Detection of Lotmaria passim, Crithidia mellificae and Replicative Forms of Deformed Wing Virus and Kashmir Bee Virus in the Small Hive Beetle (Aethina tumida)

Knowledge regarding the honey bee pathogens borne by invasive bee pests remains scarce. This investigation aimed to assess the presence in Aethina tumida (small hive beetle, SHB) adults of honey bee pathogens belonging to the following groups: (i) bacteria (Paenibacillus larvae and Melissococcus plutonius), (ii) trypanosomatids (Lotmaria passim and Crithidia mellificae), and (iii) viruses (black queen cell virus, Kashmir bee virus, deformed wing virus, slow paralysis virus, sacbrood virus, Israeli acute paralysis virus, acute bee paralysis virus, chronic bee paralysis virus). Specimens were collected from free-flying colonies in Gainesville (Florida, U.S.A.) in summer 2017. The results of the molecular analysis show the presence of L. passim, C. mellificae, and replicative forms of deformed wing virus (DWV) and Kashmir bee virus (KBV). Replicative forms of KBV have not previously been reported. These results support the hypothesis of pathogen spillover between managed honey bees and the SHB, and these dynamics require further investigation.

Virion structure and in vitro genome release mechanism of dicistrovirus Kashmir bee virus

Infections of Kashmir bee virus (KBV) are lethal for honeybees and have been associated with colony collapse disorder. KBV and closely related viruses contribute to the ongoing decline in the number of honeybee colonies in North America, Europe, Australia, and other parts of the world. Despite the economic and ecological impact of KBV, its structure and infection process remain unknown. Here we present the structure of the virion of KBV determined to a resolution of 2.8 Å. We show that the exposure of KBV to acidic pH induces a reduction in inter-pentamer contacts within capsids and the reorganization of its RNA genome from a uniform distribution to regions of high and low density. Capsids of KBV crack into pieces at acidic pH, resulting in the formation of open particles lacking pentamers of capsid proteins. The large openings of capsids enable the rapid release of genomes and thus limit the probability of their degradation by RNases. The opening of capsids may be a shared mechanism for the genome release of viruses from the family Dicistroviridae. Importance The western honeybee (Apis mellifera) is indispensable for maintaining agricultural productivity as well as the abundance and diversity of wild flowering plants. However, bees suffer from environmental pollution, parasites, and pathogens, including viruses. Outbreaks of virus infections cause the deaths of individual honeybees as well as collapses of whole colonies. Kashmir bee virus has been associated with colony collapse disorder in the US, and no cure of the disease is currently available. Here we report the structure of an infectious particle of Kashmir bee virus and show how its protein capsid opens to release the genome. Our structural characterization of the infection process determined that therapeutic compounds stabilizing contacts between pentamers of capsid proteins could prevent the genome release of the virus.

Resistance of native honey bees from Rhodope Mountains and lowland regions of Bulgaria to Nosema ceranae and viral pathogens

The Western honey bee (Apis mellifera L., Hymenoptera: Apidae) is a species of fundamental economic, agricultural and environmental importance. The aim of this study was to compare the prevalence of some parasitic and viral pathogens in local honey bees from the Rodope Mountains and plain regions. To achieve this goal, molecular screening for two of the most distributed Nosema spp. and molecular identification of six honey bee viruses – Deformed wing virus (DWV), Acute bee paralysis virus (ABPV), Chronic bee paralysis virus (CBPV), Sacbrood virus (SBV), Kashmir bee virus (KBV), and Black queen cell virus (BQCV) was performed. Molecular analysis was carried out on 168 honey bee samples from apiaries situated in three different parts of the country where a mix of different honey bee subspecies were reared. In South Bulgaria (the Rhodope Mountains), a local honey bee called Apis mellifera rodopica (a local ecotype of A. m. macedonica) was bred, while in the other two regions (plains) different introduced subspecies existed. The results showed that the samples from the lowland regions in the country were outlined with the highest prevalence (70.5%) of N. ceranae, while those from the mountainous parts had the lowest rate (5.2%). Four of the honey bee viruses were identified – DWV (10/5.9%), followed by SBV (6/3.6%) and ABPV (2/1.2%), and one case of BQCV. In conclusion, the local honey bee A. m. rodopica (despite the higher number of samples) has shown lower prevalence of both nosemosis and viral infections. Therefore, this honey bee has to be preserved as a part of the national biodiversity.

Molecular detection and phylogenetic assessment of six honeybee viruses in Apis mellifera L. colonies in Bulgaria

Honey bee colonies suffer from various pathogens, including honey bee viruses. About 24 viruses have been reported so far. However, six of them are considered to cause severe infection which inflicts heavy losses on beekeeping. The aim of this study was to investigate incidence of six honey bee viruses: deformed wing virus (DWV), acute bee paralysis virus (ABPV), chronic bee paralysis virus (CBPV), sacbrood virus (SBV), kashmir bee virus (KBV), and black queen cell virus (BQCV) by a reverse transcription polymerase chain reaction (RT-PCR). A total of 250 adult honey bee samples were obtained from 50 colonies from eight apiaries situated in three different parts of the country (South, North and West Bulgaria). The results showed the highest prevalence of DWV followed by SBV and ABPV, and one case of BQCV. A comparison with homology sequences available in GenBank was performed by phylogenetic analysis, and phylogenetic relationships were discussed in the context of newly described genotypes in the uninvestigated South Eastern region of Europe. In conclusion, the present study has been the first to provide sequencing data and phylogenetics analyses of some honey bee viruses in Bulgaria.

The first molecular detection and phylogenetic assessment of six honeybee viruses in Apis mellifera L. colonies in Bulgaria

Honey bee colonies suffer from various pathogens, including honey bee viruses. About 24 viruses have been reported so far. However, six of them are considered to cause severe infection which inflicts heavy losses on beekeeping. The aim of this study is to detect six honey bee viruses: deformed wing virus (DWV), acute bee paralysis virus (ABPV), chronic bee paralysis virus (CBPV), sacbrood virus (SBV), kashmir bee virus (KBV), and black queen cell virus (BQCV) by a Reverse transcription polymerase chain reaction (RT-PCR). A total of 50 adult honey bee samples were obtained from apiaries situated in three different parts of the country (South, North and West Bulgaria).The results showed the highest prevalence of DWV (10/20 %), followed by SBV (6/12 %) and ABPV (2/4%), and one case of BQCV. A comparison with homology sequences available in GenBank was performed by phylogenetic analysis, and phylogenetic relationships were discussed in the context of newly described genotypes in the uninvestigated South Eastern region of Europe.In conclusion, the present study has been the first to provide sequencing data and phylogenetics analyses of some honey bee viruses in Bulgaria.

The first molecular detection and phylogenetic assessment of six honeybee viruses in Apis mellifera L. colonies in Bulgaria

Honey bee colonies suffer from various pathogens, including honey bee viruses. About 24 viruses have been reported so far. However, six of them are considered to cause severe infection which inflicts heavy losses on beekeeping. The aim of this study is to detect six honey bee viruses: deformed wing virus (DWV), acute bee paralysis virus (ABPV), chronic bee paralysis virus (CBPV), sacbrood virus (SBV), kashmir bee virus (KBV), and black queen cell virus (BQCV) by a Reverse transcription polymerase chain reaction (RT-PCR). A total of 50 adult honey bee samples were obtained from apiaries situated in three different parts of the country (South, North and West Bulgaria).The results showed the highest prevalence of DWV (10/20 %), followed by SBV (6/12 %) and ABPV (2/4%), and one case of BQCV. A comparison with homology sequences available in GenBank was performed by phylogenetic analysis, and phylogenetic relationships were discussed in the context of newly described genotypes in the uninvestigated South Eastern region of Europe.In conclusion, the present study has been the first to provide sequencing data and phylogenetics analyses of some honey bee viruses in Bulgaria.

Prevalence of common honey bee pathogens at selected apiaries in Kenya, 2013/2014

The present study was part of a larger surveillance effort to identify the determinants of African honey bee health, and, particularly, to detect honey bee pathogens across Kenya, where 160 colonies were examined from 32 apiaries (five colonies/apiary). From each colony, 20 individual foragers, nurse bees, worker pupae, and drone pupae were sampled separately. These were organized as 30 foragers, 32 nurse bees, 28 worker pupae, and 10 drone pupae pools. Nucleic acid was extracted from the pooled homogenates and tested using a panel of 18 different (RT-)PCR methods targeted at detectingPaenibacillus larvae,Melissococcus plutonius,Ascophaera apis,Aspergillusspp.,Nosema ceranae, N. apis,Deformed wing virus(DWV),Varroa destructor virus 1(VDV 1),Acute bee paralysis virus(ABPV),Sacbrood virus(SBV),Israeli acute paralysis virus(IAPV),Black queen cell virus(BQCV),Chronic bee paralysis virus(CBPV), andKashmir bee virus. All amplified bands were sequenced and compared to the GenBank database. VDV 1 was the most abundant virus at 50% prevalence in the 100 bee pools. It was closely followed by DWV at 44%. The others were BQCV (36%), SBV (14%), IAPV (9%), ABPV (8%), andN. ceranae(5%). The pathogens co-existed within apiaries. VDV 1 was present in 66% of the apiaries, DWV in 69%, BQCV in 69%, SBV in 28%, IAPV in 22%, ABPV in 19%, andN. ceranaein 13%. The study concludes that these pathogens should be incorporated in honey bee disease surveillance activities in the region.