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Coxiella burnetii is a globally distributed obligate intracellular pathogen. Although often asymptomatic, infections can cause acute Q fever with influenza-like symptoms and/or severe chronic Q fever. Coxiella burnetii develops a unique replicative niche within host cells called the Coxiella-containing vacuole (CCV), facilitated by the Dot/Icm type IV secretion system translocating a cohort of bacterial effector proteins into the host. The role of some effectors has been elucidated; however, the actions of the majority remain enigmatic and the list of true effectors is disputable. This study examined CBU2016, a unique C. burnetii protein previously designated as an effector with a role in infection. We were unable to validate CBU2016 as a translocated effector protein. Employing targeted knock-out and complemented strains, we found that the loss of CBU2016 did not cause a replication defect within Hela, THP-1, J774, or iBMDM cells or in axenic media, nor did it affect the pathogenicity of C. burnetii in the Galleria mellonella infection model. The absence of CBU2016 did, however, result in a consistent decrease in the size of CCVs in HeLa cells. These results suggest that although CBU2016 may not be a Dot/Icm effector, it is still able to influence the host environment during infection., (© The Author(s) 2024. Published by Oxford University Press on behalf of FEMS.)

Coxiella burnetii is an obligate intracellular bacterium that causes Q fever, a zoonotic disease typically manifests as a severe flu-illness. After invading into the host cells, C. burnetii delivers effectors to regulate the vesicle trafficking and fusion events to form a large and mature Coxiella -containing vacuole (CCV), providing sufficient space and nutrition for its intracellular growth and proliferation. Lysosomal trafficking regulator (LYST) is a member of the Beige and Chediak-Higashi syndrome (BEACH) family, which regulates the transport of vesicles to lysosomes and regulates TLR signaling pathway, but the effect of LYST on C. burnetii infection is unclear. In this study, a series of experiments has been conducted to investigate the influence of LYST on intracellular growth of C. burnetii . Our results showed that lyst transcription was up-regulated in the host cells after C. burnetii infection, but there is no significant change in lyst expression level after infection with the Dot/Icm type IV secretion system (T4SS) mutant strain, while CCVs expansion and significantly increasing load of C. burnetii appeared in the host cells with a silenced lyst gene, suggesting LYST inhibits the intracellular proliferation of C. burnetii by reducing CCVs size. Then, the size of CCVs and the load of C. burnetii in the HeLa cells pretreated with E-64d were significantly decreased. In addition, the level of iNOS was decreased significantly in LYST knockout THP-1 cells, which was conducive to the intracellular replication of C. burnetii . This data is consistent with the phenotype of L-NMMA-treated THP-1 cells infected with C. burnetii . Our results revealed that the upregulation of lyst transcription after infection is due to effector secretion of C. burnetii and LYST inhibit the intracellular replication of C. burnetii by reducing the size of CCVs and inducing nos2 expression., 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 © 2024 Wan, Zhang, Zhao, OuYang, Yu, Xiong, Zhao and Jiao.)

Having previously shown that soluble E-cadherin (sE-cad) is found in sera of Q fever patients and that infection of BeWo cells by C. burnetii leads to modulation of the E-cad/β-cat pathway, our purpose was to identify which sheddase(s) might catalyze the cleavage of E-cad. Here, we searched for a direct mechanism of cleavage initiated by the bacterium itself, assuming the possible synthesis of a sheddase encoded in the genome of C. burnetii or an indirect mechanism based on the activation of a human sheddase. Using a straightforward bioinformatics approach to scan the complete genomes of four laboratory strains of C. burnetii , we demonstrate that C. burnetii encodes a 451 amino acid sheddase (CbHtrA) belonging to the HtrA family that is differently expressed according to the bacterial virulence. An artificial CbHtrA gene (CoxbHtrA) was expressed, and the CoxbHtrA recombinant protein was found to have sheddase activity. We also found evidence that the C. burnetii infection triggers an over-induction of the human HuHtrA gene expression. Finally, we demonstrate that cleavage of E-cad by CoxbHtrA on macrophages-THP-1 cells leads to an M2 polarization of the target cells and the induction of their secretion of IL-10, which "disarms" the target cells and improves C. burnetii replication. Taken together, these results demonstrate that the genome of C . burnetii encodes a functional HtrA sheddase and establishes a link between the HtrA sheddase-induced cleavage of E-cad, the M2 polarization of the target cells and their secretion of IL-10, and the intracellular replication of C. burnetii .

Coxiella burnetii is a bacterial pathogen that replicates within host cells by establishing a membrane-bound niche called the Coxiella -containing vacuole. Biogenesis of this compartment requires effectors of its Dot/Icm type IV secretion system. A large cohort of such effectors has been identified, but the function of most of them remain elusive. Here, by a cell-based functional screening, we identified the effector Cbu0513 (designated as CinF) as an inhibitor of NF-κB signaling. CinF is highly similar to a fructose-1,6-bisphosphate (FBP) aldolase/phosphatase present in diverse bacteria. Further study reveals that unlike its ortholog from Sulfolobus tokodaii , CinF does not exhibit FBP phosphatase activity. Instead, it functions as a protein phosphatase that specifically dephosphorylates and stabilizes IκBα. The IκBα phosphatase activity is essential for the role of CinF in C. burnetii virulence. Our results establish that C. burnetii utilizes a protein adapted from sugar metabolism to subvert host immunity., Competing Interests: The authors declare no competing interest., (Copyright © 2021 the Author(s). Published by PNAS.)

Coxiella burnetii is a bacterial pathogen that replicates in a specialised lysosome-derived organelle called the Coxiella-containing vacuole (CCV). Establishment of the CCV requires the Dot/Icm type IVB secretion system. A previous transposon mutagenesis screen identified the gene cbu1754 as being important for the intracellular replication of C. burnetii. To understand the function of the protein encoded by cbu1754, CCV maturation and intracellular replication phenotypes of a cbu1754 mutant were analysed. In contrast to vacuoles containing wild-type C. burnetii Nine Mile phase II, vacuoles containing the isogenic cbu1754 mutant were smaller and did not display detectible amounts of the autophagy protein LC3, which indicated a CCV biogenesis defect. The Cbu1754 protein was not efficiently delivered into the host cell cytosol during infection, which indicated this protein is not a Dot/Icm-translocated effector protein. Secondary structure predictions suggested that Cbu1754 could be similar to the Legionella pneumophila LvgA protein, which is a component of the Dot/Icm apparatus. Consistent with this hypothesis, production of Cbu1754 in an L. pneumophila ∆lvgA mutant restored LvgA-dependent activities. The L. pneumophila proteins LvgA, IcmS and IcmW are interacting partners that comprise a subassembly of the coupling protein complex that mediates Dot/Icm-dependent effector translocation. Similarly, the Cbu1754 protein was found to be a component of the chaperone complex containing the C. burnetii proteins IcmS and IcmW. Thus, the Cbu1754 protein is an LvgA-related protein important for Dot/Icm function and intracellular replication of C. burnetii., (© 2021 John Wiley & Sons Ltd.)

Background: Coxiella burnetii is the causative agent of Q fever which is a highly infectious zoonotic disease. C. burnetii has become one of the most important causes of abortion in livestock, which can lead to widespread abortions in these animals. There are very limited studies on the prevalence of C. burnetii infection in cases of animal abortion in Iran. The aim of this study was to investigate the occurrence of C. burnetii in ruminant abortion samples in Iran., Methods: Abortion samples from cattle, sheep and goats were collected from different parts of Iran and were tested using Real-time PCR targeting the IS1111 element of C. burnetii., Results: In this study, 36 samples (24.7%) of the 146 collected samples were positive for C. burnetii. The prevalence of C. burnetii was 21.3% (20 of 94 samples) in sheep samples. Also, 10 of 46 cattle samples (21.7%) were positive. All six goat abortion samples were positive for C. burnetii., Conclusions: The findings of the study demonstrate that C. burnetii plays an important role in domestic ruminant abortions in Iran, suggesting that more attention should be paid to the role of C. burnetii in domestic animal abortions by veterinary organizations. The risk of transmitting the infection to humans due to abortion of animals should also be considered., Competing Interests: The authors have declared that no competing interests exist.

The obligate intracellular pathogen Coxiella burnetii is the causative agent of the zoonosis Q fever. C. burnetii infection can have severe outcomes due to the development of chronic infection. To establish and maintain an infection, C. burnetii depends on a functional type IVB secretion system (T4BSS) and, thus, on the translocation of effector proteins into the host cell. Here, we showed that the C. burnetii T4BSS effector protein CaeB targets the conserved endoplasmatic reticulum (ER) stress sensor IRE1 during ER stress in mammalian and plant cells. CaeB-induced upregulation of IRE1 RNase activity was essential for CaeB-mediated inhibition of ER stress-induced cell death. Our data reveal a novel role for CaeB in ER stress signalling modulation and demonstrate that CaeB is involved in pathogenicity in vivo. Furthermore, we provide evidence that C. burnetii infection leads to modulation of the ER stress sensors IRE1 and PERK, but not ATF6 during ER stress. While the upregulation of the RNase activity of IRE1 during ER stress depends on CaeB, modulation of PERK is CaeB independent, suggesting that C. burnetii encodes several factors influencing ER stress during infection., (© 2020 The Authors. Cellular Microbiology published by John Wiley & Sons Ltd.)

The human pulmonary environment is complex, containing a matrix of cells, including fibroblasts, epithelial cells, interstitial macrophages, alveolar macrophages and neutrophils. When confronted with foreign material or invading pathogens, these cells mount a robust response. Nevertheless, many bacterial pathogens with an intracellular lifecycle stage exploit this environment for replication and survival. These include, but are not limited to, Coxiella burnetii, Legionella pneumophila, Yersinia pestis, Mycobacterium tuberculosis and Staphylococcus aureus. Currently, few human disease-relevant model systems exist for studying host-pathogen interactions during these bacterial infections in the lung. Here, we present two novel infection platforms, human alveolar macrophages (hAMs) and human precision-cut lung slices (hPCLS), along with an up-to-date synopsis of research using said models. Additionally, alternative uses for these systems in the absence of pathogen involvement are presented, such as tissue banking and further characterization of the human lung environment. Overall, hAMs and hPCLS allow novel human disease-relevant investigations that other models, such as cell lines and animal models, cannot completely provide., (© The Author(s) 2021. Published by Oxford University Press on behalf of FEMS.)

The gram-negative bacterium Coxiella burnetii is the causative agent of Query (Q) fever in humans and coxiellosis in livestock. Host genetics are associated with C. burnetii pathogenesis both in humans and animals; however, it remains unknown if specific genes are associated with severity of infection. We employed the Drosophila Genetics Reference Panel to perform a genome-wide association study to identify host genetic variants that affect host survival to C. burnetii infection. The genome-wide association study identified 64 unique variants (P < 10-5) associated with 25 candidate genes. We examined the role each candidate gene contributes to host survival during C. burnetii infection using flies carrying a null mutation or RNAi knockdown of each candidate. We validated 15 of the 25 candidate genes using at least one method. This is the first report establishing involvement of many of these genes or their homologs with C. burnetii susceptibility in any system. Among the validated genes, FER and tara play roles in the JAK/STAT, JNK, and decapentaplegic/TGF-β signaling pathways which are components of known innate immune responses to C. burnetii infection. CG42673 and DIP-ε play roles in bacterial infection and synaptic signaling but have no previous association with C. burnetii pathogenesis. Furthermore, since the mammalian ortholog of CG13404 (PLGRKT) is an important regulator of macrophage function, CG13404 could play a role in host susceptibility to C. burnetii through hemocyte regulation. These insights provide a foundation for further investigation regarding the genetics of C. burnetii susceptibility across a wide variety of hosts., (© The Author(s) 2021. Published by Oxford University Press on behalf of Genetics Society of America.)

Purpose: Dermacentor reticulatus is the second most common tick species in Poland after Ixodes ricinus. The aim of the study was to analyze the presence of pathogen DNA in D. reticulatus., Materials and Methods: Ticks were collected in The Protected Landscape Area of the Bug and Nurzec Valley (52°40' N and 22°28' E) between 2016 and 2017. End-point PCR for Borrelia burgdorferi sensu lato, Anaplasma phagocytophilum, Babesia spp., Rickettsia spp., Bartonella spp. and Coxiella burnetii detection was performed., Results: Tick-borne pathogens' DNA was detected in 11.3% of 301 ticks: B. burgdorferi s.l. in 3.6%, Babesia spp. in 6.3%, A. phagocytophilum in 0.7% and B. burgdorferi s.l.-Babesia spp. co-infection in 0.7%. In all 21 Babesia spp. positive samples, sequence analysis confirmed the presence of Babesia canis with an 80.3%-98.3% homology with the B. canis sequences in GenBank. C. burnetii, Bartonella spp., and Rickettsia spp. DNA were not detected., Conclusions: Dermacentor reticulatus from north-eastern Poland were found to carry three of the most common tick-borne pathogens (B. burgdorferi s.l., Babesia canis, A. phagocytophilum) which lead to single and mixed infections. Babesia canis was the most prevalent pathogen identified in D. reticulatus., Competing Interests: Declaration of competing interest The authors declare no conflict of interests., (Copyright © 2021 Medical University of Bialystok. Published by Elsevier B.V. All rights reserved.)

Q fever is an important zoonosis, yet it is often neglected and can present large outbreaks, as observed in the Netherlands. In the past few years, cases of Q fever have been described in Brazil; however, the epidemiological situation of Q fever in ruminants, the main reservoir of the pathogen, is unknown in this country. Our study aimed to estimate the prevalence of C. burnetii in cattle sent to slaughterhouses using an immunofluorescence assay (IFA) and quantitative real-time PCR (qPCR). From 1515 cattle serum samples collected from nine slaughterhouses, 23.8% (360/1515) were serologically positive by IFA (cutoff titer>1:64), indicating past or recent exposure to C. burnetii infection. Among the 54 cities sampled during the study, 83.3% (45/54) had at least one seropositive animal. Subsequently, all seropositive samples were submitted to qPCR for C. burnetii DNA, and 12.2% (44/360) of the sera were qPCR positive, which indicates bacteremia and suggests active or recent infection. The results highlight the risk for abattoir workers that results from exposure to contaminated aerosols produced during slaughter procedures. Moreover, the heat maps that were construction from the positive samples demonstrate the widespread distribution of C. burnetii in the State of São Paulo, Brazil and denotes the need for surveillance and preventive measures to reduce the prevalence in cattle., Competing Interests: The authors have declared that no competing interests exist.

The persistent and highly transmissible Coxiella burnetii is a neglected infection that negatively affects reproductive parameters of livestock. It is also of zoonotic importance and has been reported to cause devastating human infections globally. Domestic ruminants represent the most frequent source of human infection. Data from Nigeria are very few and outdated. There is a significant gap in up-to-date information on the exposure, spatial distribution and risk factors of infection of this important disease. The exposure to C. burnetii was determined using sensitive serological assays in cattle and small ruminants. A total of 538 animals made up of 268 cattle and 270 small ruminants were sampled from three northern Nigerian states. The proportion of cattle sampled that were seropositive from the study locations were: Kwara 14/90 (15.6%; 95% CI: 8.8-24.7); Plateau 10/106 (9.43%; 95% CI: 4.6-16.7) and Borno 4/72 (5.56%; 95% CI: 1.5-13.6) states. Lower seroprevalence was recorded among the small ruminants sampled, with positives recorded from sheep and goat sampled from only Kwara state 6/184 (3.3%; 95% CI: 1.2-7.0); while none of the small ruminants sampled from Plateau were seropositive. The results of the bivariate analysis showed that none of the tested independent variables (village, age group, sex, breed of cattle, presence of ticks, reproductive status, and management system) were statistically significant factors associated with seropositivity of cattle for antibodies to C. burnetii. Stakeholders involved in animal husbandry should be duly educated on proper disposal of birth products as well as bodily fluids in order to reduce environmental contamination, persistence and human infection., Competing Interests: The authors have declared that no competing interests exists.

Intracellular bacterial pathogens harbour genes, the closest homologues of which are found in eukaryotes. Regulator of chromosome condensation 1 (RCC1) repeat proteins are phylogenetically widespread and implicated in protein-protein interactions, such as the activation of the small GTPase Ran by its cognate guanine nucleotide exchange factor, RCC1. Legionella pneumophila and Coxiella burnetii, the causative agents of Legionnaires' disease and Q fever, respectively, harbour RCC1 repeat coding genes. Legionella pneumophila secretes the RCC1 repeat 'effector' proteins LegG1, PpgA and PieG into eukaryotic host cells, where they promote the activation of the pleiotropic small GTPase Ran, microtubule stabilisation, pathogen vacuole motility and intracellular bacterial growth as well as host cell migration. The RCC1 repeat effectors localise to the pathogen vacuole or the host plasma membrane and target distinct components of the Ran GTPase cycle, including Ran modulators and the small GTPase itself. Coxiella burnetii translocates the RCC1 repeat effector NopA into host cells, where the protein localises to nucleoli. NopA binds to Ran GTPase and promotes the nuclear accumulation of Ran(GTP), thus pertubing the import of the transcription factor NF-κB and innate immune signalling. Hence, divergent evolution of bacterial RCC1 repeat effectors defines the range of Ran GTPase cycle targets and likely allows fine-tuning of Ran GTPase activation by the pathogens at different cellular sites., (© 2020 John Wiley & Sons Ltd.)

Coxiella burnetii is a highly infectious zoonotic pathogen infecting wide range of mammals, including humans. In the present study, a total of 711 blood samples from bovines [cattle (n = 543) and buffaloes (n = 168)] from eight farms at different geographical locations in India were screened for C. burnetii targeting the IS1111 and the com1 genes. The anti-C. burnetii antibodies in serum samples were detected using indirect-ELISA kits. Also, a total of 21 parameters pertaining to animal health and farm management were identified to assess their role as possible risk factors for coxiellosis among the targeted farms. The apparent prevalence (positive for PCR and/or ELISA) for coxiellosis was reported to be 24.5% in cattle and 8.9% in buffaloes. In cattle, the detection rate of C. burnetii employing the IS1111 gene (8.5%) was found to be significantly higher (p<0.05) as compared to the com1 (6.5%) gene. The seropositivity by ELISA was higher among cattle (17.7%) than in buffaloes (8.3%). Further, on univariable analysis of risk factors, species (cattle) (OR:3.31; 95%CI:1.88-5.82), inadequate floor spacing (OR:1.64; 95%CI:1.10-2.43), mastitis (OR:2.35, 95%CI:1.45-3.81) and reproductive disorders (OR:2.54; 95%CI:1.67-3.85) were significantly (p<0.05) having high odds for coxiellosis. The multivariable logistic regression analysis of the animal level risk factors revealed that species and age were found to be significantly associated with coxiellosis. However, since the number of screened farms is limited; further research is needed with a higher number of animals to confirm the farm level odds ratio of risk factors. Quarantine and biosecurity measures including farm hygiene operations were observed to be inadequate and also the lack of awareness about coxiellosis among the farm workers. In absence of vaccination program for coxiellosis in India, robust surveillance, farm biosecurity measures and the awareness for the disease among risk groups can play an important role in the disease prevention and subsequent transmission of the pathogen., Competing Interests: The authors have declared that no competing interests exist.

Coxiella burnetii , the causative agent of Query (Q) fever in humans, is a highly infectious obligate intracellular bacterium. Following uptake into a host cell, C. burnetii replicates within a phagolysosome-derived compartment referred to as the Coxiella -containing vacuole (CCV). During infection, C. burnetii exhibits tropism for tissues related to iron storage and recycling (e.g., the liver and splenic red pulp), suggesting that pathogen physiology is linked to host iron metabolism. Iron has been described to have a limited role in C. burnetii virulence regulation, despite evidence that C. burnetii - infected host cells increase expression of transferrin receptors, thereby suggesting that active iron acquisition by the bacterium occurs upon infection. Through the use of host cell-free culture, C. burnetii was separated from the host cell in order to directly assess the role of different forms of iron in C. burnetii replication and viability, and therefore virulence. Results indicate that C. burnetii tolerates molecular iron over a broad concentration range (i.e., ∼0.001 to 1 mM) and undergoes gross loss of viability upon iron starvation. C. burnetii protein synthesis and energy metabolism, however, occur nearly uninhibited under iron concentrations not permissive to replication. Despite the apparent absence of genes related to acquisition of host-associated iron-containing proteins, C. burnetii replication is supported by hemoglobin, transferrin, and ferritin, likely due to release of iron from such proteins under acidic conditions. Moreover, chelation of host iron pools inhibited pathogen replication during infection of cultured cells. IMPORTANCE Host organisms restrict the availability of iron to invading pathogens in order to reduce pathogen replication. To counteract the host's response to infection, bacteria can rely on redundant mechanisms to obtain biologically diverse forms of iron during infection. C. burnetii appears specifically dependent on molecular iron for replication and viability and exhibits a response to iron akin to bacteria that colonize iron-rich environments. Physiological adaptation of C. burnetii to the unique acidic and degradative environment of the CCV is consistent with access of this pathogen to molecular iron., (Copyright © 2020 Sanchez and Omsland.)

Background: Following outbreaks in other parts of the Netherlands, the Dutch border region of South Limburg experienced a large-scale outbreak of human Q fever related to a single dairy goat farm in 2009, with surprisingly few cases reported from neighbouring German counties. Late chronic Q fever, with recent spikes of newly detected cases, is an ongoing public health concern in the Netherlands. We aimed to assess the scope and scale of any undetected cross-border transmission to neighbouring German counties, where individuals unknowingly exposed may carry extra risk of overlooked diagnosis., Methods: (A) Seroprevalence rates in the Dutch area were estimated fitting an exponential gradient to the geographical distribution of notified acute human Q fever cases, using seroprevalence in a sample of farm township inhabitants as baseline. (B) Seroprevalence rates in 122 neighbouring German postcode areas were estimated from a sample of blood donors living in these areas and attending the regional blood donation centre in January/February 2010 (n = 3,460). (C) Using multivariate linear regression, including goat and sheep densities, veterinary Q fever notifications and blood donor sampling densities as covariates, we assessed whether seroprevalence rates across the entire border region were associated with distance from the farm., Results: (A) Seroprevalence in the outbreak farm's township was 16.1%. Overall seroprevalence in the Dutch area was 3.6%. (B) Overall seroprevalence in the German area was 0.9%. Estimated mean seroprevalence rates (per 100,000 population) declined with increasing distance from the outbreak farm (0-19 km = 2,302, 20-39 km = 1,122, 40-59 km = 432 and ≥60 km = 0). Decline was linear in multivariate regression using log-transformed seroprevalence rates (0-19 km = 2.9 [95% confidence interval (CI) = 2.6 to 3.2], 20 to 39 km = 1.9 [95% CI = 1.0 to 2.8], 40-59 km = 0.6 [95% CI = -0.2 to 1.3] and ≥60 km = 0.0 [95% CI = -0.3 to 0.3])., Conclusions: Our findings were suggestive of widespread cross-border transmission, with thousands of undetected infections, arguing for intensified cross-border collaboration and surveillance and screening of individuals susceptible to chronic Q fever in the affected area., (© 2020 The Authors. Transboundary and Emerging Diseases published by Blackwell Verlag GmbH.)

Coxiella burnetii causes Q fever in humans and coxiellosis in animals. In humans, it causes acute febrile illnesses like influenza, pneumonia, hepatitis, and chronic illnesses such as endocarditis, vascular infection, and post-infectious fatigue syndrome. It is widely distributed worldwide, and its main reservoirs are sheep, goats, and cattle. This study aimed to determine the frequency of C. burnetii infection using molecular detection and to identify the associated factors in livestock farmers and cattle from the Magdalena Medio region of Antioquia, Colombia. Using real-time polymerase chain reaction (PCR), molecular detection was performed for the IS1111 insertion sequence of C. burnetii using genomic DNA collected from the peripheral blood of 143 livestock farmers and 192 cattle from 24 farms located in Puerto Berrío, Puerto Nare, and Puerto Triunfo. To confirm the results, bidirectional amplicon sequencing of 16S rRNA was performed in four of the positive samples. Additionally, factors associated with C. burnetii were identified using a Poisson regression with cluster effect adjustment. Real-time PCR showed positive results in 25.9% and 19.5% of livestock farmer samples and cattle samples, respectively. For livestock farmers, factors associated with C. burnetii were the area where the farm was located [Puerto Berrío, adjusted prevalence ratio (aPR): 2.13, 95% confidence interval (CI): 1.10-4.11], presence of hens (aPR: 1.47, 95% CI: 1.21-1.79), horses (aPR: 1.61, 95% CI: 1.54-1.67), and ticks (aPR: 2.36, 95% CI: 1.03-5.42) in the residence, and consumption of raw milk (aPR: 1.47, 95% CI: 1.26-1.72). For cattle, the factors associated with Coxiella genus were municipality (Puerto Nare; aPR: 0.39, 95% CI: 0.37-0.41) and time of residence on the farm (≥49 months; aPR: 2.28, 95% CI: 1.03-5.20). By analyzing sequences of the 16S rRNA molecular marker, C. burnetii infection was confirmed in livestock farmers. However, in cattle, only the presence of Coxiella-type bacteria was identified. Further research is necessary to determine the potential role that these types of bacteria have as etiological agents for disease in livestock farmers and cattle from the study area., Competing Interests: The authors declare that there is no conflict of interest in this study.

Q fever, spotted fever rickettsioses and equine piroplasmosis, are some of the most serious equine tick-borne diseases caused by Coxiella burnetii, Rickettsia spp., Babesia caballi and/or Theileria equi. This study surveyed and molecularly characterized these pathogens infecting horses in ten ranches from XUAR, China using molecular technology. Among 200 horse blood samples, 163 (81.5%) were infected with at least one of the pathogens. Rickettsia spp. was the most prevalent pathogen (n = 114, 57.0%), followed by C. burnetii (n = 79, 39.5%), T. equi (n = 79, 39.5%) and B. caballi (n = 49, 24.5%). Co-infections were observed in 61.3% of positive samples in this study. Statistically significant differences were observed between the sampling regions for C. burnetii, B. caballi and T. equi, and also in different age group for C. burnetii and T. equi. The genotype analysis indicated that C. burnetii htpB, Rickettsia spp. ompA, B. caballi rap-1, B. caballi 18S rRNA, T. equi EMA-1 and T. equi 18S rRNA gene sequences from horses in XUAR were variable. To the best of our knowledge, this study is the first report of C. burnetii and Rickettsia spp. infection and co-infected with piroplasma in horses in China. Overall, this study revealed the high infection rate of the pathogens in horses in XUAR, China. The current findings are expected to provide a basis for better tick-borne disease control in the region., Competing Interests: Declaration of Competing Interest The authors declare that they have no competing interest., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Equine abortion is a cause of severe economic loss to the equine industry. Equine herpesvirus 1 is considered a primary cause of infectious abortion in horses, however other infectious agents can also cause abortion. Abortions due to zoonotic pathogens have implications for both human and animal health. We determined the prevalence of Coxiella burnetii, Leptospira spp. and Toxoplasma gondii in 600 aborted equine foetal tissues that were submitted to our diagnostic laboratories at the University of Melbourne from 1994 to 2019. Using qPCR we found that the prevalence of C. burnetii was 4%. The highest annual incidence of C. burnetii was observed between 1997-2003 and 2016-2018. The prevalence of C. burnetii in Victoria and New South Wales was 3% and 6% respectively. All the samples tested negative for Leptospira spp. and Toxoplasma gondii DNA. Equine herpesvirus 1 DNA was detected at a prevalence of 3%. This study has provided evidence for the presence of C. burnetii in equine aborted foetal tissues in Australia, but the role of C. burnetii as potential cause of abortion in Australia requires further investigation. C. burnetii is a zoonotic disease agent that causes the disease 'Q fever' in humans. We recommend that appropriate protective measures should be considered when handling material associated with equine abortions to reduce the risk of becoming infected with C. burnetii., Competing Interests: The authors have declared that no competing interests exist.

Coxiella burnetii is an obligate intracellular bacterial pathogen responsible for severe worldwide outbreaks of the zoonosis Q fever. The remarkable resistance to environmental stress, extremely low infectious dose and ease of dissemination, contributed to the classification of C. burnetii as a class B biothreat. Unique among intracellular pathogens, C. burnetii escapes immune surveillance and replicates within large autophagolysosome-like compartments called Coxiella-containing vacuoles (CCVs). The biogenesis of these compartments depends on the subversion of several host signalling pathways. For years, the obligate intracellular nature of C. burnetii imposed significant experimental obstacles to the study of its pathogenic traits. With the development of an axenic culture medium in 2009, C. burnetii became genetically tractable, thus allowing the implementation of mutagenesis tools and screening approaches to identify its virulence determinants and investigate its complex interaction with host cells. Here, we review the key advances that have contributed to our knowledge of C. burnetii pathogenesis, leading to the rise of this once-neglected pathogen to an exceptional organism to study the intravacuolar lifestyle., (© 2020 John Wiley & Sons Ltd.)

Coxiella burnetii is an intracellular bacterium that causes acute and chronic Q fever. This unique pathogen has been historically challenging to study due to obstacles in genetically manipulating the organism and the inability of small animal models to fully mimic human Q fever. Here, we review the current state of C. burnetii research, highlighting new approaches that allow the mechanistic study of infection in disease relevant settings., Competing Interests: Declaration of Competing Interest There is no conflict of interest., (Copyright © 2019 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.)

Coxiella burnetii is an intracellular, gram-negative bacterium that causes the zoonosis Q fever. This disease typically presents as an acute flu-like illness with persistent, focalized infections occurring less frequently. Clinical outcomes of Q fever have been associated with distinct genomic groups of C. burnetii, suggesting that gene content is responsible for virulence potential. To investigate this hypothesis, the virulence of thirteen C. burnetii strains (representing genomic groups I-VI) was evaluated in a guinea pig infection model by intraperitoneal injection. Seven strains caused a sustained fever (at least two days ≥39.5°C) in at least half of the animals within each experimental group. At fourteen days post infection, animals were euthanized and additional endpoints were evaluated, including splenomegaly and serology. The magnitude of these endpoints roughly correlated with the onset, duration, and severity of fever. The most severe disease was caused by group I strains. Intermediate and no virulence were evidenced following infection with group II-V and group VI strains, respectively. Flow cytometric analysis of the mesenteric lymph nodes revealed decreased CD4 + T cell frequency following infection with highly virulent group I strains. These findings buttress the hypothesis that the pathogenic potential of C. burnetii strains correlates with genomic grouping. These data, combined with comparative genomics and genetic manipulation, will improve our understanding of C. burnetii virulence determinants.

Background: As Q fever, caused by Coxiella burnetii, is a major health challenge due to its cardiovascular complications, we aimed to detect acute Q fever valvular injury to improve therapeutic management., Methods: In the French national reference center for Q fever, we prospectively collected data from patients with acute Q fever and valvular injury. We identified a new clinical entity, acute Q fever endocarditis, defined as valvular lesion potentially caused by C. burnetii: vegetation, valvular nodular thickening, rupture of chorda tendinae, and valve or chorda tendinae thickness. To determine whether or not the disease was superimposed on an underlying valvulopathy, patients' physicians were contacted. Aortic bicuspidy, valvular stenosis, and insufficiency were considered as underlying valvulopathies., Results: Of the 2434 patients treated in our center, 1797 had acute Q fever and 48 had acute Q fever endocarditis. In 35 cases (72%), transthoracic echocardiography (TTE) identified a valvular lesion of acute Q fever endocarditis without underlying valvulopathy. Positive anticardiolipin antibodies (>22 immunoglobulin G-type phospholipid units [GPLU]) were independently associated with acute Q fever endocarditis (odds ratio [OR], 2.7 [95% confidence interval {CI}, 1.3-5.5]; P = .004). Acute Q fever endocarditis (OR, 5.2 [95% CI, 2.6-10.5]; P < .001) and age (OR, 1.7 [95% CI, 1.1-1.9]; P = .02) were independent predictors of progression toward persistent C. burnetii endocarditis., Conclusions: Systematic TTE in acute Q fever patients offers a unique opportunity for early diagnosis of acute Q fever endocarditis and for the prevention of persistent endocarditis. Transesophageal echocardiography should be proposed in men, aged >40 years, with anticardiolipin antibodies >60 GPLU when TTE is inconclusive or negative., (© The Author(s) 2019. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

Coxiellosis is a zoonotic disease caused by the obligate intracellular bacterium Coxiella burnetii affecting the productive and reproductive capabilities of animals. This study was conducted to gain insight into the seroprevalence of coxiellosis in small ruminants in seven farms of the Punjab, Pakistan. Potential risk factors were assessed. In total, 1000 serum samples (500 from sheep and 500 from goats) and 163 ticks were collected from the ruminants. All these 163 ticks were merged into 55 pools (29 pools for ticks from sheep and 26 pools for ticks from goat). Serum samples were investigated using an indirect ELISA and PCR. Coxiella burnetii DNA was detected in 29 pooled seropositive samples and 11 pooled ticks by real-time qPCR. Serological analysis revealed a prevalence of 15.6% and 15.0% in sheep and goats, respectively. A significant association was found between seropositivity and different variables like district, lactational status, reproductive status, body condition and reproductive disorders. Univariate analysis showed that detection of C. burnetii DNA in tick pools was significantly associated with the presence of ticks on sheep and goats. However, a non-significant association was found for the prevalence of C. burnetii DNA in serum pools. Hence, C. burnetii infection is prevalent in small ruminants and ticks maintained at livestock farms in Punjab, Pakistan.

Coxiella burnetii is a Gram-negative bacterium which causes Q fever, a complex and life-threatening infection with both acute and chronic presentations. C. burnetii invades a variety of host cell types and replicates within a unique vacuole derived from the host cell lysosome. In order to understand how C. burnetii survives within this intracellular niche, we have investigated the carbon metabolism of both intracellular and axenically cultivated bacteria. Both bacterial populations were shown to assimilate exogenous [13C]glucose or [13C]glutamate, with concomitant labeling of intermediates in glycolysis and gluconeogenesis, and in the TCA cycle. Significantly, the two populations displayed metabolic pathway profiles reflective of the nutrient availabilities within their propagated environments. Disruption of the C. burnetii glucose transporter, CBU0265, by transposon mutagenesis led to a significant decrease in [13C]glucose utilization but did not abolish glucose usage, suggesting that C. burnetii express additional hexose transporters which may be able to compensate for the loss of CBU0265. This was supported by intracellular infection of human cells and in vivo studies in the insect model showing loss of CBU0265 had no impact on intracellular replication or virulence. Using this mutagenesis and [13C]glucose labeling approach, we identified a second glucose transporter, CBU0347, the disruption of which also showed significant decreases in 13C-label incorporation but did not impact intracellular replication or virulence. Together, these analyses indicate that C. burnetii may use multiple carbon sources in vivo and exhibits greater metabolic flexibility than expected., (© 2019 The Author(s).)

Coxiella burnetii is an obligate intracellular pathogen that causes the zoonotic disease Q fever in humans, which can occur in either an acute or a chronic form with serious complications. The bacterium has a wide host range, including unicellular organisms, invertebrates, birds and mammals, with livestock representing the most significant reservoir for human infections. Cell culture models have been used to decipher the intracellular lifestyle of C. burnetii , and several infection models, including invertebrates, rodents and non-human primates, are being used to investigate host-pathogen interactions and to identify bacterial virulence factors and vaccine candidates. However, none of the models replicate all aspects of human disease. Furthermore, it is becoming evident that C. burnetii isolates belonging to different lineages exhibit differences in their virulence in these models. Here, we compare the advantages and disadvantages of commonly used infection models and summarize currently available data for lineage-specific virulence.

Infection with Coxiella burnetii , the causative agent of Q fever, can result in life-threatening persistent infection. Reactogenicity hinders worldwide implementation of the only licensed human Q fever vaccine. We previously demonstrated long-lived immunoreactivity in individuals with past symptomatic and asymptomatic Coxiella infection (convalescents) to promiscuous HLA class II C. burnetii epitopes, providing the basis for a novel T-cell targeted subunit vaccine. In this study, we investigated in a cohort of 22 individuals treated for persistent infection (chronic Q fever) whether they recognize the same set of epitopes or distinct epitopes that could be candidates for a therapeutic vaccine or aid in the diagnosis of persistent infection. In cultured enzyme-linked immunosorbent spot (ELISpot) assays, individuals with chronic Q fever showed strong class II epitope-specific responses that were largely overlapping with the peptide repertoire identified previously for convalescents. Five additional peptides were recognized more frequently by chronic subjects, but there was no combination of epitopes uniquely recognized by or nonreactive in subjects with chronic Q fever. Consistent with more recent/prolonged exposure, we found, however, stronger ex vivo responses by direct ELISpot to both whole-cell C. burnetii and individual peptides in chronic patients than in convalescents. In conclusion, we have validated and expanded a previously published set of candidate epitopes for a novel T-cell targeted subunit Q fever vaccine in treated patients with chronic Q fever and demonstrated that they successfully mounted a T-cell response comparable to that of convalescents. Finally, we demonstrated that individuals treated for chronic Q fever mount a broader ex vivo response to class II epitopes than convalescents, which could be explored for diagnostic purposes., (Copyright © 2019 American Society for Microbiology.)

Objective: Q fever epidemic outbreaks have been reported in French Guiana and in The Netherlands. To determine whether the C. burnetii strains involved in these epidemics had a peculiar virulence pattern, we compared the pathogenicity of the Guiana and the German strain (a clone of The Netherlands strain), in silico, in vitro, and in vivo versus the Nine Mile strain., Method: The pan-genomes of the Guiana (Cb175), German (Z3055), and the referent Nine Mile (RSA 493) C. burnetii strains were compared. In vitro, the growth rate and the morphological presentation were compared. In vivo (SCID and Balb/c mice), weight loss, histological lesions, C. burnetii bacterial load in deep organs, and serological response were reported according to each C. burnetii strain studied., Results: The Guiana strain had 77 times more missing genes and 12 times more unique genes than the German strain. The Guiana strain presented as large cell variants (LCVs) and led to the most pronounced fatality rate in SCID mice (100% at 4 weeks). The German strain presented as small cell variants (SCVs), and had an intermediate fatality rate (75% at 4 weeks). Both the Guiana and the German strains led to a significant higher serological response at 2 and 4 weeks post infection (p <0.05)., Conclusion: The Guiana strain was the most virulent strain, followed by the German strain and the referent Nine Mile strain. Unique and missing genes could be implicated but further investigations are necessary to specify their role., (Copyright © 2019 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

Most intracellular pathogens that reside in a vacuole prevent transit of their compartment to lysosomal organelles. Effector mechanisms induced by the pro-inflammatory cytokine Interferon-gamma (IFNγ) can promote the delivery of pathogen-occupied vacuoles to lysosomes for proteolytic degradation and are therefore important for host defense against intracellular pathogens. The bacterial pathogen Coxiella burnetii is unique in that, transport to the lysosome is essential for replication. The bacterium modulates membrane traffic to create a specialized autophagolysosomal compartment called the Coxiella-containing vacuole (CCV). Importantly, IFNγ signaling inhibits intracellular replication of C. burnetii, raising the question of which IFNγ-activated mechanisms restrict replication of a lysosome-adapted pathogen. To address this question, siRNA was used to silence a panel of IFNγ-induced genes in HeLa cells to identify genes required for restriction of C. burnetii intracellular replication. This screen demonstrated that Indoleamine 2,3-dioxygenase 1 (IDO1) contributes to IFNγ-mediated restriction of C. burnetii. IDO1 is an enzyme that catabolizes cellular tryptophan to kynurenine metabolites thereby reducing tryptophan availability in cells. Cells deficient in IDO1 function were more permissive for C. burnetii replication when treated with IFNγ, and supplementing IFNγ-treated cells with tryptophan enhanced intracellular replication. Additionally, ectopic expression of IDO1 in host cells was sufficient to restrict replication of C. burnetii in the absence of IFNγ signaling. Using differentiated THP1 macrophage-like cells it was determined that IFNγ-activation resulted in IDO1 production, and that supplementation of IFNγ-activated THP1 cells with tryptophan enhanced C. burnetii replication. Thus, this study identifies IDO1 production as a key cell-autonomous defense mechanism that limits infection by C. burnetii, which suggests that peptides derived from hydrolysis of proteins in the CCV do not provide an adequate supply of tryptophan for bacterial replication., Competing Interests: The authors have declared that no competing interests exist.

Background: From 2007 through 2010, a large epidemic of acute Q fever occurred in the Netherlands. Patients with cardiac valvulopathy are at high risk to develop chronic Q fever after an acute infection. This patient group was not routinely screened, so it is unknown whether all their chronic infections were diagnosed. This study aims to investigate how many chronic Q fever patients can be identified by routinely screening patients with valvulopathy and to establish whether the policy of not screening should be changed., Methods: In a cross-sectional study (2016-2017) in a hospital at the epicentre of the Q fever epidemic, a blood sample was taken from patients 18 years and older who presented with cardiac valvulopathy. The sample was tested for IgG antibodies against phase I and II of Coxiella burnetii using an immunofluorescence assay. An IgG phase II titre of ≥1:64 was considered serological evidence of a previous Q fever infection. An IgG phase I titre of ≥1:512 was considered suspicious for a chronic infection, and these patients were referred for medical examination., Results: Of the 904 included patients, 133 (15%) had evidence of a previous C. burnetii infection, of whom 6 (5%) had a chronic infection on medical examination., Conclusions: In a group of high-risk patients with a heart valve defect, we diagnosed new chronic Q fever infections seven years after the epidemic, emphasizing the need for screening of this group to prevent complications in those not yet diagnosed in epidemic areas., Competing Interests: The authors have declared that no competing interests exist.

Respiratory tract infections (RTIs) are severe acute infectious diseases, which require the timely and accurate identification of the pathogens involved so that the individual treatment plan can be selected, including optimized use of antibiotics. However, high throughput and ultrasensitive quantification of multiple nucleic acids is a challenge in a point of care testing (POCT) device. Methods: Herein, we developed a 2×3 microarray on a lateral flow strip with surface enhanced Raman scattering (SERS) nanotags encoding the nucleic acids of 11 common RTI pathogens. On account of the signal magnification of encoded SERS nanotags in addition to the high surface area to volume ratio of the nitrocellulose (NC) membrane, rapid quantification of the 11 pathogens with a broad linear dynamic range (LDR) and ultra-high sensitivity was achieved on one lateral flow microarray. Results: The limit of detection (LOD) for influenza A, parainfluenza 1, parainfluenza 3, respiratory syncytial virus, coxiella burnetii, legionella pneumophila, influenza B, parainfluenza 2, adenovirus, chlamydophila pneumoniae, and mycoplasma pneumoniae were calculated to be 0.031 pM, 0.030 pM, 0.038 pM, 0.038 pM, 0.040 pM, 0.039 pM, 0.035 pM, 0.032 pM, 0.040 pM, 0.039 pM, and 0.041 pM, respectively. The LDR of measurement of the target nucleic acids of the eleven RTI pathogens were 1 pM-50 nM, which span 5 orders of magnitude. Conclusions: We anticipate this novel approach could be widely adopted in the early and precise diagnosis of RTI and other diseases., Competing Interests: Competing Interests: The authors have declared that no competing interest exists.

Cadherins switching is a hallmark of neoplasic processes. The E-cadherin (E-cad) subtype is one of the surface molecules regulating cell-to-cell adhesion. After its cleavage by sheddases, a soluble fragment (sE-cad) is released that has been identified as a pro-carcinogenic inflammatory signal in several bacteria-induced cancers. Recently we reported that Q fever, a disease due to Coxiella burnetii infection, can be complicated by occurrence of non-Hodgkin lymphoma (NHL). Therefore, we studied E-cad switching in Q fever. The sE-cad levels were found increased in the sera of acute and persistent Q fever patients, whereas they remained at the baseline in controls groups of healthy donors, people cured of Q fever, patients suffering from unrelated inflammatory diseases, and past Q fever patients who developed NHL. These results indicate that sE-cad can be considered as a new biomarker of C. burnetii infection rather than a marker of NHL-associated to Q fever. We wondered if changes in sE-cad reflected variations in the CDH1 gene transcription. The expression of E-cad mRNA and its intracellular ligand β-catenin was down-regulated in peripheral blood mononuclear cells (PBMCs) of patients with either acute or persistent forms of Q fever. Indeed, a lower cell-surface expression of E-cad was measured in a minority (<5%) subpopulation of HLADR + /CD16 + monocytes from patients with acute Q fever. However, a very strong increase in E-cad expression was observed on more than 30% of the HLADR + /CD16 + monocytes of persistent Q fever patients, a cell subpopulation known to be a target for C. burnetii in humans. An experimental in vitro infection of healthy donors' PBMCs with C. burnetii , was performed to directly evaluate the link between C. burnetii interaction with PBMCs and their E-cad expression. A significant increase in the percentage of HLADR + /CD16 + monocytes expressing E-cad was measured after PBMCs had been incubated for 8 h with C. burnetii Nine Mile strain. Altogether, these data demonstrate that C. burnetii severely impairs the E-cad expression in circulating cells of Q fever patients.

Coxiella burnetii, the agent causing Q fever, has been associated with B-cell non-Hodgkin lymphoma (NHL). To better clarify this link, we analysed the genetic transcriptomic profile of peripheral blood leukocytes from patients with C. burnetii infection to identify possible links to lymphoma. Microarray analyses revealed that 1189 genes were expressed differently (p <.001 and fold change ≥4) in whole blood of patients with C. burnetii infection compared to controls. In addition, 95 genes expressed in patients with non-Hodgkin lymphoma (NHL) and in patients with C. burnetii persistent infection have allowed us to establish the 'C. burnetii-associated NHL signature'. Among these, 33 genes previously found modulated in C. burnetii-associated -NHL by the microarray analysis were selected and their mRNA expression levels were measured in distinct C. burnetii-induced pathologies, namely, acute Q fever, focalized persistent infection, lymphadenitis and C.burnetii-associated NHL. Specific genes involved in anti-apoptotic process were found highly expressed in leukocytes from patients with C. burnetii associated-NHL: MIR17HG, REL and SP100. This signature differed from that found for NHL-control group. Patients with C. burnetii lymphadenitis presented significant elevated levels of BCL2 and ETS1 mRNAs. Altogether, we identified a specific transcriptionnal signature for NHL during C. burnetii infection reflecting the up-regulation of anti-apoptotic processes and the fact that lymphadenitis might constitute a critical step towards lymphomagenesis., Competing Interests: The authors have declared that no competing interests exist.

Background: Head louse, Pediculus humanus capitis, is an obligatory blood-sucking ectoparasite, distributed worldwide. Phylogenetically, it occurs in five divergent mitochondrial clades (A-E); each exhibiting a particular geographical distribution. Recent studies suggest that, as in the case of body louse, head louse could be a disease vector. We aimed to study the genetic diversity of head lice collected in the Democratic Republic of the Congo (DR Congo) and to screen for louse-borne pathogens in these lice., Methods: A total of 181 head lice were collected from 27 individuals at the Monkole Hospital Center located in Kinshasa. All head lice were genotyped and screened for the presence of louse-borne bacteria using molecular methods. We searched for Bartonella quintana, Borrelia recurrentis, Rickettsia prowazekii, Anaplasma spp., Yersinia pestis, Coxiella burnetii and Acinetobacter spp., Results: Among these head lice, 67.4% (122/181) belonged to clade A and 24.3% (44/181) belonged to clade D. Additionally, for the first time in this area, we found clade E in 8.3% (15/181) of tested lice, from two infested individuals. Dual infestation with clades A and D was observed for 44.4% individuals. Thirty-three of the 181 head lice were infected only by different bacterial species of the genus Acinetobacter. Overall, 16 out of 27 individuals were infested (59.3%). Six Acinetobacter species were detected including Acinetobacter baumannii (8.3%), Acinetobacter johnsonii (1.7%), Acinetobacter soli (1.7%), Acinetobacter pittii (1.7%), Acinetobacter guillouiae (1.1%), as well as a new potential species named "Candidatus Acinetobacter pediculi"., Conclusions: To our knowledge, this study reports for the first time, the presence of clade E head lice in DR Congo. This study is also the first to report the presence of Acinetobacter species DNAs in human head lice in DR Congo.

Coxiellosis is a zoonotic disease caused by the ubiquitous bacteria Coxiella burnetii, which can be spread either through ticks or through body fluids. In humans the infection is characterized by a febrile disease; ruminants may abort and reduce their milk yield, causing serious production losses for the farmer. In India, the disease has been known to be present since the 1970s, but little is known about the epidemiology in most states. In this cross-sectional survey in the two states of Assam and Bihar, 520 households were interviewed for risk factors, and serum samples from 744 dairy animals were analyzed using ELISA as well as PCR. Out of the farms, 17.4% had at least one seropositive animal, with significantly higher seroprevalence in Bihar (27.1%) than Assam (5.8%); and significantly more sero-positive farms in urban areas (23.1%) than rural (12.2%). On an individual animal level, 14.1% were seropositive, with higher prevalence among buffaloes than cows (28.0% versus 13.6%). Out of the seropositive animals, 10.6% had aborted during the last three years, and 37.5% had experienced problems with repeat breeding: both higher than in non-seropositive animals. In conclusion, this study indicates that coxiellosis is potentially an important cause of reproductive failures and production losses in dairy animals. The high prevalence, especially in urban areas, is a public health risk. Further research is needed to elucidate the epidemiology and identify mitigation options that could work in the different settings of different Indian states., (Copyright © 2019. Published by Elsevier B.V.)

The present review aims to compile the currently available literature since 1936 according the sources of infection of the Q fever pathogen (Coxiella (C.) burnetii) as well as the transmission from animal to man and also from human to human. In terms of quality and validity, the existing publications were reviewed systematically. For this purpose, firstly a structured literature search was carried out using various databases and search engines supplemented by a manual literature search. For critical appraisal, 1444 relevant publications were identified for the moment and evaluated. A total of 73 publications describing a transmission of C. burnetii from animals to man or a human-to-human transmission were discovered. The identified publications are 29 case series, two case reports, 21 cohort studies and 21 case-control studies. With regard to the sources of infection, 25 publications describing the transmission of C. burnetii from sheep to humans could be identified., (Copyright © 2019. Published by Elsevier Ltd.)

Human Q fever is caused by the intracellular bacterial pathogen Coxiella burnetii Q fever presents with acute flu-like and pulmonary symptoms or can progress to chronic, severe endocarditis. After human inhalation, C. burnetii is engulfed by alveolar macrophages and transits through the phagolysosomal maturation pathway, resisting the acidic pH of lysosomes to form a parasitophorous vacuole (PV) in which to replicate. Previous studies showed that C. burnetii replicates efficiently in primary human alveolar macrophages (hAMs) in ex vivo human lung tissue. Although C. burnetii replicates in most cell types in vitro , the pathogen does not grow in non-hAM cells of human lung tissue. In this study, we investigated the interaction between C. burnetii and other pulmonary cell types apart from the lung environment. C. burnetii formed a prototypical PV and replicated efficiently in human pulmonary fibroblasts and in airway, but not alveolar, epithelial cells. Atypical PV expansion in alveolar epithelial cells was attributed in part to defective recruitment of autophagy-related proteins. Further assessment of the C. burnetii growth niche showed that macrophages mounted a robust interleukin 8 (IL-8), neutrophil-attracting response to C. burnetii and ultimately shifted to an M2-polarized phenotype characteristic of anti-inflammatory macrophages. Considering our findings together, this study provides further clarity on the unique C. burnetii -lung dynamic during early stages of human acute Q fever., (Copyright © 2019 American Society for Microbiology.)

The entry of pathogens into nonphagocytic host cells has received much attention in the past three decades, revealing a vast array of strategies employed by bacteria and viruses. A method of internalization that has been extensively studied in the context of viral infections is the use of the clathrin-mediated pathway. More recently, a role for clathrin in the entry of some intracellular bacterial pathogens was discovered. Classically, clathrin-mediated endocytosis was thought to accommodate internalization only of particles smaller than 150 nm; however, this was challenged upon the discovery that Listeria monocytogenes requires clathrin to enter eukaryotic cells. Now, with discoveries that clathrin is required during other stages of some bacterial infections, another paradigm shift is occurring. There is a more diverse impact of clathrin during infection than previously thought. Much of the recent data describing clathrin utilization in processes such as bacterial attachment, cell-to-cell spread and intracellular growth may be due to newly discovered divergent roles of clathrin in the cell. Not only does clathrin act to facilitate endocytosis from the plasma membrane, but it also participates in budding from endosomes and the Golgi apparatus and in mitosis. Here, the manipulation of clathrin processes by bacterial pathogens, including its traditional role during invasion and alternative ways in which clathrin supports bacterial infection, is discussed. Researching clathrin in the context of bacterial infections will reveal new insights that inform our understanding of host-pathogen interactions and allow researchers to fully appreciate the diverse roles of clathrin in the eukaryotic cell., (Copyright © 2019 American Society for Microbiology.)

Microtubules (Mts) are dynamic cytoskeleton structures that play a key role in vesicular transport. The Mts-mediated transport depends on motor proteins named kinesins and the dynein/dynactin motor complex. The Rab7 adapter protein FYCO1 controls the anterograde transport of the endocytic compartments through the interaction with the kinesin KIF5. Rab7 and its partner RILP induce the recruitment of dynein/dynactin to late endosomes regulating its retrograde transport to the perinuclear area to fuse with lysosomes. The late endosomal-lysosomal fusion is regulated by the HOPS complex through its interaction with RILP and the GTPase Arl8. Coxiella burnetii (Cb), the causative agent of Q fever, is an obligate intracellular pathogen, which generates a large compartment with autophagolysosomal characteristics named Cb-containing vacuole (CCV). The CCV forms through homotypic fusion between small non-replicative CCVs (nrCCV) and through heterotypic fusion with other compartments, such as endosomes and lysosomes. In this work, we characterise the role of Mts, motor proteins, RILP/Rab7 and Arl8 on the CCV biogenesis. The formation of the CCV was affected when either the dynamics and/or the acetylation state of Mts were modified. Similarly, the overexpression of the dynactin subunit non-functional mutants p150Glued and RILP led to the formation of small nrCCVs. This phenomenon is not observed in cells overexpressing WT proteins, the motor KIF5 or its interacting protein FYCO1. The formation of the CCV was normal in infected cells that overexpressed Arl8 alone or together with hVps41 (a HOPS subunit) or in cells co-overexpressing hVps41 and RILP. The dominant negative mutant of Arl8 and the non-functional hVps41 inhibited the formation of the CCV. When the formation of CCV was affected, the bacterial multiplication diminished. Our results suggest that nrCCVs recruit the molecular machinery that regulate the Mts-dependent retrograde transport, Rab7/RILP and the dynein/dynactin system, as well as the tethering processes such as HOPS complex and Arl8 to finally originate the CCV where C. burnetii multiplies., Competing Interests: The authors have declared that no competing interests exist.

Laboratory testing for Q fever (Coxiella burnetii) is essential for a differential diagnosis, yet little is known about Q fever diagnostic testing practices in the United States. We retrospectively analyzed Q fever immunoglobulin G (IgG) indirect immunofluorescence assay (IFA) testing data between 1/1/2012-10/31/2016 from ARUP, LabCorp, Mayo Medical Laboratories, and Quest Diagnostics. Data included IgG phase I and phase II titers, patient age and sex, and state and date of specimen collection. On average, 12,821 specimens were tested for Q fever annually by the participating laboratories. Of 64,106 total specimens, 84.1% tested negative for C. burnetii-specific antibodies. Positive titers ranged from 16 to 262,144 against both phase I and phase II antigens. Submission of specimens peaked during the summer months, and more specimens were submitted from the West North Central division. Testing occurred more frequently in males (53%) and increased with age. In conclusion, few U.S. Q fever cases are reported, despite large volumes of diagnostic specimens tested. Review of commercial laboratory data revealed a lack of paired serology samples and patterns of serology titers that differ from case reporting diagnostic criteria.

Ticks are important vectors of pathogens affecting humans and animals worldwide. They do not only carry pathogens but diverse commensal and symbiotic microorganisms are also present in ticks. A molecular screening for tick-borne pathogens and endosymbionts was carried out in Ixodes ricinus, Dermacentor reticulatus and Haemaphysalis inermis questing ticks collected in Slovakia. The presence of Rickettsia spp., Coxiella burnetii, Coxiella-like and Francisella-like microorganisms was evaluated by PCR in 605 individuals and by randomly sequencing 66 samples. Four species of rickettsiae (R. raoultii, R. slovaca, R. helvetica and R. monacensis) were identified and reported with an overall prevalence range between 0.4 and 50.3% (±8.0) depending on tick species, sex and locality. Partial sequencing of the gltA gene of 5 chosen samples in H. inermis showed 99% identity with Candidatus Rickettsia hungarica. The total prevalence of C. burnetii in ticks was 2.2 ± 1.7%; bacteria were confirmed in I. ricinus and D. reticulatus ticks. The sequences from 2 D. reticulatus males and 1 I. ricinus female ticks were compared to GenBank submissions and a 99.8% match was obtained with the pathogenic C. burnetii. Coxiella-like endosymbionts were registered in all three species of ticks from all studied sites with an average prevalence of 32.7 ± 3.7%. A phylogenetic analysis of this Coxiella sp. showed that it does not group with the pathogenic C. burnetii. The prevalence of Francisella-like microorganisms in questing ticks was 47.9 ± 3.9%, however H. inermis (n = 108) were not infested. Obtained sequences were 98% identical with previously identified Francisella-like endosymbionts in D. reticulatus and I. ricinus. Coxiella-like and Francisella-like microorganisms were identified for the first time in Slovakia, they might be considered as a non-pathogenic endosymbiont of I. ricinus, D. reticulatus and H. inermis, and future investigations could aim to assess their role in these ticks. However, this work provided further data and broadened our knowledge on bacterial pathogens and endosymbionts present in ticks in Slovakia to help understanding co-infestations, combined treatments and public health issues linked to tick bites., (Copyright © 2018 Elsevier GmbH. All rights reserved.)

Lipid A is an essential basal component of lipopolysaccharide of most Gram-negative bacteria. Inhibitors targeting LpxC, a conserved enzyme in lipid A biosynthesis, are antibiotic candidates against Gram-negative pathogens. Here we report the characterization of the role of lipid A in Coxiella burnetii growth in axenic media, monkey kidney cells (BGMK and Vero), and macrophage-like THP-1 cells by using a potent LpxC inhibitor -LPC-011. We first determined the susceptibility of C. burnetii LpxC to LPC-011 in a surrogate E. coli model. In E. coli , the minimum inhibitory concentration (MIC) of LPC-011 against C. burnetii LpxC is < 0.05 μg/mL, a value lower than the inhibitor's MIC against E. coli LpxC. Considering the inhibitor's problematic pharmacokinetic properties in vivo and Coxiella 's culturing time up to 7 days, the stability of LPC-011 in cell cultures was assessed. We found that regularly changing inhibitor-containing media was required for sustained inhibition of C. burnetii LpxC in cells. Under inhibitor treatment, Coxiella has reduced growth yields in axenic media and during replication in non-phagocytic cells, and has a reduced number of productive vacuoles in such cells. Inhibiting lipid A biosynthesis in C. burnetii by the inhibitor was shown in a phase II strain transformed with chlamydial kdtA . This exogenous KdtA enzyme modifies Coxiella lipid A with an α-Kdo-(2 → 8)-α-Kdo epitope that can be detected by anti- chlamydia genus antibodies. In inhibitor-treated THP-1 cells, Coxiella shows severe growth defects characterized by poor vacuole formation and low growth yields. Coxiella progenies prepared from inhibitor-treated cells retain the capability of normally infecting all tested cells in the absence of the inhibitor, which suggests a dispensable role of lipid A for infection and early vacuole development. In conclusion, our data suggest that lipid A has significance for optimal development of Coxiella -containing vacuoles, and for robust multiplication of C. burnetii in macrophage-like THP-1 cells. Unlike many bacteria, C. burnetii replication in axenic media and non-phagocytic cells was less dependent on normal lipid A biosynthesis.

Q fever vaccine uptake among veterinary nurses in Australia is low, suggesting veterinarians are not recommending the vaccination to veterinary personnel. This study aimed to determine the willingness of veterinarians to recommend Q fever vaccination to veterinary personnel and to identify factors influencing Q fever vaccine uptake by veterinary nurses in Australia. An online cross sectional survey targeted veterinarians and veterinary nurses in Australia in 2014. Responses were analysed using multivariable logistic regression. Factors significantly (p<0.05) associated with a willingness to recommend the vaccination, expressed by 35% (95% CI 31-38%) of veterinarians (n = 828), were (1) being very concerned for colleagues regarding Coxiella burnetii (OR 4.73), (2) disagreeing the vaccine is harmful (OR 3.80), (3) high Q fever knowledge (OR 2.27), (4) working within small animal practice (OR 1.67), (5) disagreeing the vaccine is expensive (OR 1.55), and (6) age, with veterinarians under 39 years most likely to recommend vaccination. Of the veterinary nursing cohort who reported a known Q fever vaccination status (n = 688), 29% (95% CI 26-33%) had sought vaccination. This was significantly (p<0.05) associated with (1) agreeing the vaccine is important (OR 8.34), (2) moderate/high Q fever knowledge (OR 5.51), (3) working in Queensland (OR 4.00), (4) working within livestock/mixed animal practice (OR 3.24), (5) disagreeing the vaccine is expensive (OR 1.86), (6) strong reliance on work culture for biosecurity information (OR 2.5), (7) perceiving personal exposure to Coxiella burnetii to be at least low/moderate (OR 2.14), and (8) both agreeing the vaccine is safe and working within a corporate practice structure (OR 4.28). The study identified the need for veterinarians to take greater responsibility for workplace health and safety promotion, and calls for better education of veterinary personnel to raise awareness of the potential for occupational exposure to C. burnetii and improve the perception of the Q fever vaccine as being important, safe and cost-effective., Competing Interests: The authors have declared that no competing interests exist.

Background: The American dog tick, Dermacentor variabilis, is an important vector of pathogens to humans, wildlife and domestic animals in North America. Although this tick species is widely distributed in the USA and Canada, knowledge of its range-wide phylogeographic patterns remains incomplete., Methods: We carried out a phylogenetic analysis of D. variabilis using samples collected from 26 USA states and five Canadian provinces. Tick samples (n = 1053 in total) originated from two main sources: existing archives (2000-2011), and new collections made from 2012 to 2013. We sequenced a 691 bp fragment of the cox1 gene from a subset (n = 332) of geographically diverse D. variabilis. DNA extracted from individual ticks (n = 1053) was also screened for a Francisella-like endosymbiont, using a targeted 16S rRNA sequencing approach, and important pathogens (Rickettsia spp. and Coxiella burnetii), using species-specific quantitative PCR assays., Results: Maximum parsimony analysis of cox1 sequences revealed two major groups within D. variabilis with distinct geographical distributions: one from the eastern USA/Canada (Group 1) and one from the west coast states of the USA (California and Washington; Group 2). However, genetic subdivisions within both of these two major groups were weak to moderate and not tightly correlated with geography. We found molecular signatures consistent with Francisella-like endosymbionts in 257 of the DNA extracts from the 1053 individual ticks, as well as Rickettsia spp. and Coxiella burnetii in a small number of ticks (n = 29 and 2, respectively). Phylogenetic patterns for Francisella-like endosymbionts, constructed using sequence data from the bacterial 16S rRNA locus, were similar to those for D. variabilis, with two major groups that had a nearly perfect one-to-one correlation with the two major groups within D. variabilis., Conclusions: Our findings reveal a distinct phylogenetic split between the two major D. variabilis populations. However, high levels of genetic mixture among widely separated geographical localities occur within each of these two major groups. Furthermore, our phylogenetic analyses provide evidence of long-term tick-symbiont co-evolution. This work has implications for understanding the dispersal and evolutionary ecology of D. variabilis and associated vector-borne diseases.

Background: Q fever is a zoonotic disease caused by Coxiella burnetii. This bacterium survives harsh conditions and attaches to dust, suggesting environmental dispersal is a risk factor for outbreaks. Spatial epidemiology studies collating evidence on Q fever geographical contamination gradients are needed, as human cases without occupational exposure are increasing worldwide., Methods: We used a systematic literature search to assess the role of distance from ruminant holdings as a risk factor for human Q fever outbreaks. We also collated evidence for other putative drivers of C. burnetii geographical dispersal., Results: In all documented outbreaks, infective sheep or goats, not cattle, was the likely source. Evidence suggests a prominent role of airborne dispersal; Coxiella burnetii travels up to 18 km on gale force winds. In rural areas, highest infection risk occurs within 5 km of sources. Urban outbreaks generally occur over smaller distances, though evidence on attack rate gradients is limited. Wind speed / direction, spreading of animal products, and stocking density may all contribute to C. burnetii environmental gradients., Conclusions: Q fever environmental gradients depend on urbanization level, ruminant species, stocking density and wind speed. While more research is needed, evidence suggests that residential exclusion zones around holdings may be inadequate to contain this zoonotic disease, and should be species-specific.

Type IV secretion systems (T4SS) are multi-protein complexes in a number of bacterial pathogens that can translocate proteins and DNA to the host. Most T4SSs function in conjugation and translocate DNA; however, approximately 13% function to secrete proteins, delivering effector proteins into the cytosol of eukaryotic host cells. Upon entry, these effectors manipulate the host cell's machinery for their own benefit, which can result in serious illness or death of the host. For this reason recognition of T4SS effectors has become an important subject. Much previous work has focused on verifying effectors experimentally, a costly endeavor in terms of money, time, and effort. Having good predictions for effectors will help to focus experimental validations and decrease testing costs. In recent years, several scoring and machine learning-based methods have been suggested for the purpose of predicting T4SS effector proteins. These methods have used different sets of features for prediction, and their predictions have been inconsistent. In this paper, an optimal set of features is presented for predicting T4SS effector proteins using a statistical approach. A thorough literature search was performed to find features that have been proposed. Feature values were calculated for datasets of known effectors and non-effectors for T4SS-containing pathogens for four genera with a sufficient number of known effectors, Legionella pneumophila, Coxiella burnetii, Brucella spp, and Bartonella spp. The features were ranked, and less important features were filtered out. Correlations between remaining features were removed, and dimensional reduction was accomplished using principal component analysis and factor analysis. Finally, the optimal features for each pathogen were chosen by building logistic regression models and evaluating each model. The results based on evaluation of our logistic regression models confirm the effectiveness of our four optimal sets of features, and based on these an optimal set of features is proposed for all T4SS effector proteins.

Coxiella burnetii is the causative agent of human Q fever, a debilitating flu-like illness that can progress to chronic disease presenting as endocarditis. Following inhalation, C. burnetii is phagocytosed by alveolar macrophages and generates a lysosome-like replication compartment termed the parasitophorous vacuole (PV). A type IV secretion system (T4SS) is required for PV generation and is one of the pathogen's few known virulence factors. We previously showed that C. burnetii actively recruits autophagosomes to the PV using the T4SS but does not alter macroautophagy. In the current study, we confirmed that the cargo receptor p62/sequestosome 1 (SQSTM-1) localizes near the PV in primary human alveolar macrophages infected with virulent C. burnetii p62 and LC3 typically interact to select cargo for autophagy-mediated degradation, resulting in p62 degradation and LC3 recycling. However, in C. burnetii -infected macrophages, p62 was not degraded when cells were starved, suggesting that the pathogen stabilizes the protein. In addition, phosphorylated p62 levels increased, indicative of activation, during infection. Small interfering RNA experiments indicated that p62 is not absolutely required for intracellular growth, suggesting that the protein serves a signaling role during infection. Indeed, the Nrf2-Keap1 cytoprotective pathway was activated during infection, as evidenced by sustained maintenance of Nrf2 levels and translocation of the protein to the nucleus in C. burnetii -infected cells. Collectively, our studies identify a new p62-regulated host signaling pathway exploited by C. burnetii during intramacrophage growth., (Copyright © 2018 American Society for Microbiology.)