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Endemic Q fever in small ruminants remains an ongoing challenge for veterinary and human public health agencies. Though surveillance programs are implemented in Belgium, infection patterns and vaccination profiles, driving variables, as well as geographical clustering were not presented until now. Based on data from a decade of bulk tank milk analysis between 2009 and 2019, shedding in dairy goat herds declined from 16% (8/50) to 6% (10/162), whereas seroprevalence remained between 32% and 40%. Merely up to two shedding dairy sheep flocks were detected until 2019; seroprevalence peaked in 2017 (43%, 12/28) and declined thereafter. The number of animals in the holding influenced significantly (p = .048) the likelihood of shedding, whereas other established risk factors such as uncovered manure, high abortion rates and diversified farm structure could not be confirmed to significantly affect infection on Belgian herd level. Intermittent, incomplete and unsynchronized vaccinated herds shed Coxiella burnetii significantly more often and longer (p < .001) than continuously, complete and synchronized vaccinated herds. Spatial analyses revealed restricted but matching, homogenous clusters with ≤35 km diameter, concentrated in the coastal region close to the border to the Netherlands from 2009 to 2012, and broadened, heterogeneous clusters with ≥45 km diameter between 2014 and 2016 spreading south-west. Though the majority of human cases was notified in this region, the animal clusters could not be allied with Q fever cases. The impact of environmental factors as well as the role of wildlife, rodents and ticks on the transmission between flocks and to humans remains to be elucidated to harness additional epidemiological drivers of Q fever in Belgium. In conclusion, attempts to reduce the burden of Q fever in Belgium should particularly focus on the timely, complete and synchronized vaccination of flocks, including the breeding sire, and particularity in high-risk areas., (© 2021 Wiley-VCH GmbH.)

Q fever is a zoonotic disease caused by the bacteria Coxiella burnetii. Domestic ruminants are the primary source for human infection, and the identification of likely contamination routes from the reservoir animals the critical point to implement control programs. This study shows that Q fever is detected in Belgium in abortion of cattle, goat and sheep at a different degree of apparent prevalence (1.93%, 9.19%, and 5.50%, respectively). In addition, and for the first time, it is detected in abortion of alpaca ( Vicugna pacos ), raising questions on the role of these animals as reservoirs. To determine the relationship between animal and human strains, Multiple Locus Variable-number Tandem Repeat Analysis (MLVA) (n=146), Single-Nucleotide Polymorphism (SNP) (n=92) and Whole Genome Sequencing (WGS) (n=4) methods were used to characterize samples/strains during 2009-2019. Three MLVA clusters (A, B, C) subdivided in 23 subclusters (A1-A12, B1-B8, C1-C3) and 3 SNP types (SNP1, SNP2, SNP6) were identified. The SNP2 type/MLVA cluster A was the most abundant and dispersed genotype over the entire territory, but it seemed not responsible for human cases, as it was only present in animal samples. The SNP1/MLVA B and SNP6/MLVA C clusters were mostly found in small ruminant and human samples, with the rare possibility of spillovers in cattle. SNP1/MLVA B cluster was present in all Belgian areas, while the SNP6/MLVA C cluster appeared more concentrated in the Western provinces. A broad analysis of European MLVA profiles confirmed the host-species distribution described for Belgian samples. In silico genotyping (WGS) further identified the spacer types and the genomic groups of C. burnetii Belgian strains: cattle and goat SNP2/MLVA A isolates belonged to ST61 and genomic group III, while the goat SNP1/MLVA B strain was classified as ST33 and genomic group II. In conclusion, Q fever is widespread in all Belgian domestic ruminants and in alpaca. We determined that the public health risk in Belgium is likely linked to specific genomic groups (SNP1/MLVA B and SNP6/MLVA C) mostly found in small ruminant strains. Considering the concordance between Belgian and European results, these considerations could be extended to other European countries., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Tomaiuolo, Boarbi, Fancello, Michel, Desqueper, Grégoire, Callens, Fretin, Devriendt, Cox and Mori.)

Coxiella burnetii is a globally distributed zoonotic γ-proteobacterium with an obligatory intracellular lifestyle. It is the causative agent of Q fever in humans and of coxiellosis among ruminants, although the agent is also detected in ticks, birds, and various other mammalian species. Requirements for intracellular multiplication together with the necessity for biosafety level 3 facilities restrict the cultivation of C. burnetii to specialized laboratories. Development of a novel medium formulation enabling axenic growth of C. burnetii has facilitated fundamental genetic studies. This review provides critical insights into direct diagnostic methods currently available for C. burnetii. It encompasses molecular detection methods, isolation, and propagation of the bacteria and its genetic characterization. Differentiation of C. burnetii from Coxiella-like organisms is an essential diagnostic prerequisite, particularly when handling and analyzing ticks.

Q fever, a worldwide zoonosis, is an arousing public health concern in many countries since the recent Dutch outbreak. An emerging C. burnetii clone, genotype CbNL01, was identified as responsible for the Dutch human Q fever cluster cases. Since 2009, Q fever surveillance in the goat industry was implemented by the Belgian authorities. The herd prevalence (December 2009-March 2013) ranged between 6.3 and 12.1%. Genotypic analysis highlighted the molecular diversity of the Belgian strains from goats and identified an emerging CbNL01-like genotype. This follow-up allowed the description of shedding profiles in positive farms which was either continuous (type I) and associated to the CbNL01-like genotype; or intermittent (type II) and linked to other genotypes. Despite the circulation of a CbNL01-like strain, the number of notified Belgian human cases was very low. The mandatory vaccination (in June 2011) on positive dairy goat farms in Belgium, contributed to a decrease in shedding., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Coxevac® is the EMA-approved veterinary vaccine for the protection of cattle and goats against Q fever, a zoonotic bacterial disease due to Coxiella burnetii. Since Coxevac® reduces bacterial shedding and clinical symptoms but does not prevent infection, novel, ready-to-use vaccine formulations are needed to increase its immunogenicity. Here, a goat vaccination-challenge model was used to evaluate the impact of the commercially available saponin-based QuilA® adjuvant on Coxevac® immunity. Upon challenge, the QuilA®-Coxevac® group showed a stronger immune response reflected in a higher magnitude of total IgG and an increase in circulating and splenic CD8+ T-cells compared to the Coxevac® and challenged-control groups. The QuilA®-Coxevac® group was characterized by a targeted Th1-type response (IFNγ, IP10) associated with increased transcripts of CD8+ and NK cells in spleens and γδ T cells in bronchial lymph nodes. Coxevac® vaccinated animals presented an intermediate expression of Th1-related genes, while the challenged-control group showed an immune response characterized by pro-inflammatory (IL1β, TNFα, IL12), Th2 (IL4 and IL13), Th17 (IL17A) and other immunoregulatory cytokines (IL6, IL10). An intriguing role was observed for γδ T cells, which were of TBX21- and SOX4-types in the QuilA®-Coxevac® and challenged control group, respectively. Overall, the addition of QuilA® resulted in a sustained Th1-type activation associated with an increased vaccine-induced bacterial clearance of 33.3% as compared to Coxevac® only. QuilA® could be proposed as a readily-applied veterinary solution to improve Coxevac® efficacy against C. burnetii infection in field settings. [ABSTRACT FROM AUTHOR]

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Ruminants are considered the commonest animal reservoir for human infection of Coxiella burnetii, the Q fever causative agent. Considering the recently described importance of human Q fever in Greece, we aimed at providing the first comprehensive direct evidence of C. burnetii in dairy cows in Greece, including the genetic characterization of strains. The 462 examined dairy farms represented all geographical areas of Greece. One bulk tank milk sample was collected from every farm and tested for the presence of C. burnetii. Molecular genotyping of strains, performed directly on samples, revealed the existence of two separate clades characterized by single nucleotide polymorphism (SNP) genotypes of type 1 and type 2. The two clades were clearly distinguished in multiple locus variable-number tandem repeat analysis (MLVA) by two discriminative loci: MS30 and MS28. Whereas MLVA profiles of SNP-type 2 clade were closely related to strains described in other European cattle populations, the MLVA profile observed within the SNP type 1 clade highlighted a peculiar genetic signature for Greece, related to genotypes found in sheep and goats in Europe. The shedding of C. burnetii bearing this genotype might have yet undefined human epidemiological consequences. Surveillance of the genetic distribution of C. burnetii from different sources is needed to fully understand the epidemiology of Q fever in Greece. [ABSTRACT FROM AUTHOR]