Occurrence and prevalence of seven bee viruses in Apis mellifera and Apis cerana apiaries in China

Populations of Apis mellifera and Apis cerana in China were surveyed for seven bee viruses: acute bee paralysis virus (ABPV), black queen cell virus (BQCV), chronic bee paralysis virus (CBPV), deformed wing virus (DWV), Kashmir bee virus (KBV), sacbrood virus (SBV), and Isreal acute paralysis virus (IAPV). No KBV was detected from any samples of the two species. In A. mellifera, DWV was the most prevalent virus, but in A. cerana, SBV was the dominant. Simultaneous multiple infections of viruses were common in both species. This is the first report of detection of IAPV and CBPV in A. cerana.     

北京地区六种蜜蜂病毒病的流行病学研究

【目的】对蜜蜂的6种病毒:以色列急性麻痹病毒(Israeli acute paralysis virus,IAPV)、残翅病毒(Deformed wing virus,DWV)、囊状幼虫病病毒(Sacbrood virus,SBV)、急性蜜蜂麻痹病毒(Acute bee paralysis virus,ABPV)、黑蜂王台病毒(Black queen cell virus,BQCV)、慢性麻痹病毒(Chronic bee paralysis virus,CBPV)在北京地区的流行情况进行调查,以期为该地区蜜蜂病毒病的防控提供一定的理论依据。【方法】应用多重RT-PCR法确定上述6种病毒在该地区的感染情况,并通过序列分析确定特异性。【结果】在所有检测样本中均未检测到急性麻痹病病毒和慢性麻痹病病毒,感染率最高的是以色列急性麻痹病毒,其次是残翅病毒。检测的样本普遍存在混合感染。【结论】以色列急性麻痹病毒、残翅病毒、囊状幼虫病病毒、黑蜂王台病毒4种病毒可能在北京地区广泛分布。     

Prevalence of the microsporidian Nosema ceranae in honeybee (Apis mellifera) apiaries in Central Italy

Nosema ceranae and Nosema apis are microsporidia which play an important role in the epidemiology of honeybee microsporidiosis worldwide. Nosemiasis reduces honeybee population size and causes significant losses in honey production. To the best of our knowledge, limited information is available about the prevalence of nosemiasis in Italy. In this research, we determined the occurrence of Nosema infection in Central Italy. Thirty-eight seemingly healthy apiaries (2 to 4 hives each) were randomly selected and screened from April to September 2014 (n = 11) or from May to September 2015 (n = 27). The apiaries were located in six areas of Central Italy, including Lucca (n = 11), Massa Carrara (n = 9), Pisa (n = 9), Leghorn (n = 7), Florence (n = 1), and Prato (n = 1) provinces. Light microscopy was carried out according to current OIE recommendations to screen the presence of microsporidiosis in adult worker honeybees. Since the morphological characteristics of N. ceranae and N. apis spores are similar and can hardly be distinguished by optical microscopy, all samples were also screened by multiplex polymerase chain reaction (M-PCR) assay based on 16S rRNA-gene-targeted species-specific primers to differentiate N. ceranae from N. apis. Furthermore, PCR-positive samples were also sequenced to confirm the species of amplified Nosema DNA. Notably, Nosema spores were detected in samples from 24 out of 38 (63.2%, 95% CI: 47.8–78.5%) apiaries. Positivity rates in single provinces were 10/11, 8/9, 3/9, 1/7, or 1/1 (n = 2). A full agreement (Cohen's Kappa = 1) was assessed between microscopy and M-PCR. Based on M-PCR and DNA sequencing results, only N. ceranae was found. Overall, our results highlighted that N. ceranae infection occurs frequently in the cohort of honeybee populations that was examined despite the lack of clinical signs. These findings suggest that colony disease outbreaks might result from environmental factors that lead to higher susceptibility of honeybees to this microsporidian.     

Prevalence of pathogenic bee viruses in Hungarian apiaries: situation before joining the European Union

A survey on the occurrence of six honeybee-pathogenic viruses was carried out using one-step RT-PCR assays. Samples were collected between 1999 and 2004 in 52 Hungarian apiaries located in different regions of the country. The results of the assays on samples of adult honeybees and Varroa destructor mites were compared to similar surveys from France and Austria. The study demonstrates geographical differences in the prevalence of honeybee viruses between Hungary and the older EU member states. The results could serve as a basis for monitoring further changes in the distribution of honeybee viruses in Europe.     

Prevalence of honeybee viruses in the Czech Republic and coinfections with other honeybee disease

Six bee viruses, which occur in Apis mellifera, were monitored in the Czech Republic between 2006 and 2009. Samples of larvae and pupae collected from hives where American foulbrood was detected were screened for bee viruses and in the 125 samples of larvae, there was no confirmed case of a larva infected with both American foulbrood and a bee virus. Of 145 samples infected with the protozoan Nosema apis, there were 23 cases of coinfections with the BQCV virus, 18 with the DWV virus and 11 with the ABPV virus. All coinfections with three or four viruses were also statistically significant apart from the one between ABPV with CBPV and DWV. The PCA ordination diagram indicates that BQCV occurs mainly with Nosema apis and DWV mainly with ABPV.     

Occurrence of Malassezia species in healthy and dermatologically diseased dogs

The presence of Malassezia spp. yeasts was investigated in dermatological specimens of 224 dogs, 164 dermatologically diseased and 60 normal dogs. Subjects included in the study were of different breed, age, sex and habitat. Malassezia spp. positive cultures were obtained in 142 (63.4%) specimens: 67.6% from dermatologically diseased subjects and 51.6% from healthy dogs. Malassezia pachydermatis, either as a pure culture or in association with lipid-dependent species, was identified in 138 (97%) specimens. Malassezia furfur was identified in 69 (48.6%) specimens and was associated with other Malassezia species in 68 dogs, as a pure culture in one subject: at the best of our knowledge, this species was identified before as the sole species from canine dermatitis. Malassezia sympodialis was identified in 11 (7.7%) specimens, always in association with other species: it was never isolated from kennel dogs. Statistical analysis of data showed a very significant difference (P < 0.01) in the prevalence of isolation of Malassezia spp. between animals with and without dermatological signs, and in the distribution of cultural burden between diseased and healthy dogs. A statistically significant difference (P<0.05) was also detected in the group of animals between 1- and 5-years of age. No significant difference was found between male and female dogs.     

Occurrence of proctolin in six orders of insects

Nine representatives of six orders of insects (Orthoptera, Diptera, Coleoptera, Hemiptera, Hymenoptera, Lepidoptera) were extracted and partially processed by means used in the recent isolation of proctolin, a pentapeptide transmitter candidate in insects. Each insect yielded a substance with pharmacological activity on cockroach proctodeal muscle similar to that of proctolin. Like the responses evoked by proctolin and nerve stimulation, responses to the purified extracts were inhibited by tyramine. All of the active substances behaved as proctolin when subjected to paper chromatography or high voltage paper electrophoresis at pH 6.4 and 3.5. Proctolin appears to be widely and perhaps universally present in the Insecta occurring in most at levels of 2 to 9 μg/kg body weight.     

Abundance and diversity of viruses in six Delaware soils

The importance of viruses in marine microbial ecology has been established over the past decade. Specifically, viruses influence bacterial abundance and community composition through lysis and alter bacterial genetic diversity through transduction and lysogenic conversion. By contrast, the abundance and distribution of viruses in soils are almost completely unknown. This study describes the abundance and diversity of autochthonous viruses in six Delaware soils: two agricultural soils, two coastal plain forest soils, and two piedmont forest soils. Viral abundance was measured using epifluorescence microscopy, while viral diversity was assessed from morphological data obtained through transmission electron microscopy. Extracted soil virus communities were dominated by bacteriophages that demonstrated a wide range of capsid diameters (20 nm to 160 nm) and morphologies, including filamentous forms and phages with elongated capsids. The reciprocal Simpson's index suggests that forest soils harbor more diverse assemblages of viruses, particularly in terms of morphological distribution. Repeated extractions of virus-like particles (VLPs) from soils indicated that the initial round of extraction removes approximately 70% of extractable viruses. Higher VLP abundances were observed in forest soils (1.31 x 10(9) to 4.17 x 10(9) g(-1) dry weight) than in agricultural soils (8.7 x 10(8) to 1.1 x 10(9) g(-1) dry weight). Soil VLP abundance was significantly correlated to moisture content (r = 0.988) but not to soil texture. Land use (agricultural or forested) was significantly correlated to both bacterial (r = 0.885) and viral (r = 0.812) abundances, as were soil organic matter and water content. Thus, land use is a significant factor influencing viral abundance and diversity in soils.     

A comparison of serological techniques for detecting and identifying honeybee viruses

The sensitivity and specificity of conventional Ouchterlony gel-diffusion, immuno-osmoelectrophoresis (IO), immune serum electron microscopy (ISEM), “decoration,” radioimmunoassay (RIA), and enzyme-linked immunosorbent assay (ELISA) tests for detecting black queen cell virus (BQCV), chronic bee paralysis virus (CBPV), Kashmir bee virus (KBV), and sacbrood virus (SBV) particles in extracts of diseased honeybees were compared. A “slow” ISEM method detected virus particles in extracts of individuals or groups of individuals diluted to 10^?3 and 10^?4, respectively, whereas the IO method and a “fast” ISEM method using protein A were one-tenth as sensitive, and Ouchterlony gel-diffusion tests were only one-thousandth as sensitive. Using the antibody “decoration” technique, mixtures of serologically unrelated virus particles could be resolved. RIA and ELISA were found to be one thousand times more sensitive than ISEM in detecting the particles of BQCV, CBPV, KBV, and SBV; however, nonspecific reactions occurred when using RIA with very dilute particle suspensions, and this made dilution endpoints difficult to assess, but this did not occur when using the ELISA method. There was little difference in the effectiveness of rabbit or hen antisera in the tests, except when protein A was used as it does not combine with hen antibodies.     

Occurrence and distribution of viruses infecting potato in Russia

Potato viral disease has been a major problem in potato production worldwide including Russia. Here, we detected Potato Virus M (PVM), P (PVP), S (PVS), Y (PVY), and X (PVX) and Potato Leaf Roll Virus (PLRV) by RT-PCR on potato leaves and tubers from the Northwestern (NW), Volga (VF), and Far Eastern (FE) federal districts of Russia. Each sample was co-infected with up to five viruses. RT-PCR disclosed all six viruses in NW, three in VF, and five in FE. Phylogenetic analyses of PVM and PVS strains resolved all PVM isolates in Group O (ordinary) and all PVS isolates in Group O. Seven PVY strains were detected, and they included only recombinants. PVY recombinants were thus the dominant potato virus strains in Russia, although they widely varied among the regions. Our research provides insights into the geographical distribution and genetic variability of potato viruses in Russia.     

Occurrence of temperature-sensitive influenza A viruses in nature.

The origin and characteristics of the first naturally occurring temperature-sensitive (ts) strain of influenza A virus identified in 1973, Xia-ts, are described. Natural ts strains were found to occur in the early egg passage material of all influenza A subtypes examined, but the proportion of ts virus varied from 8.3% for old H1N1 virus (1949 to 1957) to 82.4% for recent H3N2 virus (1979 to 1980). A number of strains were found to be composed of a mixture of ts and wild-type (ts+) particles. Six natural ts strains with different shutoff temperatures and one ts+ strain of the H1N1 subtype were tested in antibody-free volunteers. Strains with a shutoff temperature of 38 degrees C or lower caused very mild symptoms, whereas those with a shutoff temperature of 39 degrees C and the ts+ strain were much more reactogenic. By complementation tests against a set of prototype WSN ts mutants with a defined genetic lesion, the ts lesion of two H3N2 viruses (HK/8/68 and Xia-ts) was located on the NP gene and that of two H1N1 viruses (Tianjin/78/77 and Beijing/1/79) was located on the M protein gene. The present study demonstrates the widespread occurrence in nature of influenza viruses of different degrees of temperature sensitivity and presumably of different degrees of virulence.     

Occurrence of fungally transmitted wheat mosaic viruses in China

A soil-borne mosaic disease of winter wheat in Sichuan, Shaanxi, Hubei and Henan provinces was associated with infection by a virus with filamentous particles and that in Shandong, Anhui, Jiangsu and Zhejiang provinces by co-infection with this virus and soil-borne wheat mosaic virus. The virus with filamentous particles was identified serologically, by particle sizes, cytopathology and the molecular weights of the coat protein and the two RNA species to be either wheat spindle streak mosaic virus (WSSMV) or wheat yellow mosaic virus. These viruses are probably isolates of the same virus and the name WSSMV is preferred. In baiting tests using infested soil, the dilution endpoints for detecting WSSMV were 1/625-1/15625, and for the fungus vector, Polymyxa graminis, 1/3125-1/15625.     

Survey of six bee viruses using RT-PCR in Northern Thailand

Six honey bee viruses were surveyed using RT-PCR in Northern Thailand where about 80% of Thai apiaries are located. Tested samples were found to be positive for deformed wing virus (DWV), acute bee paralysis virus (ABPV), sacbrood virus (SBV) and Kashmir bee virus (KBV). In the collected samples, neither chronic bee paralysis virus nor black queen cell virus nucleic acids could be detected. It was found that DWV was the most widespread and ABPV was the second most prevalent. Kashmir bee virus was found only in the Lampang province where high infestation of Varroa destructor mite occurred. Tropilaelaps, European foulbrood, and Chalkbrood diseases were found in some apiaries.     

中缅树鼩自然感染六种病毒的血清流行病学

中缅树鼩作为一种新型实验动物,在医学生物学上,尤其是病毒学方面的应用受到越来越多的重视.实验动物自身病毒感染会影响动物健康和干扰实验结果,甚至危害实验人员生命安全.所以,实验动物病毒检测一直是动物质量控制的重要部分.中缅树鼩研究迄今缺乏清晰的病毒自然感染资料.为调查中缅树鼩的病毒感染状况,采集野生俘获和人工繁殖的中缅树鼩血清样本272份,全部血清样本通过ELISA方法对乙型肝炎病毒(HBV)表面抗原,丙型肝炎病毒(HCV)总抗体,以及戊型肝炎病毒(HEV)、腺病毒(ADV)、单纯疱疹病毒1型(HSV-1)和2型(HSV-2)的IgG抗体进行了检测.结果表明,ELISA初筛HBV表面抗原有3份阳性样本,但通过乙型肝炎两对半定量检测进一步确认为阴性:抗HCV抗体和抗HEV、ADV、HSV-1 IgO抗体检测均为阴性;抗HSV-2 IgG检测有1份阳性样本.提示仪抗原或抗体血清学指标检测树鼩肝炎结果并不能反应个体携带病毒的状态,应该再进行病毒学指标确认.同时建议中缅树嗣繁殖群应进行HSV-2的筛选,以便杜绝和控制该病毒的感染.     

The prevalence of the honeybee brood pathogens Ascosphaera apis,Paenibacillus larvae and Melissococcus plutonius in Spanish apiaries determined with a new multiplex PCR assay

The microorganisms Ascosphaera apis, Paenibacillus larvae and Melissococcus plutonius are the three most important pathogens that affect honeybee brood. The aim of the present study was to evaluate the prevalence of these pathogens in honeybee colonies and to elucidate their role in the honeybee colony losses in Spain. In order to get it, a multiplex polymerase chain reaction (PCR) assay was developed to simultaneously amplify the16S ribosomal ribonucleic acid (rRNA) gene of P. larvae and M. plutonius, and the 5.8S rRNA gene of A. apis. The multiplex PCR assay provides a quick and specific tool that successfully detected the three infectious pathogens (P. larvae, M. plutonius and A. apis) in brood and adult honeybee samples without the need for microbiological culture. This technique was then used to evaluate the prevalence of these pathogens in Spanish honeybee colonies in 2006 and 2007, revealing our results a low prevalence of these pathogens in most of the geographic areas studied.     

Varroa mites in the apiaries of Campania region

Mites in the genus Varroa are obligate ectoparasites of honey bee populations worldwide. Recent evidence from morphological, geographical, and especially genetic variation has spurred an important revision of Varroa taxonomy. Specifically, mitochondrial DNA (mtDNA) evidence suggests that the main mite pest on western honey bees (Apis mellifera) is not Varroa jacobsoni, as first described, but a distinct species now named Varroa destructor. Genetic markers also have been used to support a taxonomic basis for regional differences in how Varroa mites impact honey bees. Recent morphometric and molecular studies confirmed the presence of the species V. destructor also in the apiaries of the Campania region of southern Italy. In the three-year period 2001-2003 a survey was conducted in 118 municipalities of the five provinces of the Campania region in order to add data to the limited epidemiological information available regarding Varroa destructor in this zone. The level of infestation by the mite was assessed on a total of 521 apiaries (241 apiaries were inspected on 2001, 154 on 2002, and 126 on 2003). In each apiary, 100 comb cells were examined and in each province the level of infestation was calculated using the following formula: (number of Varroa specimens/number of open comb cells) x 100. In order to display the level of infestation, Geographical Information Systems were used in order to draw parasitological maps.     

Occurrence of papaya ringspot potyvirus and cucurbit viruses in Nepal

A survey of papaya and 10 cucurbitaceous vegetables (ashgourd, zucchini, watermelon, cucumber, pumpkin, bottlegourd, snakegourd, spongegourd, bittergourd and choyote) during 1989 and from 1992 to 1994 in more than 68 locations (both experimental plots and farmers' fields) covering 18 terai and inner-terai districts of Nepal, indicated that these crops were heavily affected with various virus-like symptoms. The most commonly observed symptoms were severe mosaic, leaf distortion, oily streaks or spots on papaya; leaf distortion, blisters and shoe stringing on zucchini; and mosaic or yellow mosaic, blisters, and leaf distortion on other cucurbits. Average incidence of plants with symptoms ranged from 75% to 100% on papaya; 85% to 100% on zucchini; 4% to 100% on cucumber; 4% to 100% on pumpkin and 10–100% on bottlegourd, choyote and watermelon. The virus isolated from papaya and zucchini was confirmed as papaya ringspot potyvirus — watermelon strain (PRSV-W). It was also detected in survey samples from ashgourd, bittergourd, snakegourd, spongegourd, zucchini, watermelon, bottlegourd and cucumber. Leaf extracts of some cucumber, choyote, pumpkin, zucchini and snakegourd samples reacted with cucumber mosaic cucumovirus (CMV) and zucchini yellow mosaic potyvirus (ZYMV) antisera. Leaf extracts of ashgourd, cucumber and pumpkin reacted with antibodies against cucurbit aphid-borne yellow luteovirus (CABW). No samples reacted with antiserum to watermelon mosaic-2 potyvirus (WMV-2) or squash mosaic potyvirus (SqMV). Some papaya and most cucurbits leaf samples cross-reacted with antibodies against Moroccan (Mor) and Algerian (Alg) isolates of WMV. The Nepalese PRSV isolate was related to but distinct from a PRSV-W type strain from France. This is the first report on the identity of ZYMV and CABW in Nepal.