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Q fever epidemiological investigations of the likely sources of contamination may involve Coxiella burnetii MLVA for direct and rapid typing from clinical samples. However, little information is available with regards to PCR amplification failures in C. burnetii MLVA typing. This paper focuses on difficulties encountered with MLVA loci that may impact the interpretation of MLVA data and shows that some loci may constitute hotspots for mutational events. MLVA genotyping, using 17 different loci, was used on vaginal swabs (VS) from clinically infected animals as described elsewhere (Chmielewski et al., 2009). Amplicons of interest were sequenced and identified using the BLAST software by comparison with sequences available in GenBank. All VS samples produced MLVA patterns. However, amplification failures or unexpected sizes amplicons (>to 1.5 kbp), making the interpretation of MLVA complicated, were also observed. Sequencing of these amplicons revealed the presence of IS 1111 element insertion. In this C. burnetii MLVA study some difficulties encountered with genotyping are highlighted and the role of IS 1111 element in genome plasticity is confirmed. Finally, the need for the selection of a set of VNTRs for an efficient MLVA scheme and the question of standardization and harmonization for comparable MLVA typing data are raised again. [ABSTRACT FROM AUTHOR]

In alpine pasture, interspecies transmission has recently been incriminated in the epidemiology of pestivirus infection. The aim of this study was to investigate pestivirus infections in wild and domestic ruminants sharing pastures in the French Southern Alps. Animal sera were screened for pestivirus antibodies against the pestivirus NS3 protein by a commercial blocking enzyme linked immunosorbent assay (ELISA). All 38 domestic herds tested were positive for pestivirus-specific antibodies. Individual sero-prevalence reached 76.5% (95% confidence interval [95% CI]: [74.2–78.8%]) of the 1383 sheep tested. For wild ruminants, 38.7% (95% CI: [33.8–43.9%]) of the 369 chamois tested, 28.7% (95% CI: [17.4–38.1%]) of the 72 roe deer, and 22.2% (95% CI: [6.5–37.9%]) of the 27 mouflons were seropositive. Virus screening was carried out on spleen samples from hunted wild animals ( n = 160) and from 15 domestic ruminants (clinically suspected to be persistently infected animals), by a conventional reverse transcription-polymerase chain reaction (RT-PCR). Three pestivirus strains were isolated from the sheep samples positive by RT-PCR. The viruses were classified in the BDV-3, BDV-Tunisian and BDV-6 genotypes. For the first time, one strain (RUPI-05 strain) was isolated from an alpine chamois and clustered in the BDV-6 genotype, showing in the 5′-UTR region 92% of identity with the ovine isolate from the same area. Thus, an active circulation of pestiviruses was demonstrated in both wild and domestic ungulates from the French Southern Alps. The results suggest that interspecies transmission between sheep and chamois probably occur. [ABSTRACT FROM AUTHOR]

Nosemosis is a microsporidian disease causing mortality and weakening of honey bee colonies, especially in the event of co-exposure to other sources of stress. As a result, the disease is regulated in some countries. Reliable and harmonised diagnosis is crucial to ensure the quality of surveillance and research results. For this reason, the first European Interlaboratory Comparison (ILC) was organised in 2017 in order to assess both the methods and the results obtained by National Reference Laboratories (NRLs) in counting Nosema spp. spores by microscopy. Implementing their own routine conditions of analysis, the 23 participants were asked to perform an assay on a panel of ten positive and negative samples of crushed honey bee abdomens. They were asked to report results from a qualitative and quantitative standpoint. The assessment covered specificity, sensitivity, trueness and precision. Quantitative results were analysed in compliance with international standards NF ISO 13528 (2015) and NF ISO 5725-2 (1994). Three results showed a lack of precision and five a lack of trueness. However, overall results indicated a global specificity of 98% and a global sensitivity of 100%, thus demonstrating the advanced performance of the microscopic methods applied to Nosema spores by the NRLs. Therefore, the study concluded that using microscopy to detect and quantify spores of Nosema spp. was reliable and valid. • The ILC objective was to test the accuracy of the methods for counting Nosema spores. • EU-NRLs methods were based on the same principle set out in OIE Terrestrial manual. • The results of the 23 participants showed a good specificity, sensitivity, trueness and precision. [ABSTRACT FROM AUTHOR]

Aethina tumida (small hive beetle, SHB) was first detected in September 2014 in Calabria region, southern Italy, and in a single apiary in Sicily in November 2014. In September 2015, SHB was again recorded in Calabria, and in 2016, only sentinel honey bee nucleus colonies were found to be infested. Its phylogenetic relationship and possible origin were investigated comparing the cox1 sequences with the corresponding region available in the GenBank database. The neighbour-joining method revealed that the first Italian specimen belonged to a group also containing an African specimen from Cameroon. The Italian specimens differ from the SHBs spread worldwide and are split into two different groups: group B1 includes the AfricCam3 sequence and the first SHB identified in Calabria; group B2 includes specimens from Calabria and the only one from Sicily which share identical cox1 sequences. SHB in Italy appears to have been introduced from Africa and includes independent or contemporary incursions in the two concerned regions. The most likely scenario is that SHB was introduced into Calabria followed by man-mediated migration to Sicily. [ABSTRACT FROM AUTHOR]

Abstract: Coxiella burnetii is an intracellular bacterium that causes a worldwide zoonosis, the Q fever. Currently, to diagnose the infection in ruminants, whole cell antigens-based ELISAs are used. In this study a heat shock protein, the HspB, was evaluated for its ability to be recognized by the goat immune system and its capacity to sign a stage of infection. The htpB gene of C. burnetii was cloned and sequenced. A high identity (>90%) was observed among the htpB genes of four ruminant strains tested. A recombinant protein was expressed in a prokaryotic expression system. The rHspB protein was used to determine the IgG reactivity by ELISA. Sera from experimentally and naturally infected goats were tested. The rHspB is recognized early during the infection course, at 18 days post-infection. Moreover, 80–90% of the animals tested were positive at 39–60dpi. In addition, animals presenting a reactivation of the infection displayed a higher reactivity, statistically significant (p <0.05), than that of the animals in latent infection. These findings suggest that the rHspB could be a good candidate for the development of an ELISA test making possible the detection of recent C. burnetii infection in goats as well as reactivation in those with latent infection. [Copyright &y& Elsevier]

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. [ABSTRACT FROM AUTHOR]