Reliability of Morphological and PCR Methods for the Official Diagnosis of <i>Aethina tumida</i> (Coleoptera: Nitidulidae): A European Inter-Laboratory Comparison

Simple Summary Aethina tumida, also called the Small Hive Beetle, is an insect that multiplies primarily in honeybee hives, causing honey losses and weakening colonies. It is native to sub-Saharan Africa and was introduced into different countries and continents over the last 20 years, posing a threat to beekeeping internationally. In case of introduction into a new area, officially approved laboratories (certified by government services) carry out analyses to confirm the outbreak. The reliability of the results is essential in the implementation of management measures. Therefore, a study was organised at the European level to compare the results between official laboratories for two types of methods, used routinely for the identification of A. tumida: morphological examination (form and structure) and DNA testing (genetics). The 22 participants analysed in a blinded way a panel of 12 samples (positive and negative samples). The results were very satisfactory, with the exception of one participant who encountered several anomalies for negative samples and especially for DNA tests, probably related to his inexperience with the method. This study proved the ability of laboratories and analytical methods to identify A. tumida, which is a key element in monitoring and managing this risk. The Small Hive Beetle (Aethina tumida Murray, 1867) is an invasive scavenger of honeybees. Originally endemic in sub-Saharan Africa, it is regulated internationally in order to preserve the areas still free from this species. To ensure the reliability of official diagnoses in case of introduction, an inter-laboratory comparison was organised on the identification of A. tumida by morphology and real-time PCR. Twenty-two National Reference Laboratories in Europe participated in the study and analysed 12 samples with adult coleopterans and insect larvae. The performance of the laboratories was evaluated in terms of sensitivity and specificity. Sensitivity was satisfactory for all the participants and both types of methods, thus fully meeting the diagnostic challenge of confirming all truly positive cases as positive. Two participants encountered specificity problems. For one, the anomaly was minor whereas, for the other, the issues concerned a larger number of results, especially real-time PCR, which probably were related to inexperience with this technique. The comparison demonstrated the reliability of official diagnosis, including the entire analytical process of A. tumida identification: from the first step of the analysis to the expression of opinions. The performed diagnostic tools, in parallel with field surveillance, are essential to managing A. tumida introduction

Shift in virus composition in honeybees (Apis mellifera) following worldwide invasion by the parasitic mite and virus vector Varroa destructor

Invasive vectors can induce dramatic changes in disease epidemiology. While viral emergence following geographical range expansion of a vector is well known, the influence a vector can have at the level of the host's pathobiome is less well understood. Taking advantage of the formerly heterogeneous spatial distribution of the ectoparasitic mite Varroa destructor that acts as potent virus vector among honeybees Apis mellifera, we investigated the impact of its recent global spread on the viral community of honeybees in a retrospective study of historical samples. We hypothesized that the vector has had an effect on the epidemiology of several bee viruses, potentially altering their transmissibility and/or virulence, and consequently their prevalence, abundance, or both. To test this, we quantified the prevalence and loads of 14 viruses from honeybee samples collected in mite-free and mite-infested populations in four independent geographical regions. The presence of the mite dramatically increased the prevalence and load of deformed wing virus, a cause of unsustainably high colony losses. In addition, several other viruses became more prevalent or were found at higher load in mite-infested areas, including viruses not known to be actively varroa-transmitted, but which may increase opportunistically in varroa-parasitized bees

SLU:s verksamhet med bin

Under 2014/15 var vi sammanlagt sju personer i den grupp på SLU som arbetar med olika aspekter på hälsa och sjukdomar hos honungsbin. Gruppen utökades under sommaren 2014 med två nya forskarstuderande/doktorander; Sepideh Lamei som ska studera mjölksyrabakteriers betydelse för bins hälsa, och Srinivas Thaduri som ska studera sammansättningen av mikroorganismer hos den gotländska population av bin som överlevt och klarar sig utan behandling mot varroakvalstret (de så kallade Bondbina). En kortfattad beskrivning av deras respektive projekt kan ni läsa om längre ned i texten

Virus Diversity and Loads in Crickets Reared for Feed: Implications for Husbandry

Insects generally have high reproductive rates leading to rapid population growth and high local densities; ideal conditions for disease epidemics. The parasites and diseases that naturally regulate wild insect populations can also impact when these insects are produced commercially, on farms. While insects produced for human or animal consumption are often reared under high density conditions, very little is known about the microbes associated with these insects, particularly those with pathogenic potential. In this study we used both target-free and targeted screening approaches to explore the virome of two cricket species commonly reared for feed and food, Acheta domesticus and Gryllus bimaculatus. The target-free screening of DNA and RNA from a single A. domesticus frass sample revealed that only 1% of the nucleic acid reads belonged to viruses, including known cricket, insect, bacterial and plant pathogens, as well as a diverse selection of novel viruses. The targeted screening revealed relatively high levels of Acheta domesticus densovirus, invertebrate iridovirus 6 and a novel iflavirus, as well as low levels of Acheta domesticus volvovirus, in insect and frass samples from several retailers. Our findings highlight the value of multiple screening approaches for a comprehensive and robust cricket disease monitoring and management strategy. This will become particularly relevant as-and-when cricket rearing facilities scale up and transform from producing insects for animal feed to producing insects for human consumption

Genome Characterization, Prevalence and Distribution of a Macula-Like Virus from Apis mellifera and Varroa destructor

Around 14 distinct virus species-complexes have been detected in honeybees, each with one or more strains or sub-species. Here we present the initial characterization of an entirely new virus species-complex discovered in honeybee (Apis mellifera L.) and varroa mite (Varroa destructor) samples from Europe and the USA. The virus has a naturally poly-adenylated RNA genome of about 6500 nucleotides with a genome organization and sequence similar to the Tymoviridae (Tymovirales; Tymoviridae), a predominantly plant-infecting virus family. Literature and laboratory analyses indicated that the virus had not previously been described. The virus is very common in French apiaries, mirroring the results from an extensive Belgian survey, but could not be detected in equally-extensive Swedish and Norwegian bee disease surveys. The virus appears to be closely linked to varroa, with the highest prevalence found in varroa samples and a clear seasonal distribution peaking in autumn, coinciding with the natural varroa population development. Sub-genomic RNA analyses show that bees are definite hosts, while varroa is a possible host and likely vector. The tentative name of Bee Macula-like virus (BeeMLV) is therefore proposed. A second, distantly related Tymoviridae-like virus was also discovered in varroa transcriptomes, tentatively named Varroa Tymo-like virus (VTLV)

Tracing probiotics in salami using PCR

Starter cultures of different bacteria strains like lactic acid producing bacteria, Staphylococcus and Kocuria are used when making salami. Starter cultures give the sausage specific flavours and improve the quality and ripening of the final product. Probiotic strains can also be added during the production of salami. Studies have shown that probiotics are good for health and are therefore added to food, such as fermented sausages. In order to work as a probiotic strain, the bacteria have to survive during the production process, storage and through the whole human gastrointestinal tract. The aim of this study was to trace the probiotic strains Lactobacillus casei and Lactobacillus paracasei in salami samples to see if they had survived the production process. Methods used were DNA extraction, PCR, colony PCR and gel electrophoresis. Out of 100 samples in duplicate run in PCR, probiotics were found in only 3 of them. To see if screening of probiotics directly from plates was possible, a colony PCR was done. Colony PCR was made on colonies of two different strains of Lactobacillus casei, Lactobacillus paracasei and Lactobacillus sakei. From each bacteria strain, 5 colonies were analysed. Result showed that colony PCR, to screen for probiotic is a possible method

Persistence of subclinical deformed wing virus infections in honeybees following <i>Varroa</i> mite removal and a bee population turnover

<div><p>Deformed wing virus (DWV) is a lethal virus of honeybees (<i>Apis mellifera</i>) implicated in elevated colony mortality rates worldwide and facilitated through vector transmission by the ectoparasitic mite <i>Varroa destructor</i>. Clinical, symptomatic DWV infections are almost exclusively associated with high virus titres during pupal development, usually acquired through feeding by <i>Varroa</i> mites when reproducing on bee pupae. Control of the mite population, generally through acaricide treatment, is essential for breaking the DWV epidemic and minimizing colony losses. In this study, we evaluated the effectiveness of remedial mite control on clearing DWV from a colony. DWV titres in adult bees and pupae were monitored at 2 week intervals through summer and autumn in acaricide-treated and untreated colonies. The DWV titres in Apistan treated colonies was reduced 1000-fold relative to untreated colonies, which coincided with both the removal of mites and also a turnover of the bee population in the colony. This adult bee population turnover is probably more critical than previously realized for effective clearing of DWV infections. After this initial reduction, subclinical DWV titres persisted and even increased again gradually during autumn, demonstrating that alternative non-<i>Varroa</i> transmission routes can maintain the DWV titres at significant subclinical levels even after mite removal. The implications of these results for practical recommendations to mitigate deleterious subclinical DWV infections and improving honeybee health management are discussed.</p></div

Results of the maximum likelihood repeated measures model analysis.

<p>Results of the maximum likelihood repeated measures model analysis.</p

Average DWV titres in adult bees.

<p>The average DWV titres with standard error bars in adult bees in Apistan treated colonies (light bars) and untreated control colonies (dark bars) are presented on the right y-axis. The symptomatic bee with deformed wings and the asymptomatic bee represent the difference in titre amounts between clinical and sub-clinical DWV on the right y-axis. The average mite infestation rates with standard error bars in treated colonies (light red lines) and untreated colonies (dark red lines) are presented on the left y-axis. The sampling points are presented progressively in weeks (and months) in relation to when the Apistan treatment was administered.</p

Average DWV titres in pupae.

<p>The average DWV titres with standard error bars in pupae in Apistan treated colonies (light bars) and untreated control colonies (dark bars) are presented on the right y-axis. The symptomatic bee with deformed wings and the asymptomatic bee represent the difference in titre amounts between clinical and sub-clinical DWV on the right y-axis. The average mite infestation rates with standard error bars in treated colonies (light red lines) and untreated colonies (dark red lines) are presented on the left y-axis. The sampling points are presented progressively in weeks (and months) in relation to when the Apistan treatment was administered.</p