Diversity of bartonellae in mites (Acari: Mesostigmata: Macronyssidae and Spinturnicidae) of boreal forest bats: Association of host specificity of mites and habitat selection of hosts with vector potential.

Research into various bacterial pathogens that can be transmitted between different animals and may have zoonotic potential has led to the discovery of different strains of Bartonella sp. in bats and their associated ectoparasites. Despite their enormous species diversity, only a few studies have focussed on the detection of bacterial pathogens in insectivorous bats of boreal forests and their associated Macronyssidae and Spinturnicidae mites. We collected and molecularly analysed mite samples from forest-dwelling bat species distributed all along the boreal belt of the Palearctic, from Central Europe to Far East. Ectoparasitic mites were pooled for DNA extraction and DNA amplification polymerase chain reaction (PCRs) were conducted to detect the presence of various bacterial (Anaplasmataceae, Bartonella sp., Rickettsia sp., Mycoplasma sp.) and protozoal (Hepatozoon sp.) pathogens. Bartonella sp. DNA was detected in four different mite species (Macronyssidae: Steatonyssus periblepharus and Spinturnicidae: Spinturnix acuminata, Sp. myoti and Sp. mystacinus), with different prevalences of the targeted gene (gltA, 16-23S ribosomal RNA intergenic spacer and ftsZ). Larger pools (>5 samples pooled) were more likely to harbour Bartonella sp. DNA, than smaller ones. In addition, cave-dwelling bat hosts and host generalist mite species are more associated with Bartonella spp. presence. Spinturnicidae mites may transmit several distinct Bartonella strains, which cluster phylogenetically close to Bartonella species known to cause diseases in humans and livestock. Mites with ubiquitous presence may facilitate the long-term maintenance (and even local recurrence) of Bartonella-infestations inside local bat populations, thus acting as continuous reservoirs for Bartonella spp in bats.

Rickettsial pathogens drive microbiota assembly in Hyalomma marginatum and Rhipicephalus bursa ticks.

Most tick-borne pathogens (TBPs) are secondarily acquired by ticks during feeding on infected hosts, which imposes 'priority effect' constraints, as arrival order influences the establishment of new species in a microbial community. Here we tested whether once acquired, TBPs contribute to bacterial microbiota functioning by increasing community stability. For this, we used Hyalomma marginatum and Rhipicephalus bursa ticks collected from cattle in different locations of Corsica and combined 16S rRNA amplicon sequencing and co-occurrence network analysis, with high-throughput pathogen detection, and in silico removal of nodes to test for impact of rickettsial pathogens on network properties. Despite its low centrality, Rickettsia showed preferential connections in the networks, notably with a keystone taxon in H. marginatum, suggesting facilitation of Rickettsia colonisation by the keystone taxon. In addition, conserved patterns of community assembly in both tick species were affected by Rickettsia removal, suggesting that privileged connections of Rickettsia in the networks make this taxon a driver of community assembly. However, Rickettsia removal had minor impact on the conserved 'core bacterial microbiota' of H. marginatum and R. bursa. Interestingly, networks of the two tick species with Rickettsia have similar node centrality distribution, a property that is lost after Rickettsia removal, suggesting that this taxon drives specific hierarchical interactions between bacterial microbes in the microbiota. The study indicates that tick-borne Rickettsia play a significant role in the tick bacterial microbiota, despite their low centrality. These bacteria are influential and contribute to the conservation of the 'core bacterial microbiota' while also promoting community stability.

First molecular characterization of Dirofilaria Immitis in Cuba.

BACKGROUND: Dirofilarioses are widespread diseases caused by mosquito-borne nematodes of the family Onchocercidae, genus Dirofilaria. The major etiologic agent of canine dirofilariosis in the American continent is the zoonotic parasite Dirofilaria immitis. Existing reports of filarioid nematodes in Cuba are based solely on morphological and immunological analysis which do not allow unambiguous identification and/or direct detection of causal agents. RESULTS: Here we present the molecular characterization of filarioid nematodes found in a dog in Cuba. Based on the molecular and phylogenetic analysis of the 5.8S-ITS2-28S region and cox1 gene fragments, the worms were unambiguously classified as D. immitis. Sequence analysis showed high identity of the gene fragments in this study with others previously obtained from D. immitis found in dogs, wolfs and jackals but also from mosquito vectors of D. immitis. CONCLUSIONS: Further studies are guarantee to better understand the epidemiological impact of canine dirofilariosis in Cuba as well as the competence of different species of culicid mosquitoes as vectors of Dirofilaria in the country.

Mediterranean spotted fever-like illness caused by Rickettsia sibirica mongolitimonae, North Macedonia, June 2022.

Mediterranean spotted fever-like illness (MSF-like illness) is a tick-borne disease caused by Rickettsia sibirica mongolitimonae first reported in France more than 25 years ago. Until today, more than 50 cases of MSF-like illness have been reported in different regions of Europe and Africa, highlighting variable clinical manifestation. Here we report a case of MSF-like illness following a bite from a Hyalomma tick in the Skopje region of North Macedonia.

Metabolic markers of protein maldigestion after a 15N test meal in minipigs with pancreatic exocrine insufficiency.

The effect of pancreatic exocrine insufficiency (PEI) on protein malabsorption is little documented, partly due to methodological barriers. We aimed to validate biomarkers of protein malabsorption using a 15N test meal in a minipig model of PEI. Six pancreatic duct-ligated minipigs were used as a model of PEI and four nonoperated animals as a control. All animals were equipped with an ileocecal reentrant cannula. Minipigs were given a test meal containing [15N]casein. The PEI animals repeated the test three times, in the absence of any pancreatic enzymes, or after pancreatic substitution at two levels [ A or B: 7,500 or 75,000 (lipase) and 388 or 3881 (protease) FIP U]. Ileal chyme, urine, and blood were collected postprandially. Nitrogen and 15N were measured in digestive and metabolic pools. We obtained a gradient of ileal protein digestibility from 29 ± 11% in PEI to 89 ± 6% in the controls and a dose- dependent response of enzymes. Insulin and gastric inhibitory polypeptide secretions were decreased by PEI, an effect that was counteracted with the enzymes at level B. The total recovery of 15N in urinary urea and plasma proteins was 14 ± 5.1% in the control group and decreased to 5.5 ± 2.1% by PEI. It was dose dependently restored by the treatment. Both 15N recovery in plasma and urine were correlated to protein digestibility. We confirm that the 15N transfer in those pools is a sensitive marker of protein malabsorption. Nevertheless, an optimization of the test meal conditions would be necessary in the view of implementing a clinical test. NEW & NOTEWORTHY We designed an intervention study to create a gradient of ileal protein digestibility in minipigs with pancreatic exocrine insufficiency and to validate reliable metabolic markers using a 15N oral meal test. 15N recovery in plasma proteins and to a higher extent in urine was sensitive to protein malabsorption. This test is minimally invasive and could be used to reveal protein malabsorption in patients.

High True Ileal Digestibility but Not Postprandial Utilization of Nitrogen from Bovine Meat Protein in Humans Is Moderately Decreased by High-Temperature, Long-Duration Cooking.

BACKGROUND: Meat protein digestibility can be impaired because of indigestible protein aggregates that form during cooking. When the aggregates are subsequently fermented by the microbiota, they can generate potentially harmful compounds for the colonic mucosa. OBJECTIVE: This study evaluated the quantity of bovine meat protein escaping digestion in the human small intestine and the metabolic fate of exogenous nitrogen, depending on cooking processes. METHODS: Sixteen volunteers (5 women and 11 men; aged 28 ± 8 y) were equipped with a double lumen intestinal tube positioned at the ileal level. They received a test meal exclusively composed of 120 g of intrinsically (15)N-labeled bovine meat, cooked either at 55°C for 5 min (n = 8) or at 90°C for 30 min (n = 8). Ileal effluents and blood and urine samples were collected over an 8-h period after the meal ingestion, and (15)N enrichments were measured to assess the digestibility of meat proteins and the transfer of dietary nitrogen into the metabolic pools. RESULTS: Proteins tended to be less digestible for the meat cooked at 90°C for 30 min than at 55°C for 5 min (90.1% ± 2.1% vs. 94.1% ± 0.7% of ingested N; P = 0.08). However, the particle number and size in ileal digesta did not differ between groups. The appearance of variable amounts of intact fibers was observed by microscopy. The kinetics of (15)N appearance in plasma proteins, amino acids, and urea were similar between groups. The amount of exogenous nitrogen lost through deamination did not differ between groups (21.2% ± 0.8% of ingested N). CONCLUSIONS: Cooking bovine meat at a high temperature for a long time can moderately decrease protein digestibility compared with cooking at a lower temperature for a short time and does not affect postprandial exogenous protein metabolism in young adults. The study was registered at www.clinicaltrials.gov as NCT01685307.

Tick paralysis induced by Ixodes gibbosus: enigmatic cases in domestic mammals from Cyprus.

Tick paralysis is a potentially fatal condition caused by toxins produced and secreted by tick salivary glands. This survey presents clinical and epidemiological observations of tick paralysis cases in domestic animals in Cyprus. Local veterinarians report typical tick paralysis cases occurring in goats, sheep, dogs, and cats. The animals suffering from paralysis are free from other neurological diseases, have blood and biochemical parameters within normal ranges, and recover fast by simply removing the ticks found predominantly on the head and around the neck. Tick paralysis cases occur in a specific geographic area of Cyprus (Akamas peninsula), from September through March, but not every year. Instead, the phenomenon has 2 periodic cycles of occurrence, a 3- and a 7-year cycle. The 2 cycles are differentiated by severity based on the number of affected animals and the resulting losses. As described for other tick-borne diseases, these cyclic patterns may be attributed to external factors, self-oscillations of the disease system, or the combined action of these mechanisms. Ticks collected from a recent paralysis case in a goat were morphologically and molecularly identified as Ixodes gibbosus. Efforts should be made to characterize the specific toxins involved in tick paralysis and to develop a vaccine, which could prevent significant losses of small ruminants, especially in free-ranging farming systems, a prevalent management approach observed in Cyprus and various regions worldwide.

Dynamic nesting of Anaplasma marginale in the microbial communities of Rhipicephalus microplus.

Interactions within the tick microbiome involving symbionts, commensals, and tick-borne pathogens (TBPs) play a pivotal role in disease ecology. This study explored temporal changes in the microbiome of Rhipicephalus microplus, an important cattle tick vector, focusing on its interaction with Anaplasma marginale. To overcome limitations inherent in sampling methods relying on questing ticks, which may not consistently reflect pathogen presence due to variations in exposure to infected hosts in nature, our study focused on ticks fed on chronically infected cattle. This approach ensures continuous pathogen exposure, providing a more comprehensive understanding of the nesting patterns of A. marginale in the R. microplus microbiome. Using next-generation sequencing, microbiome dynamics were characterized over 2 years, revealing significant shifts in diversity, composition, and abundance. Anaplasma marginale exhibited varying associations, with its increased abundance correlating with reduced microbial diversity. Co-occurrence networks demonstrated Anaplasma's evolving role, transitioning from diverse connections to keystone taxa status. An integrative approach involving in silico node removal unveils the impact of Anaplasma on network stability, highlighting its role in conferring robustness to the microbial community. This study provides insights into the intricate interplay between the tick microbiome and A. marginale, shedding light on potential avenues for controlling bovine anaplasmosis through microbiome manipulation.

Tick-borne diseases at the crossroads of the Middle East and central Europe.

OBJECTIVES: The Balkan Peninsula, acting as a crossroad between central Europe and the Middle East, presents diverse ecosystems supporting various tick species capable of transmitting TBDs. This study focuses on Serbia and North Macedonia, both endemic for TBDs, aiming to investigate human-biting ticks' prevalence, TBD prevalence, and major TBPs in blood samples. PATIENTS AND METHODS: This prospective observational study was conducted in 2022 at two medical centers, involving 45 patients from Novi Sad, Serbia, and 17 patients from Skopje, North Macedonia. All participants had either a tick still attached or had had one removed within the preceding 48 h. The study consisted in clinical evaluations of patients and testing of patient samples and ticks for tick-borne pathogens using a High-Throughput pathogen detection system based on microfluidic real-time PCR. In addition, the study assessed the genetic diversity of the identified pathogens. RESULTS: Ixodes ricinus was the most prevalent tick species, with varying infestation rates across various body parts. Tick species and feeding times differed between Novi Sad and Skopje. TBPs were prevalent, with Rickettsia spp. dominant in Skopje and a mix including Rickettsia aeschlimannii, Rickettsia monacensis, Anaplasma phagocytophilum, and Borrelia afzelii in Novi Sad. Subclinical bacteremia occurred in 8.06% of cases, mostly involving Anaplasma spp. Clinical manifestations, primarily local hypersensitivity reactions, were observed in six patients. Phylogenetic analysis confirmed R. aeschlimannii and R. monacensis identity, highlighting genetic differences in gltA gene sequences. CONCLUSIONS: This study sheds light on the prevalence and diversity of TBPs in tick-infested individuals from Serbia and North Macedonia, contributing valuable insights into the epidemiology of TBDs in the Balkan region.

One health approach to study human health risks associated with Dermanyssus gallinae mites.

Despite the significant health risks associated with Dermanyssus gallinae infestations in humans, they are often overlooked. This study investigated a household case of D. gallinae infestation and explored the resulting clinical manifestations and risk of infection in family members. Microfluidic PCR was employed for high-throughput screening of pathogens in collected mites and blood samples from both chickens and family members. Morphological and molecular examinations confirmed the identity of the mites as D. gallinae sensu stricto (s.s.), with evidence indicating recent blood feeding. Results indicated that the mites exclusively harbored various pathogens, including Bartonella spp., Ehrlichia spp., Apicomplexa, and Theileria spp. Blood samples from family members and poultry tested negative for these pathogens, suggesting a potential reservoir role for D. gallinae. The study further identified haplotypes of D. gallinae, classifying them into D. gallinae s.s., cosmopolitan haplogroup A. Serological analysis revealed elevated IgE seroreactivity against mite proteins in the family member with bite lesions. Antibodies against Bartonella spp. were detected in this individual, indicating exposure to the pathogen. In summary, this study sheds light on the clinical manifestations, pathogen detection, and genetic characterization of D. gallinae infestations, underscoring the necessity of adopting comprehensive approaches to manage such infestations effectively.

Differential nested patterns of Anaplasma marginale and Coxiella-like endosymbiont across Rhipicephalus microplus ontogeny.

Understanding the intricate ecological interactions within the microbiome of arthropod vectors is crucial for elucidating disease transmission dynamics and developing effective control strategies. In this study, we investigated the ecological roles of Coxiella-like endosymbiont (CLE) and Anaplasma marginale across larval, nymphal, and adult stages of Rhipicephalus microplus. We hypothesized that CLE would show a stable, nested pattern reflecting co-evolution with the tick host, while A. marginale would exhibit a more dynamic, non-nested pattern influenced by environmental factors and host immune responses. Our findings revealed a stable, nested pattern characteristic of co-evolutionary mutualism for CLE, occurring in all developmental stages of the tick. Conversely, A. marginale exhibited variable occurrence but exerted significant influence on microbial community structure, challenging our initial hypotheses of its non-nested dynamics. Furthermore, in silico removal of both microbes from the co-occurrence networks altered network topology, underscoring their central roles in the R. microplus microbiome. Notably, competitive interactions between CLE and A. marginale were observed in nymphal network, potentially reflecting the impact of CLE on the pathogen transstadial-transmission. These findings shed light on the complex ecological dynamics within tick microbiomes and have implications for disease management strategies.

Microfluidic PCR and network analysis reveals complex tick-borne pathogen interactions in the tropics.

BACKGROUND: Ixodid ticks, particularly Rhipicephalus sanguineus s.l., are important vectors of various disease-causing agents in dogs and humans in Cuba. However, our understading of interactions among tick-borne pathogens (TBPs) in infected dogs or the vector R. sanguineus s.l. remains limited. This study integrates microfluidic-based high-throughput real-time PCR data, Yule's Q statistic, and network analysis to elucidate pathogen-pathogen interactions in dogs and ticks in tropical western Cuba. METHODS: A cross-sectional study involving 46 client-owned dogs was conducted. Blood samples were collected from these dogs, and ticks infesting the same dogs were morphologically and molecularly identified. Nucleic acids were extracted from both canine blood and tick samples. Microfluidic-based high-throughput real-time PCR was employed to detect 25 bacterial species, 10 parasite species, 6 bacterial genera, and 4 parasite taxa, as well as to confirm the identity of the collected ticks. Validation was performed through end-point PCR assays and DNA sequencing analysis. Yule's Q statistic and network analysis were used to analyse the associations between different TBP species based on binary presence-absence data. RESULTS: The study revealed a high prevalence of TBPs in both dogs and R. sanguineus s.l., the only tick species found on the dogs. Hepatozoon canis and Ehrlichia canis were among the most common pathogens detected. Co-infections were observed, notably between E. canis and H. canis. Significant correlations were found between the presence of Anaplasma platys and H. canis in both dogs and ticks. A complex co-occurrence network among haemoparasite species was identified, highlighting potential facilitative and inhibitory roles. Notably, H. canis was found as a highly interconnected node, exhibiting significant positive associations with various taxa, including A. platys, and E. canis, suggesting facilitative interactions among these pathogens. Phylogenetic analysis showed genetic diversity in the detected TBPs. CONCLUSIONS: Overall, this research enhances our understanding of TBPs in Cuba, providing insights into their prevalence, associations, and genetic diversity, with implications for disease surveillance and management.

Tick Activity, Host Range, and Tick-Borne Pathogen Prevalence in Mountain Habitats of the Western Carpathians, Poland.

In mountainous regions, diverse ecosystems provide a habitat for numerous species of organisms. In this study, we focused on ixodid ticks and their presence in the Western Carpathians, Poland. Our objectives were to investigate the impact of environmental factors on tick occurrence and activity, the prevalence of vectored pathogens, and tick hosts, and their role as reservoir organisms for tick-borne pathogens (TBPs). To this end, we collected ticks from the vegetation and from animals (Apodemus agrarius, A. flavicollis, Capreolus capreolus, Microtus spp., Myodes glareolus, Ovis aries). In addition, we collected blood samples from rodents. The collected material underwent molecular analysis, utilizing the high-throughput microfluidic real-time PCR technique, to detect the presence of TBPs. Our findings confirmed the occurrence of only two species of ixodid ticks in the study area: the dominant Ixodes ricinus, and Dermacentor reticulatus with very limited abundance. Temperature significantly influenced tick activity, and the number of I. ricinus nymphs varied with altitude. We also observed a circadian pattern of questing activity in I. ricinus ticks. The main hosts for juvenile tick stages were M. glareolus and A. agrarius, while adult stages were frequently found on C. capreolus. I. ricinus ticks collected from the vegetation were often infected with Rickettsia helvetica (up to 35.71%), Borrelia afzelii (up to 28.57%), and Ehrlichia spp. (up to 9.52%). In contrast, juvenile stages frequently carried Bartonella spp. (up to 10.00%), Mycoplasma spp. (up to 16.67%) and R. helvetica (up to 16.67%). Moreover, we detected genetic material of Mycoplasma spp. (up to 100.00%), Ehrlichia spp. (up to 35.71%), Bartonella spp. (up to 25.00%), and Borrelia spp. (up to 6.25%) in rodent blood samples. The obtained results indicate A. agrarius and M. glareolus as reservoir animals for TBPs in the studied region.

Disparate dynamics of pathogen prevalence in Ixodes ricinus and Dermacentor reticulatus ticks occurring sympatrically in diverse habitats.

Ixodes ricinus and Dermacentor reticulatus ticks are important reservoirs and vectors of pathogens. The aim of the present study was to investigate the dynamic of the prevalence and genetic diversity of microorganisms detected in these tick species collected from two ecologically diverse biotopes undergoing disparate long-term climate condition. High-throughput real time PCR confirmed high prevalence of microorganisms detected in sympatrically occurring ticks species. D. reticulatus specimens were the most often infected with Francisella-like endosymbiont (FLE) (up to 100.0%) and Rickettsia spp. (up to 91.7%), while in case of I. ricinus the prevalence of Borreliaceae spirochetes reached up to 25.0%. Moreover, pathogens belonging to genera of Bartonella, Anaplasma, Ehrlichia and Babesia were detected in both tick species regardless the biotope. On the other hand, Neoehrlichia mikurensis was conformed only in I. ricinus in the forest biotope, while genetic material of Theileria spp. was found only in D. reticulatus collected from the meadow. Our study confirmed significant impact of biotope type on prevalence of representatives of Borreliaceae and Rickettsiaceae families. The most common co-infection detected in D. reticulatus was Rickettsia spp. + FLE, while Borreliaceae + R. helvetica was the most common in I. ricinus. Additionally, we found significant genetic diversity of R. raoultii gltA gene across studied years, however such relationship was not observed in ticks from studied biotopes. Our results suggest that ecological type of biotope undergoing disparate long-term climate conditions have an impact on prevalence of tick-borne pathogens in adult D. reticulatus and I. ricinus.

Differential detection of tick-borne pathogens in human platelets and whole blood using microfluidic PCR.

The tick-borne pathogens (TBPs) with adhesive phenotype can use platelets for dissemination and colonization of distant tissues and organs, and it has been shown that they can be found concentrated in the platelet fraction of blood. This study shows the differential presence of TBPs in samples of human platelet fraction (n = 68), whole blood samples (n = 68) and ticks collected (n = 76) from the same individuals, using an unbiased high-throughput pathogen detection microfluidic system. The clinical symptoms were characterized in enrolled patients. In patients with suspected TBP infection, serological assays were conducted to test for the presence of antibodies against specific TBPs. Tick species infesting humans were identified as Ixodes ricinus, Dermacentor reticulatus, and Haemaphysalis punctata. Eight patients developed local skin lesions at the site of the tick bite including non-specific lesions, itching sensation at the lesion site, and eschar. Most common TBPs detected in platelet fraction were Borrelia spielmanii and Rickettsia sp., followed by Borrelia afzelii and Anaplasma phagocytophilum. Multiple infections with three TBPs were detected in platelet fraction. In whole blood, most common TBPs detected were Anaplasma spp. and A. phagocytophilum, followed by Rickettsia spp. and B. afzelii. In ticks, the most common TBP detected was Rickettsia spp., followed by Borrelia spp. and Anaplasma spp. Overall, nine different pathogens with variable prevalence were identified using species-specific primers, and the most common was Rickettsia helvetica. In three patients, there were no coincidences between the TBPs detected in whole blood and tick samples. Only in one patient was detected A. phagocytophilum in both, whole blood and tick samples. These results suggest the unequal detection of TBPs in whole blood, platelet fraction and ticks collected, from the same individual. The results justify the use of both whole blood and platelet fraction for molecular diagnosis of TBPs in patients.

Detection of bacterial and protozoan pathogens in individual bats and their ectoparasites using high-throughput microfluidic real-time PCR.

Among the most studied mammals in terms of their role in the spread of various pathogens with possible zoonotic effects are bats. These are animals with a very complex lifestyle, diet, and behavior. They are able to fly long distances, thus maintaining and spreading the pathogens they may be carrying. These pathogens also include vector-borne parasites and bacteria that can be spread by ectoparasites such as ticks and bat flies. In the present study, high-throughput screening was performed and we detected three bacterial pathogens: Bartonella spp., Neoehrlichia mikurensis and Mycoplasma spp., and a protozoan parasite: Theileria spp. in paired samples from bats (blood and ectoparasites). In the samples from the bat-arthropod pairs, we were able to detect Bartonella spp. and Mycoplasma spp. which also showed a high phylogenetic diversity, demonstrating the importance of these mammals and the arthropods associated with them in maintaining the spread of pathogens. Previous studies have also reported the presence of these pathogens, with one exception, Neoehrlichia mikurensis, for which phylogenetic analysis revealed less genetic divergence. High-throughput screening can detect more bacteria and parasites at once, reduce screening costs, and improve knowledge of bats as reservoirs of vector-borne pathogens. IMPORTANCE The increasing number of zoonotic pathogens is evident through extensive studies and expanded animal research. Bats, known for their role as reservoirs for various viruses, continue to be significant. However, new findings highlight the emergence of Bartonella spp., such as the human-infecting B. mayotimonensis from bats. Other pathogens like N. mikurensis, Mycoplasma spp., and Theileria spp. found in bat blood and ectoparasites raise concerns, as their impact remains uncertain. These discoveries underscore the urgency for heightened vigilance and proactive measures to understand and monitor zoonotic pathogens. By deepening our knowledge and collaboration, we can mitigate these risks, safeguarding human and animal well-being.

Microbiota perturbation by anti-microbiota vaccine reduces the colonization of Borrelia afzelii in Ixodes ricinus.

BACKGROUND: Ticks can transmit a broad variety of pathogens of medical importance, including Borrelia afzelii, the causative agent of Lyme borreliosis in Europe. Tick microbiota is an important factor modulating, not only vector physiology, but also the vector competence. Anti-microbiota vaccines targeting keystone taxa of tick microbiota can alter tick feeding and modulate the taxonomic and functional profiles of bacterial communities in the vector. However, the impact of anti-microbiota vaccine on tick-borne pathogen development within the vector has not been tested. RESULTS: Here, we characterized the Ixodes ricinus microbiota modulation in response to B. afzelii infection and found that the pathogen induces changes in the microbiota composition, its beta diversity and structure of bacterial community assembly. Tick microbiota perturbation by anti-microbiota antibodies or addition of novel commensal bacteria into tick midguts causes departures from the B. afzelii-induced modulation of tick microbiota which resulted in a lower load of the pathogen in I. ricinus. Co-occurrence networks allowed the identification of emergent properties of the bacterial communities which better defined the Borrelia infection-refractory states of the tick microbiota. CONCLUSIONS: These findings suggest that Borrelia is highly sensitive to tick microbiota perturbations and that departure from the modulation induced by the pathogen in the vector microbiota pose a high cost to the spirochete. Network analysis emerges as a suitable tool to identify emergent properties of the vector microbiota associated with infection-refractory states. Anti-microbiota vaccines can be used as a tool for microbiota perturbation and control of important vector-borne pathogens. Video Abstract.

Unravelling the Diversity of Microorganisms in Ticks from Australian Wildlife.

Ticks and tick-borne pathogens pose a significant threat to the health and welfare of humans and animals. Our knowledge about pathogens carried by ticks of Australian wildlife is limited. This study aimed to characterise ticks and tick-borne microorganisms from a range of wildlife species across six sites in Victoria, Australia. Following morphological and molecular characterisation (targeting 16S rRNA and cytochrome c oxidase I), tick DNA extracts (n = 140) were subjected to microfluidic real-time PCR-based screening for the detection of microorganisms and Rickettsia-specific real-time qPCRs. Five species of ixodid ticks were identified, including Aponomma auruginans, Ixodes (I.) antechini, I. kohlsi, I. tasmani and I. trichosuri. Phylogenetic analyses of 16S rRNA sequences of I. tasmani revealed two subclades, indicating a potential cryptic species. The microfluidic real-time PCR detected seven different microorganisms as a single (in 13/45 ticks) or multiple infections (27/45). The most common microorganisms detected were Apicomplexa (84.4%, 38/45) followed by Rickettsia sp. (55.6%, 25/45), Theileria sp. (22.2% 10/45), Bartonella sp. (17.8%, 8/45), Coxiella-like sp. (6.7%, 3/45), Hepatozoon sp. (2.2%, 1/45), and Ehrlichia sp. (2.2%, 1/45). Phylogenetic analyses of four Rickettsia loci showed that the Rickettsia isolates detected herein potentially belonged to a novel species of Rickettsia. This study demonstrated that ticks of Australian wildlife carry a diverse array of microorganisms. Given the direct and indirect human-wildlife-livestock interactions, there is a need to adopt a One Health approach for continuous surveillance of tick-associated pathogens/microorganisms to minimise the associated threats to animal and human health.

Emerging tick-borne spotted fever group rickettsioses in the Balkans.

The impact of tick-borne pathogens (TBPs) on human health has increased in the last decades, since the incidence of emerging and re-emerging infectious and zoonotic tick-borne diseases has increased worldwide. Tick-borne rickettsiae of the Spotted Fever group (SFGR) are considered as emerging pathogens that can infect humans and cause a variety of non-specific clinical symptoms. Here, we report nine cases of atypical tick-borne diseases (9/460; 1.95%) that occurred over a period of four months (from 15 April 2021 to 16 August 2021) in Serbia, from which five cases were classified as confirmed SFGR infection, two cases as probable SFGR infection and two cases as suspected SFGR infection. Within cases of confirmed SFGR infection, R. helvetica was detected as the causative agent in two cases. The most common clinical finding was non-expanding persistent circular redness, followed by eschar and enlargement of regional lymph nodes, and pain at lesion site. Rickettsia outer membrane protein B (ompB) and citrate synthase (gltA) gene fragments were amplified from clinical samples and ticks attached to patients and IgG reacting with Rickettsia conorii antigen were detected in sera samples of patients, which are highly suggestive of exposure to SFGR. Surveillance and monitoring of rickettsial diseases in Serbia should continue and extended to new areas due to the increasing trend of clinical infections caused by SFGR in the country.

Dynamics of Infections in Cattle and Rhipicephalus microplus: A Preliminary Study.

Tick-borne pathogens (TBPs) pose a significant threat to livestock, including bovine species. This study aimed to investigate TBPs in cattle and ticks across four sampling points, utilizing real-time microfluidic PCR. The results revealed that Rhipicephalus microplus ticks were found infesting all animals. Among the detected TBPs in cattle, Anaplasma marginale was the most frequently identified, often as a single infection, although mixed infections involving Rickettsia felis, uncharacterized Rickettsia sp., and Anaplasma sp. were also observed. In ticks, A. marginale was predominant, along with R. felis, Rickettsia sp., and Ehrlichia sp. It is noteworthy that although A. marginale consistently infected all cattle during various sampling times, this pathogen was not detected in all ticks. This suggests a complex dynamic of pathogen acquisition by ticks. A phylogenetic analysis focused on the identification of Anaplasma species using amplified 16S rDNA gene fragments revealed the presence of A. marginale and Anaplasma platys strains in bovines. These findings underscore the presence of multiple TBPs in both cattle and ticks, with A. marginale being the most prevalent. Understanding the dynamics and phylogenetics of TBPs is crucial for developing effective control strategies to mitigate tick-borne diseases in livestock.