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Surveillance of Coxiella burnetii Shedding in Three Naturally Infected Dairy Goat Herds after Vaccination, Focusing on Bulk Tank Milk and Dust Swabs

Q fever outbreaks on three dairy goat farms (A–C) were monitored after the animals had been vaccinated with an inactivated Coxiella burnetii phase I vaccine. The antibody response was measured before vaccination by serum samples with two C. burnetii phase-specific ELISAs to characterize the disease status. Shedding was determined by vaginal swabs during three kidding seasons and monthly bulk tank milk (BTM) samples. Dust swabs from one windowsill of each barn and from the milking parlors were collected monthly to evaluate the indoor exposure.

These samples were analyzed by qPCR. The phase-specific serology revealed an acute Q fever infection in herd A, whereas herds B and C had an ongoing and past infection, respectively. In all three herds, vaginal shedders were present during three kidding seasons. In total, 50%, 69%, and 15% of all collected BTM samples were C. burnetii positive in herds A, B, and C, respectively. Barn dust contained C. burnetii DNA in 71%, 45%, and 50% of examined swabs collected from farms A, B, and C, respectively. The largest number of C. burnetii positive samples was obtained from the milking parlor (A: 91%, B: 72%, C: 73%), indicating a high risk for humans to acquire Q fever during milking activity.

Abstract taken from: Benjamin U Bauer, Clara Schoneberg, T Louise Herms, Martin Runge, Martin Ganter, Surveillance of Coxiella burnetii Shedding in Three Naturally Infected Dairy Goat Herds after Vaccination, Focusing on Bulk Tank Milk and Dust Swabs, Vet Sci. 2022 Feb 24;9(3):102. doi: 10.3390/vetsci9030102.

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Q fever expertise among human and veterinary health professionals in Germany – A stakeholder analysis of knowledge gaps

Q fever is a zoonosis caused by Coxiella burnetii. In Germany, the common sources of human infections include small ruminants that excrete the pathogen. Q fever in humans can be asymptomatic or nonspecific. However, severe disease progression is also possible, which can lead to death. Q fever in small ruminants is usually asymptomatic, although reproductive disorders may occur. To protect humans from Q fever, it is important that human and veterinary health professionals (practitioners/health authority employees) have comprehensive knowledge of the diagnosis, control and prevention of Q fever, and its zoonotic potential. To ensure and enhance this understanding, this stakeholder analysis assessed Q fever expertise in human and veterinary health professionals in Germany and investigated how these knowledge gaps can best be resolved.

For this purpose, an online survey and two focus group discussions were conducted with 836 and 18 participants, respectively. Knowledge gaps are due to a lack of awareness of Q fever, especially among human health practitioners. Moreover, colleagues who have heard about Q fever still lack the necessary cross-species knowledge to successfully diagnose, control and prevent this zoonosis. Additionally, differences exist between stakeholders regarding their work context and the region in which they work. In this study, stakeholders in southwestern Germany had slightly better Q fever knowledge than their colleagues in North-eastern Germany.

In addition, information sources aimed at resolving knowledge gaps involve direct conversations between the stakeholders, as well as reading materials and seminars. Each of these information sources should focus on interdisciplinary resources to strengthen the cooperation between human and veterinary health professionals and to raise awareness of the strengths of each stakeholder group. These results have already been implemented by the Q-GAPS project, with goals of raising awareness of Q fever and filling knowledge gaps.

Abstract taken from: Fenja Winter und Amely Campe, A stakeholder analysis of knowledge gaps, PLoS One, 2022 Mar 3;17(3):e0264629.

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The Coxiella burnetii T4SS effector protein AnkG hijacks the 7SK small nuclear ribonucleoprotein complex for reprogramming host cell transcription

Inhibition of host cell apoptosis is crucial for survival and replication of several intracellular bacterial pathogens. To interfere with apoptotic pathways, some pathogens use specialized secretion systems to inject bacterial effector proteins into the host cell cytosol. One of these pathogens is the obligate intracellular bacterium Coxiella burnetii, the etiological agent of the zoonotic disease Q fever.

In this study, we analyzed the molecular activity of the anti-apoptotic T4SS effector protein AnkG (CBU0781) to understand how C. burnetii manipulates host cell viability. We demonstrate by co- and RNA-immunoprecipitation that AnkG binds to the host cell DExD box RNA helicase 21 (DDX21) as well as to the host cell 7SK small nuclear ribonucleoprotein (7SK snRNP) complex, an important regulator of the positive transcription elongation factor b (P-TEFb). The co-immunoprecipitation of AnkG with DDX21 is probably mediated by salt bridges and is independent of AnkG-7SK snRNP binding, and vice versa. It is known that DDX21 facilitates the release of P-TEFb from the 7SK snRNP complex.

Consistent with the documented function of released P-TEFb in RNA Pol II pause release, RNA sequencing experiments confirmed AnkG-mediated transcriptional reprogramming and showed that expression of genes involved in apoptosis, trafficking, and transcription are influenced by AnkG. Importantly, DDX21 and P-TEFb are both essential for AnkG-mediated inhibition of host cell apoptosis, emphasizing the significance of the interaction of AnkG with both, the DDX21 protein and the 7SK RNA. In line with a critical function of AnkG in pathogenesis, the AnkG deletion C. burnetii strain was severely affected in its ability to inhibit host cell apoptosis and to generate a replicative C. burnetii-containing vacuole. In conclusion, the interference with the activity of regulatory host cell RNAs mediated by a bacterial effector protein represent a novel mechanism through which C. burnetii modulates host cell transcription, thereby enhancing permissiveness to bacterial infection.

Source: PLoS Pathogens 18 (2022)

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Inactivation kinetics of Coxiella burnetii during high-temperature short-time pasteurization of milk:

 The Gram-negative, obligate intracellular bacterium Coxiella burnetii is the causative organism of the zoonosis Q fever and is known for its resistance toward various intra and extracellular stressors. Infected ruminants such as cattle, sheep, and goats can shed the pathogen in their milk. Pasteurization of raw milk was introduced for the inactivation of C. burnetii and other milk-borne pathogens. Legal regulations for the pasteurization of milk are mostly based on recommendations of the Codex Alimentarius. As described there, C. burnetii is considered as the most heat-resistant non-sporeforming bacterial pathogen in milk and has to be reduced by at least 5 log10-steps during the pasteurization process. However, the corresponding inactivation data for C. burnetii originate from experiments performed more than 60 years ago. Recent scientific findings and the technological progress of modern pasteurization equipment indicate that C. burnetii is potentially more effectively inactivated during pasteurization than demanded in the Codex Alimentarius. In the present study, ultra-high heat-treated milk was inoculated with different C. burnetii field isolates and subsequently heat-treated in a pilot-plant pasteurizer. Kinetic inactivation data in terms of D- and z-values were determined and used for the calculation of heat-dependent log reduction. With regard to the mandatory 5 log10-step reduction of the pathogen, the efficacy of the established heat treatment regime was confirmed, and, in addition, a reduction of the pasteurization temperature seems feasible.

Source: Marcel Wittwer, Philipp Hammer, Martin Runge, Peter Valentin-Weigand, Heinrich Neubauer, Klaus Henning und Katja Mertens-Scholz, Inactivation kinetics of Coxiella burnetii during high-temperature short-time pasteurization of milk
Frontiers in Microbiology, Vol. 12, 2022

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Cover story: A Q fever outbreak on a mixed dairy farm – a one health approach

 A Q fever outbreak on a dairy goat/cattle farm was investigated with regard to the One Health concept. Samples from all domestic animals and the farmer's family were collected, including goats with different reproductive statuses. The serological analysis revealed current infections in goats, whereas cows had ongoing infections. The possible influence of gestation and hormonal status on Coxiella burnetii infection in goats is discussed. A new cattle-associated Coxiella burnetii genotype C16 was identified in vaginal swabs and placentas from ruminants. Feline, canine, and human sera also tested positive for Coxiella burnetii antibodies. We conclude that the cattle herd may have transmitted Coxiella to the pregnant goats, resulting in zoonotic spread of Q fever to cats, dogs, and humans on the affected farm.

Source: Benjamin U. Bauer, Michael R. Knittler, T. Louise Herms, Dimitrios Frangoulidis, Svea Matthiesen, Dennis Tappe, Martin Runge and Martin Ganter
Cover story: A Q Fever Outbreak in a Mixed Dairy Farm - A one Health Approach
Publikation: Multispecies Q Fever Outbreak in a Mixed Dairy Goat and Cattle Farm Based on a New Bovine-Associated Genotype of Coxiella burnetii
Vet. Sci., Volume 8, Issue 11 (November 2021)

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SchafeQ fever outbreak at Charité in Berlin:

 According to a news story published by "Berliner Morgenpost" staff members were affected by a q fever outbreak at Charité in Mid-July 2021 after animal experiments with pregnant sheep. 20 staff members of the „Forschungseinrichtung für experimentelle Medizin" at the Virchow Clinique were infected with Coxiella burnetii when carrying out the animal experiments.

The trigger for the q fever specific diagnostics among staff members was obviously the detection of the pathogen in one of the experimental animals at the beginning of August 2021. At that time, it was possible to detect antibodies in staff members of the Charité, which indicated an acute q fever infection. Upon request by "Berliner Morgenpost" the Charité pointed out that all individuals who became ill have recovered in the meantime.

In addition, a Charité spokesman stated that the experimental animals came from breeding facilities in Brandenburg, where no q fever has been detected so far. Clinical signs have been absent in the sheep and there hasn't been any suspicion that the animals had been infected in the first place. Immediately after a first suspected infection within the Charité the holding of the research facilities was quarantined. Yet another consequence from the q fever outbreak is that stricter quality criteria regarding hygiene in the animal stock, supply chain and risk assessment will be paid attention to in future when purchasing animals, in order to reduce the risk of introducing zoonotic diseases. There hasn't been any obligation to check for the agent of q fever in farm animals regularly, so far.

Remarks:
Fortunately, institutions from the Q-GAPS research project (National Reference Laboratories for q fever, human and veterinary medicine, Lower State Office for Consumer Protection and Food Safety, Food and Veterinary Institute Braunschweig/Hannover, (LAVES)) also contributed to the analysis of the q fever outbreak described above. Based on the experience and knowledge of the Q-GAPS partners a guideline is being drawn up which contains information including recommendations for action concerning the overall topic q fever. The article about the outbreak in an establishment for animal use cited here shows an example of the necessity and importance of such a professional gathering of information and consulting. Therefore, it should be taken into account as to whether further protective measurements have to be taken or procedures for risk mitigation have to be considered, if necessary, not only in holdings of origin but also in research projects along these lines using experimental animals. For example, a regular q fever monitoring in as well as in experimental animals would be advisable, especially, as q fever infections can often be asymptomatic.

Image Sheep: © Stiftung Tierärztliche Hochschule Hannover

Sources:
Berliner Morgenpost: Tierseuchen-Ausbruch an der Charité (in German, for a fee)

BZ: Q-Fieber! 20 Charité-Mitarbeiter mit Tier-Seuche infiziert

High Prevalence and New Genotype of Coxiella burnetii in Ticks Infesting Camels in Somalia:

 Coxiella burnetii is the causative agent of Q fever. It can infect animals, humans, and birds, as well as ticks, and it has a worldwide geographical distribution. To better understand the epidemiology of C. burnetii in Somalia, ticks infesting camels were collected from five different regions, including Bari, Nugaal, Mudug, Sool, and Sanaag, between January and March 2018.

Collected ticks were tested for C. burnetii and Coxiella-like endosymbiont DNA by using IS1111, icd, and Com1-target PCR assays. Moreover, sequencing of the 16S-rRNA was conducted. Molecular characterization and typing were done by adaA-gene analysis and plasmid-type identification. Further typing was carried out by 14-marker Multi-Locus Variable-Number Tandem Repeats (MLVA/VNTR) analysis. The investigated ticks (n = 237) were identified as Hyalomma spp. (n = 227, 95.8%), Amblyomma spp. (n = 8, 3.4%), and Ripicephalus spp. (n = 2, 0.8%), and 59.1% (140/237) of them were positive for Coxiella spp. While Sanger sequencing and plasmid-type identification revealed a C. burnetii that harbours the QpRS-plasmid, MLVA/VNTR genotyping showed a new genotype which was initially named D21. In conclusion, this is the first report of C. burnetii in ticks in Somalia. The findings denote the possibility that C. burnetii is endemic in Somalia. Further epidemiological studies investigating samples from humans, animals, and ticks within the context of “One Health” are warranted.

 Abstract taken from Dimitrios Frangoulidis; Claudia Kahlhofer; Ahmed Shire Said; Abdinasir Yusuf Osman; Lidia Chitimia-Dobler; Yassir Adam Shuaib
Pathogens 2021, Volume 10, Issue 6, 741

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A Straightforward Hypoxic Cell Culture Method Suitable for Standard Incubators:

 We present a new and straightforward method by which standard cell culture plates can be sealed off from ambient air and be placed under controlled hypoxic cell culture conditions without costly or highly specialized materials. The method was established on a murine cell culture system using the dendritic cell line JAWS II but can be readily adapted to other cell cultures. The procedure was designed to be easy to implement in cell culture laboratories with standard incubators and requires only readily available materials, resources, and consumables, such as six-well plates, degassed culture medium, CoCl2, a vacuum sealer, etc., and no further complicated laboratory equipment. The simple hypoxic cell culture method presented here is technically reliable and experimentally safe. As it can be performed in any standard incubator, it is suitable for use at both low and higher biosafety levels.

 Straightforward Hypoxic Cell Culture Method

Abstract taken from Svea Matthiesen, Rico Jahnke, Michael R. Knittler, Methods Protoc. 2021, 4(2), 25; https://doi.org/10.3390/mps4020025

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The Prevalence of Coxiella burnetii in Hard Ticks in Europe and Their Role in Q Fever Transmission Revisited - A Systematic Review

The zoonosis Q fever is caused by the obligate intracellular bacterium Coxiella burnetii. Besides the main transmission route via inhalation of contaminated aerosols, ticks are discussed as vectors since the first isolation of the pathogen from a Dermacentor andersonii tick. The rare detection of C. burnetii in ticks and the difficult differentiation of C. burnetii from Coxiella-like endosymbionts (CLEs) are questioning the relevance of ticks in the epidemiology of Q fever.

In this review, literature databases were systematically searched for recent prevalence studies concerning C. burnetii in ticks in Europe and experimental studies evaluating the vector competence of tick species. A total of 72 prevalence studies were included and evaluated regarding DNA detection methods and collection methods, country, and tested tick species. Specimens of more than 25 different tick species were collected in 23 European countries. Overall, an average prevalence of 4.8% was determined. However, in half of the studies, no Coxiella-DNA was detected. In Southern European countries, a significantly higher prevalence was observed, possibly related to the abundance of different tick species here, namely Hyalomma spp. and Rhipicephalus spp. In comparison, a similar proportion of studies used ticks sampled by flagging and dragging or tick collection from animals, under 30% of the total tick samples derived from the latter. There was no significant difference in the various target genes used for the molecular test. In most of the studies, no distinction was made between C. burnetii and CLEs. The application of specific detection methods and the confirmation of positive results are crucial to determine the role of ticks in Q fever transmission. Only two studies were available, which assessed the vector competence of ticks for C. burnetii in the last 20 years, demonstrating the need for further research.

Abstract taken from: Sophia Körner, Gustavo R Makert, Sebastian Ulbert, Martin Pfeffer, Katja Mertens-Scholz, The Prevalence of Coxiella burnetii in Hard Ticks in Europe and Their Role in Q Fever Transmission Revisited - A Systematic Review, Front Vet Sci. 2021 Apr 26; 8: 655715

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Humoral immune response to Q fever vaccination of three sheep flocks naturally pre-infected with Coxiella burnetii:

 Q fever is a zoonotic disease caused by the bacterium Coxiella burnetii; Coxiella-infected ruminants are the main reservoir shedding the pathogen during abortion or parturition through birth products. Germany has a long history of small-scale Q fever epidemics in the human population mostly associated with lambing sheep. Therefore, fast and efficient control measures are essentially required to prevent transmission from infected sheep flocks to humans.

In our present study, three sheep flocks were vaccinated with an inactivated C. burnetii phase I vaccine after a field infection with C. burnetii was diagnosed. Serum samples and vaginal swabs were collected at different time points to evaluate the extent of the outbreak and the consequences of the vaccination. The serum samples were examined by phase-specific IgG phase I and phase II ELISAs and a commercial ELISA, simultaneously detecting both phase variations. Moreover, vaginal swabs were analysed by qPCR. The fourth flock with no Q fever history and non-vaccinated animals were used as a control group to evaluate the phase-specific ELISAs.

The inactivated C. burnetii phase I vaccine induced an IgG phase II response and boosted the humoral immune reaction against natural preinfections. Furthermore, the longevity of vaccine-induced antibodies seems to depend on previous infections. Around 16 months after primary vaccination, mainly IgG phase I antibodies were detectable. Vaccination did not prevent shedding at the next lambing season. Most interestingly, the phasespecific ELISAs revealed more C. burnetii positive animals than the blended ELISA-Assay. Taken together, phase-specific ELISAs are suitable tools to provide insights into natural- or vaccine-induced humoral immune responses to C. burnetii in sheep.

Original Abstract, © 2021 Elsevier Ltd. All rights reserved.

Source: Benjamin U. Bauer, Michael R. Knittler, T. Louise Prüfer, Annika Wolf, Svea Matthiesen, Martin Runge, Martin Ganter (2021) Humoral immune response to Q fever vaccination of three sheep flocks naturally pre-infected with Coxiella burnetii. Vaccine, Volume 39, Issue 10, 5 March 2021, Pages 1499-1507, https://doi.org/10.1016/j.vaccine.2021.01.062

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