Species-level Microbiota of Biting Midges and Ticks from Poyang Lake.

Objective: The purpose of this study was to investigate the bacterial communities of biting midges and ticks collected from three sites in the Poyang Lake area, namely, Qunlu Practice Base, Peach Blossom Garden, and Huangtong Animal Husbandry, and whether vectors carry any bacterial pathogens that may cause diseases to humans, to provide scientific basis for prospective pathogen discovery and disease prevention and control. Methods: Using a metataxonomics approach in concert with full-length 16S rRNA gene sequencing and operational phylogenetic unit (OPU) analysis, we characterized the species-level microbial community structure of two important vector species, biting midges and ticks, including 33 arthropod samples comprising 3,885 individuals, collected around Poyang Lake. Results: A total of 662 OPUs were classified in biting midges, including 195 known species and 373 potentially new species, and 618 OPUs were classified in ticks, including 217 known species and 326 potentially new species. Surprisingly, OPUs with potentially pathogenicity were detected in both arthropod vectors, with 66 known species of biting midges reported to carry potential pathogens, including Asaia lannensis and Rickettsia bellii, compared to 50 in ticks, such as Acinetobacter lwoffii and Staphylococcus sciuri. We found that Proteobacteria was the most dominant group in both midges and ticks. Furthermore, the outcomes demonstrated that the microbiota of midges and ticks tend to be governed by a few highly abundant bacteria. Pantoea sp7 was predominant in biting midges, while Coxiella sp1 was enriched in ticks. Meanwhile, Coxiella spp., which may be essential for the survival of Haemaphysalis longicornis Neumann, were detected in all tick samples. The identification of dominant species and pathogens of biting midges and ticks in this study serves to broaden our knowledge associated to microbes of arthropod vectors. Conclusion: Biting midges and ticks carry large numbers of known and potentially novel bacteria, and carry a wide range of potentially pathogenic bacteria, which may pose a risk of infection to humans and animals. The microbial communities of midges and ticks tend to be dominated by a few highly abundant bacteria.

Exploring the diversity of tick-borne pathogens: The case of bacteria (Anaplasma, Rickettsia, Coxiella and Borrelia) protozoa (Babesia and Theileria) and viruses (Orthonairovirus, tick-borne encephalitis virus and louping ill virus) in the European continent.

Ticks are the main vectors for the transmission of bacterial, protist and viral pathogens in Europe affecting wildlife and domestic animals. However, some of them are zoonotic and can cause serious, sometimes fatal, problems in human health. A systematic review in PubMed/MEDLINE database was conducted to determine the spatial distribution and host and tick species ranges of a selection of tick-borne bacteria (Anaplasma spp., Borrelia spp., Coxiella spp., and Rickettsia spp.), protists (Babesia spp. and Theileria spp.), and viruses (Orthonairovirus, and flaviviruses tick-borne encephalitis virus and louping ill virus) on the European continent in a five-year period (November 2017 - November 2022). Only studies using PCR methods were selected, retrieving a total of 429 articles. Overall, up to 85 species of the selected tick-borne pathogens were reported from 36 European countries, and Anaplasma spp. was described in 37% (159/429) of the articles, followed by Babesia spp. (34%, 148/429), Borrelia spp. (34%, 147/429), Rickettsia spp. (33%, 142/429), Theileria spp. (11%, 47/429), tick-borne flaviviruses (9%, 37/429), Orthonairovirus (7%, 28/429) and Coxiella spp. (5%, 20/429). Host and tick ranges included 97 and 50 species, respectively. The highest tick-borne pathogen diversity was detected in domestic animals, and 12 species were shared between humans, wildlife, and domestic hosts, highlighting the following zoonotic species: Anaplasma phagocytophilum, Babesia divergens, Babesia microti, Borrelia afzelii, Borrelia burgdorferi s.s., Borrelia garinii, Borrelia miyamotoi, Crimean-Congo hemorrhagic fever virus, Coxiella burnetii, Rickettsia monacensis and tick-borne encephalitis virus. These results contribute to the implementation of effective interventions for the surveillance and control of tick-borne diseases.

Wild ungulates as sentinels of flaviviruses and tick-borne zoonotic pathogen circulation: an Italian perspective.

BACKGROUND: Vector-borne zoonotic diseases are a concerning issue in Europe. Lyme disease and tick-borne encephalitis virus (TBEV) have been reported in several countries with a large impact on public health; other emerging pathogens, such as Rickettsiales, and mosquito-borne flaviviruses have been increasingly reported. All these pathogens are linked to wild ungulates playing roles as tick feeders, spreaders, and sentinels for pathogen circulation. This study evaluated the prevalence of TBEV, Borrelia burgdorferi sensu lato, Rickettsia spp., Ehrlichia spp., and Coxiella spp. by biomolecular screening of blood samples and ticks collected from wild ungulates. Ungulates were also screened by ELISA and virus neutralization tests for flaviviral antibody detection. RESULTS: A total of 274 blood samples were collected from several wild ungulate species, as well as 406 Ixodes ricinus, which were feeding on them. Blood samples tested positive for B. burgdorferi s.l. (1.1%; 0-2.3%) and Rickettsia spp. (1.1%; 0-2.3%) and showed an overall flaviviral seroprevalence of 30.6% (22.1-39.2%): 26.1% (17.9-34.3%) for TBEV, 3.6% (0.1-7.1%) for Usutu virus and 0.9% (0-2.7%) for West Nile virus. Ticks were pooled when possible and yielded 331 tick samples that tested positive for B. burgdorferi s.l. (8.8%; 5.8-11.8%), Rickettsia spp. (26.6%; 21.8-31.2%) and Neoehrlichia mikurensis (1.2%; 0-2.4%). TBEV and Coxiella spp. were not detected in either blood or tick samples. CONCLUSIONS: This research highlighted a high prevalence of several tick-borne zoonotic pathogens and high seroprevalence for flaviviruses in both hilly and alpine areas. For the first time, an alpine chamois tested positive for anti-TBEV antibodies. Ungulate species are of particular interest due to their sentinel role in flavivirus circulation and their indirect role in tick-borne diseases and maintenance as Ixodes feeders and spreaders.

Detection of Coxiella antibodies in ruminants using a SucB recombinant antigen.

The detection of Coxiella burnetii in ruminants remains challenging despite the use of new technology and the accumulation of novel knowledge. Serology tools, the primary methods of infection surveillance in veterinary medicine, have limitations. We used recombinant antigen production to develop an ELISA based on the SucB protein, one of the major immunodominant antigens described in humans and laboratory animals. We produced the antigen successfully in an Escherichia coli heterologous system, confirmed by sequencing and mass spectrometry, and seen as a band of ~50 kDa in SDS-PAGE and on western blot analysis. We compared the performance of the recombinant ELISA with a commercial ELISA. We observed agreement of 83.5% and a substantial Cohen κ value of 0.67 in our pilot study.

Coxiella co-opts the Glutathione Peroxidase 4 to protect the host cell from oxidative stress-induced cell death.

The causative agent of human Q fever, Coxiella burnetii, is highly adapted to infect alveolar macrophages by inhibiting a range of host responses to infection. Despite the clinical and biological importance of this pathogen, the challenges related to genetic manipulation of both C. burnetii and macrophages have limited our knowledge of the mechanisms by which C. burnetii subverts macrophages functions. Here, we used the related bacterium Legionella pneumophila to perform a comprehensive screen of C. burnetii effectors that interfere with innate immune responses and host death using the greater wax moth Galleria mellonella and mouse bone marrow-derived macrophages. We identified MceF (Mitochondrial Coxiella effector protein F), a C. burnetii effector protein that localizes to mitochondria and contributes to host cell survival. MceF was shown to enhance mitochondrial function, delay membrane damage, and decrease mitochondrial ROS production induced by rotenone. Mechanistically, MceF recruits the host antioxidant protein Glutathione Peroxidase 4 (GPX4) to the mitochondria. The protective functions of MceF were absent in primary macrophages lacking GPX4, while overexpression of MceF in human cells protected against oxidative stress-induced cell death. C. burnetii lacking MceF was replication competent in mammalian cells but induced higher mortality in G. mellonella, indicating that MceF modulates the host response to infection. This study reveals an important C. burnetii strategy to subvert macrophage cell death and host immunity and demonstrates that modulation of the host antioxidant system is a viable strategy to promote the success of intracellular bacteria.

Molecular detection of Coxiella burnetii and Coxiella species in rats and chickens from poultry farms in North West Province, South Africa.

BACKGROUND: Coxiella burnetii is a bacterial pathogen that causes query fever and coxiellosis in humans and animals, respectively. There is a scarcity of studies on the prevalence of C. burnetii infections in rats and chickens in South Africa. OBJECTIVE: The aim of this study was to determine the occurrence of C. burnetii in rats and chickens sampled from poultry farms in the North West Province of South Africa. METHODS: DNA was extracted from rodent kidneys (n = 68) and chicken faeces (n = 52). Two rodent pest species, namely Rattus rattus and Rattus tanezumi, were identified by analysis of CO1 gene sequences. Detection of C. burnetii was carried out using polymerase chain reaction assays targeting 23S rRNA, 16S rRNA and IS111 markers. RESULTS: C. burnetii was detected in 16.2%, 8.8% and 25% of R. rattus, R. tanezumi and chickens, respectively. CONCLUSIONS: The findings in this study demonstrate that rodents and chickens are harbouring C. burnetii at sampled poultry farms. There should be frequent screening for C. burnetii in poultry operations. The likelihood of future transmission between rodents and chickens, including humans, also needs to be investigated.

Culture-Negative Endocarditis in an Immunocompromised Patient: A Case of Suspected Bartonella and Coxiella Co-Infection.

We present the case of a 23-year-old man with a previous deceased-donor renal transplant maintained on tacrolimus and prednisone who developed culture-negative endocarditis (CNE) of the mitral and aortic valves. He was suspected of being co-infected with Bartonella henselae and Coxiella burnetii, confirmed with serology testing. He was successfully managed with appropriate antibiotics and dual valve replacement.

Antimicrobial resistance gene lack in tick-borne pathogenic bacteria.

Tick-borne infections, including those of bacterial origin, are significant public health issues. Antimicrobial resistance (AMR), which is one of the most pressing health challenges of our time, is driven by specific genetic determinants, primarily by the antimicrobial resistance genes (ARGs) of bacteria. In our work, we investigated the occurrence of ARGs in the genomes of tick-borne bacterial species that can cause human infections. For this purpose, we processed short/long reads of 1550 bacterial isolates of the genera Anaplasma (n = 20), Bartonella (n = 131), Borrelia (n = 311), Coxiella (n = 73), Ehrlichia (n = 13), Francisella (n = 959) and Rickettsia (n = 43) generated by second/third generation sequencing that have been freely accessible at the NCBI SRA repository. From Francisella tularensis, 98.9% of the samples contained the FTU-1 beta-lactamase gene. However, it is part of the F. tularensis representative genome as well. Furthermore, 16.3% of them contained additional ARGs. Only 2.2% of isolates from other genera (Bartonella: 2, Coxiella: 8, Ehrlichia: 1, Rickettsia: 2) contained any ARG. We found that the odds of ARG occurrence in Coxiella samples were significantly higher in isolates related to farm animals than from other sources. Our results describe a surprising lack of ARGs in these bacteria and suggest that Coxiella species in farm animal settings could play a role in the spread of AMR.

What do we know about the microbiome of I. ricinus?

I. ricinus is an obligate hematophagous parasitic arthropod that is responsible for the transmission of a wide range of zoonotic pathogens including spirochetes of the genus Borrelia, Rickettsia spp., C. burnetii, Anaplasma phagocytophilum and Francisella tularensis, which are part the tick´s microbiome. Most of the studies focus on "pathogens" and only very few elucidate the role of "non-pathogenic" symbiotic microorganisms in I. ricinus. While most of the members of the microbiome are leading an intracellular lifestyle, they are able to complement tick´s nutrition and stress response having a great impact on tick´s survival and transmission of pathogens. The composition of the tick´s microbiome is not consistent and can be tied to the environment, tick species, developmental stage, or specific organ or tissue. Ovarian tissue harbors a stable microbiome consisting mainly but not exclusively of endosymbiotic bacteria, while the microbiome of the digestive system is rather unstable, and together with salivary glands, is mostly comprised of pathogens. The most prevalent endosymbionts found in ticks are Rickettsia spp., Ricketsiella spp., Coxiella-like and Francisella-like endosymbionts, Spiroplasma spp. and Candidatus Midichloria spp. Since microorganisms can modify ticks' behavior, such as mobility, feeding or saliva production, which results in increased survival rates, we aimed to elucidate the potential, tight relationship, and interaction between bacteria of the I. ricinus microbiome. Here we show that endosymbionts including Coxiella-like spp., can provide I. ricinus with different types of vitamin B (B2, B6, B7, B9) essential for eukaryotic organisms. Furthermore, we hypothesize that survival of Wolbachia spp., or the bacterial pathogen A. phagocytophilum can be supported by the tick itself since coinfection with symbiotic Spiroplasma ixodetis provides I. ricinus with complete metabolic pathway of folate biosynthesis necessary for DNA synthesis and cell division. Manipulation of tick´s endosymbiotic microbiome could present a perspective way of I. ricinus control and regulation of spread of emerging bacterial pathogens.

Molecular detection of Coxiella-like endosymbionts in Rhipicephalus microplus from north India.

Apart from the tick-borne pathogens affecting human and animal health, ticks also harbor various non-pathogenic endosymbionts with dynamic ecological interactions. These endosymbionts are unexplored from the Indian ticks; hence this pilot study was conducted. Seventy-nine ticks were collected from Nainital district of Uttarakhand state of north India and were identified as Rhipicephalus microplus morphologically and by molecular analysis. PCR and sequence analysis were carried out to detect the presence of Rickettsia-like, Coxiella-like and Francisella-like endosymbionts in these ticks. Based on the partial 16S rRNA gene sequence, Coxiella-like endosymbiont (CLE) was detected in the adult and other life-cycle stages of ticks with 96.6-97.7% nucleotide sequence identity with the published CLE sequences from GenBank. The phylogenetic analysis revealed that the CLE from R. microplus were clustered with the CLE from other Rhipicephalus species. All these CLE formed distinct clades from the pathogenic Coxiella burnetii. None of the tick samples was found positive for Rickettsia-like and Francisella-like endosymbionts in the present study. We also demonstrated the vertical transmission of CLE from surface sterilized and laboratory reared fully engorged adult females to the eggs and the larvae. However, large scale studies are to be conducted to detect various endosymbionts and endosymbiont-tick associations in the Indian tick species and to explore these associations for tick and tick-borne disease control.

Molecular detection of Coxiella-like endosymbionts and absence of Coxiella burnetii in Amblyomma mixtum from Veracruz, Mexico.

Ticks are obligate ectoparasites associated with a wide range of vertebrate hosts, including domestic animals. Moreover, ticks are capable of transmitting many pathogens such as Coxiella. To date, Coxiella burnetii, the etiological agent of coxiellosis or Q fever, is the only valid species of the genera. Nevertheless, a wide range of agents denominated Coxiella-like have been detected in recent studies, mainly associated with ticks. The pathogenicity of these Coxiella-like agents is controversial as some of them can infect both birds and humans. In Mexico, knowledge about Q fever is scarce and limited to historical serological records, and there is an overall lack of molecular proof of any agent of the genus Coxiella circulating in the country. Therefore, the aim of this study was to detect the presence of Coxiella in ticks associated with cattle in all 10 regions of Veracruz, Mexico. To accomplish this objective, first, we identified ticks collected from cattle and horses in Veracruz. Then, for Coxiella detection, DNA extraction from ticks and PCR amplification of the 16S-rDNA of Coxiella was performed. Finally, we performed a phylogenetic reconstruction to determine the Coxiella lineages detected. From the 10 regions sampled we collected 888 ticks grouped in 180 pools, and only five Amblyomma mixtum from the locality of Castán, and one from Los Angeles from Tuxpan were found positive, which represents a frequency of 20% for each locality. This study represents the first attempt at molecular detection of Coxiella in ticks associated with cattle in the state of Veracruz, the major livestock producer in the country. The findings of the present study are relevant as they establish a precedent regarding the circulation of Coxiella-like agents, as well as the absence in three municipalities of the state of Veracruz of C. burnetii, an abortive agent of livestock importance.

The diversity and evolutionary relationships of ticks and tick-borne bacteria collected in China.

BACKGROUND: Ticks (order Ixodida) are ectoparasites, vectors and reservoirs of many infectious agents affecting humans and domestic animals. However, the lack of information on tick genomic diversity leaves significant gaps in the understanding of the evolution of ticks and associated bacteria. RESULTS: We collected > 20,000 contemporary and historical (up to 60 years of preservation) tick samples representing a wide range of tick biodiversity across diverse geographic regions in China. Metagenomic sequencing was performed on individual ticks to obtain the complete or near-complete mitochondrial (mt) genome sequences from 46 tick species, among which mitochondrial genomes of 23 species were recovered for the first time. These new mt genomes data greatly expanded the diversity of many tick groups and revealed five cryptic species. Utilizing the same metagenomic sequence data we identified divergent and abundant bacteria in Haemaphysalis, Ixodes, Dermacentor and Carios ticks, including nine species of pathogenetic bacteria and potentially new species within the genus Borrelia. We also used these data to explore the evolutionary relationship between ticks and their associated bacteria, revealing a pattern of long-term co-divergence relationship between ticks and Rickettsia and Coxiella bacteria. CONCLUSIONS: In sum, our study provides important new information on the genetic diversity of ticks based on an analysis of mitochondrial DNA as well as on the prevalence of tick-borne pathogens in China. It also sheds new light on the long-term evolutionary and ecological relationships between ticks and their associated bacteria.

Tropical and Temperate Lineages of Rhipicephalus sanguineus s.l. Ticks (Acari: Ixodidae) Host Different Strains of Coxiella-like Endosymbionts.

Nonpathogenic bacteria likely play important roles in the biology and vector competence of ticks and other arthropods. Coxiella, a gram-negative gammaproteobacterium, is one of the most commonly reported maternally inherited endosymbionts in ticks and has been associated with over 40 tick species. Species-specific Coxiella-like endosymbionts (CLEs) have been reported in the brown dog tick, Rhipicephalus sanguineus sensu lato (Acari: Ixodidae), throughout the world, while recent research suggests low Coxiella diversity among tick species. We investigated CLE diversity among R. sanguineus s.l. ticks across Arizona. We detected 37 recurrent sequence variants (SVs) of the symbiont, indicating greater diversity in these symbiotic bacteria than previously reported. However, two SVs accounted for the vast majority of 16S rRNA amplicon reads. These two dominant CLEs were both closely related to Candidatus C. mudrowiae, an identified symbiont of Rhipicephalus turanicus ticks. One strain strongly associated with the tropical lineage of R. sanguineus s.l. while the other was found almost exclusively in the temperate lineage, supporting the conclusion that CLEs are primarily vertically transmitted. However, occasional mismatches between tick lineage and symbiont SV indicate that horizontal symbiont transfer may occur, perhaps via cofeeding of ticks from different lineages on the same dog. This study advances our understanding of CLE diversity in Rh. sanguineus s.l.

Spotted fever group Rickettsia, Anaplasma and Coxiella-like endosymbiont in Haemaphysalis ticks from mammals in Thailand.

Ticks are ectoparasites of vertebrates and vectors of various pathogenic microorganisms. In this study, the presence of bacteria and protozoa was evaluated by PCR and DNA sequencing in 233 mammal ticks collected from 8 provinces in Thailand. Sequence and phylogenetic analyses of partial rickettsial ompA, ompB, sca4 and partial Coxiella 16S rRNA, GroEL, rpoB genes clearly revealed, for the first time, a co-infection of SFG Rickettsia belonging to R. massiliae subgroup and Coxiella-like endosymbiont (CLE), Cox-hein, in a male of Haemaphysalis heinrichi tick infesting Burmese ferret-badger in Loei province. Moreover, a male of H. hystricis tick infesting the same host was infected with another CLE, Cox-hys. Based on the 16S rRNA gene sequence, Anaplasma sp., closely related to Anaplasma bovis was also detected in a male of H. heinrichi infesting the same Burmese ferret-badger. In addition, the third CLE, Cox-asia, found in H. asiatica collected from Asian palm civet in Chiang Rai province, was different from both Cox-hein and Cox-hys. This study provided important data and broadened our knowledge on tick-borne pathogens and endosymbionts in Thailand and Southeast Asia.

Growth dynamics and tissue localization of a Coxiella-like endosymbiont in the tick Haemaphysalis longicornis.

A Coxiella-like endosymbiont (Coxiella-LE hereinafter) stably infects and influences Haemaphysalis longicornis development, indicating a mutualistic relationship of Coxiella-LE and ticks. To further elucidate the patterns of growth dynamics and tissue localization of Coxiella-LE in H. longicornis, 16S rRNA high-throughput sequencing, quantitative PCR (qPCR), and fluorescence in situ hybridization (FISH) were used in this study. The density of Coxiella-LE varied among different tick life stages, and fed female ticks had the highest density, followed by unfed female and unfed larval ticks. In the four organs that were dissected from fed female ticks, the ovary carried the highest density of Coxiella-LE, which was significantly different from salivary glands, midgut and Malpighian tubules. The high abundance of Coxiella-LE in fed female ticks and in the ovaries of fed female ticks in the bacterial microbiota analyses further confirmed that Coxiella-LE rapidly proliferates in the ovary after blood feeding. The ovaries continued to develop after engorgement and oviposition began on day 5, with a significant decrease in the density of Coxiella-LE in the ovaries occurring on day 7. FISH results indicated that Coxiella-LE is mainly colonized in the cytoplasm of the oocyte and proliferates with oogenesis. Coxiella-LE was expelled from the body with the mature oocyte, ensuring its vertical transmission. In the Malpighian tubules at different days after engorgement, the white flocculent materials were increasing, and the density of Coxiella-LE raised significantly on day 7. Unlike the localization pattern in the ovary, Coxiella-LE was initially distributed in a mass and continually increased during the development of Malpighian tubules until it filled the Malpighian tubules. These findings provide new insights on the growth dynamics and tissue localization of Coxiella-LE in ticks and are useful for further investigation on the interactions of symbiont and ticks .

Coxiella-like bacteria in Haemaphysalis wellingtoni ticks associated with Great Hornbill, Buceros bicornis.

Birds are known to be the most mobile hosts and are therefore considered to be hosts with potential to contribute to the long-distance spread and transmission of tick-borne pathogens. In the present study, ticks were collected from a hornbill nest at Chaiyaphum Province, Thailand. They were screened for the presence of Coxiella bacteria using conventional PCR. The evolutionary relationships of positive Coxiella-like bacteria (CLB) were analysed based on the gene sequences of 16S rRNA, groEL and rpoB. Among all 22 tested ticks, CLB infections were found in 2 Haemaphysalis wellingtoni individuals. In a phylogenetic analysis, the Coxiella 16S rRNA gene detected in this study formed a separate clade from sequences found in ticks of the same genus. In contrast, the phylogenetic relationships based on groEL and rpoB revealed that these two genes from H. wellingtoni ticks grouped with CLB from the same tick genus (Haemaphysalis). This study is the first to report the presence of CLB in H. wellingtoni ticks associated with the Great Hornbill, Buceros bicornis in Thailand. Three genes of CLB studied herein were grouped separately with Coxiella burnetii (pathogenic strain). The effects of CLB in the ticks and Buceros bicornis require further investigation.

Ehrlichia, Coxiella and Bartonella infections in rodents from Guizhou Province, Southwest China.

Rodents are generally recognized to be the reservoir hosts of a great many zoonotic pathogens. In some areas of China, rodent-borne pathogens, as well as the role of rodents in the natural cycle of these pathogens, are still poorly investigated. To increase our knowledge on the distribution and epidemiology of rodent-borne bacterial pathogens, 81 rodent liver samples were collected in three locations of Guizhou province located in Southwest China, and screened for the presence of Ehrlichia, Coxiella, and Bartonella in them. A putative novel Ehrlichia species was identified in 5 Berylmys bowersi samples (100%, 5/5). Its 16S rRNA, gltA, and groEL genes have highest 99.84%, 89.11%, and 98.02% identities to those from known Ehrlichia species, and form distinct clades in the phylogenetic trees. Herein we name it "Candidatus Ehrlichia zunyiensis". Bartonella was tested positive in 8 A. agrarius (striped field mouse), 2 A. chevrieri (Chevrier's field mouse), 1 R. norvegicus (Norway rat), 1 N. confucianus, and 1 N. lotipes, with a total positive rate of 16.05% (13/81). Sequence analysis indicated high genetic diversity in these Bartonella strains. Unexpectedly, two Coxiella strains were identified from the rodents (1 Niviventer confucianus and 1 Mus pahari). Genetic and phylogenetic analysis indicated that both of them are closely related to the Coxiella endosymbiont of ticks. This result supported previous conjectures that vertebrate hosts such as rodents may play a role in the preservation and transmission of Coxiella endosymbiont of ticks.

Characterization of recombinant Ybgf protein for the detection of Coxiella antibodies in ruminants.

Q fever remains a One Health problem, posing a zoonotic threat and causing significant economic losses to the livestock industry. The advancement of detection tools is critical to the effective control of infection. In humans, laboratory investigations depend largely on the immunofluorescence assay, considered the gold standard. In contrast, serologic tools routinely used for veterinary screening have several gaps, resulting in interpretations that are frequently misleading. We investigated the potential application of recombinant Ybgf antigen (r-Ybgf), a periplasmic protein described as one of the most immunodominant antigens in humans, in an indirect ELISA. Following successful expression in the prokaryotic system and the preliminary evaluation of immunoreactivity in western blot, we used r-Ybgf to develop an in-house ELISA using serum samples from sheep, goats, and cattle, which were tested in parallel with an Idexx ELISA kit. The results obtained with the 2 tests were compared, and r-Ybgf performed favorably, with 81.8% sensitivity and 90.1% specificity and substantial agreement, as revealed by receiver operating characteristic analysis. Moreover, we evaluated the serologic response against phase I (PhI) and phase II (PhII) antigens, and r-Ybgf antigen induced by vaccination, using phase-specific ELISAs. The dynamics of antibody response showed a significant increase in reactivity against PhI and PhII, but not against r-Ybgf, antigens. This property may be very useful given the absence of a protocol for the differentiation of infected from vaccinated animals.

Recently Evolved Francisella-Like Endosymbiont Outcompetes an Ancient and Evolutionarily Associated Coxiella-Like Endosymbiont in the Lone Star Tick (Amblyomma americanum) Linked to the Alpha-Gal Syndrome.

Background: Ticks are hematophagous arthropods that transmit various bacterial, viral, and protozoan pathogens of public health significance. The lone star tick (Amblyomma americanum) is an aggressive human-biting tick that transmits bacterial and viral pathogens, and its bites are suspected of eliciting the alpha-gal syndrome, a newly emerged delayed hypersensitivity following consumption of red meat in the United States. While ongoing studies have attempted to investigate the contribution of different tick-inherent factors to the induction of alpha-gal syndrome, an otherwise understudied aspect is the contribution of the tick microbiome and specifically obligate endosymbionts to the establishment of the alpha-gal syndrome in humans. Materials and Methods: Here we utilized a high-throughput metagenomic sequencing approach to cataloging the entire microbial communities residing within different developmental stages and tissues of unfed and blood-fed ticks from laboratory-maintained ticks and three new geographical locations in the United States. The Quantitative Insights Into Microbial Ecology (QIIME2) pipeline was used to perform data analysis and taxonomic classification. Moreover, using a SparCC (Sparse Correlations for Compositional data) network construction model, we investigated potential interactions between members of the microbial communities from laboratory-maintained and field-collected ticks. Results: Overall, Francisellaceae was the most dominant bacteria identified in the microbiome of both laboratory-raised and field-collected Am. americanum across all tissues and developmental stages. Likewise, microbial diversity was seen to be significantly higher in field-collected ticks compared with laboratory-maintained ticks as seen with a higher number of both Operational Taxonomic Units and measures of species richness. Several potential positive and negative correlations were identified from our network analysis. We observed a strong positive correlation between Francisellaceae, Rickettsiaceae, and Midichloriaceae in both developmental stages and tissues from laboratory-maintained ticks, whereas ovarian tissues had a strong positive correlation of bacteria in the family Xanthobacteraceae and Rhizobiaceae. A negative interaction was observed between Coxiellaceae and Francisellaceae in Illinois, and all the bacteria detected from ticks from Delaware were negatively correlated. Conclusion: This study is the first to catalog the microbiome of Am. americanum throughout its developmental stages and different tissue niches and report the potential replacement of Coxiellaceae by Francisellaceae across developmental stages and tissues tested except in ovarian tissues. These unique and significant findings advance our knowledge and open a new avenue of research to further understand the role of tick microbiome in tick-borne diseases and develop a holistic strategy to control alpha-gal syndrome.