was performed from obtained RT-PCR products. The genetic comparison of identified positive samples of bumblebees and detected honeybee field strains of ABPV, BQCV, SBV, and LSV demonstrated 98.74% to...