Seasonal pattern of questing ticks and prevalence of pathogenic Rickettsia and Anaplasmataceae in Khao Yai national park, Thailand.

BACKGROUND: Tick-borne diseases (TBD) are considered neglected diseases in Thailand with disease burden likely underestimated. To assess risk for emerging TBD in Thailand, the seasonality of questing tick and pathogen prevalence were studied in Khao Yai National Park, a top tourist destination. METHODS: During 2019, questing ticks around tourist attractions were systematically collected bimonthly and analyzed for Rickettsia and Anaplasmataceae bacterial species by polymerase chain reaction and DNA sequencing. RESULTS: Larvae and nymphs of questing ticks peaked in Khao Yai National Park during the late rainy-winter season, though no specific trends were observed in adult ticks. Winter (November to February) was the highest risk for human tick-bites due to higher numbers of both ticks and visitors. Of the total 5916 ticks analyzed (651 pools), Anaplasma phagocytophilum, Neoehrlichia mikurensis, Ehrlichia ewingii, and Ehrlichia chaffeensis were detected at low rates (≤0.05%). There was a higher prevalence of human rickettsioses (0.2-7%) in ticks surveyed with Rickettsia tamurae, Rickettsia raoultii, and Rickettsia montana the major species. Amblyomma ticks had the highest prevalence of Rickettsia (85%, 35/44 Amblyomma adults), in which only R. tamurae and R. raoultii were found in Amblyomma with mixed species infections common. We report the first detection of R. africae-like and N. mikurensis in Ixodes granulatus adults in Thailand, suggesting I. granulatus as a potential vector for these pathogens. CONCLUSION: This study demonstrated the risk of emerging TBD in Thailand and underscores the need for tick-bite prevention among tourists in Thailand.

Anaplasma phagocytophilum infection rates in questing and host-attached ticks: a global systematic review and meta-analysis.

Anaplasma phagocytophilum causes a multi-organ non-specific febrile illness referred to as human granulocytic anaplasmosis. The epidemiologic risk of the pathogen is underestimated despite human encroachment into the natural habitats of ticks. In this study, we performed a systematic review and meta-analysis to determine the global infection rates and distribution of A. phagocytophilum in tick vectors. We pooled data using the random-effects model, assessed individual study quality using the Joanna Briggs Institute critical appraisal instrument for prevalence studies and determined heterogeneity and across study bias using Cochran's Q-test and Egger's regression test respectively. A total of 126 studies from 33 countries across 4 continents reported A. phagocytophilum estimated infection rate of 4.76% (9453/174,967; 95% CI: 3.96, 5.71). Estimated IRs across sub-groups varied significantly (p <0.05) with a range of 1.95 (95% CI: 0.63, 5.86) to 7.15% (95% CI: 5.31, 9.56). Country-based IRs ranged between 0.42 (95% CI: 0.22, 0.80) in Belgium and 37.54% (95% CI: 0.72, 98.03) in Norway. The highest number of studies on A. phagocytophilum were in Europe (82/126) by continent and the USA (33/126) by country. The risk of transmitting this pathogens from ticks to animals and humans exist and therefore, we recommend the use of chemical and biological control measures as well as repellents and protective clothing by occupationally exposed individuals to curtail further transmission of the pathogen to humans and animals.

Abundance of Ixodes ricinus Ticks (Acari: Ixodidae) and the Diversity of Borrelia Species in Northeastern Poland.

Monitoring the abundance of ticks and the prevalence of pathogens in ticks is an important activity in assessing the risk of tick-borne diseases and helps to develop preventive measures. This study aimed to estimate the density of Ixodes ricinus, the prevalence of Borrelia species, and their diversity in northeastern Poland. The overall mean I. ricinus density was 9.7 ticks/100 m2. There were no differences between years, subregions, or habitats of study. The Borrelia infection rate was higher in females (22.6%) and males (14.3%) than in nymphs 5.5% (MIR). The most infected ticks came from the eastern subregion (10.1%) where the incidence of borreliosis among the inhabitants was over 20% higher than in the other subregions. In the infected ticks, B. afzelii (38.3%) and B. garinii (34.5%) were predominant. B. bavariensis was confirmed in I. ricinus in Poland for the first time. The most polymorphic was B. garinii. B. miyamotoi (belonged to the European type) was identified as a mono-infection in 0.9% of ticks and in 1.5% as a co-infection with B. afzelii and with B. garinii. Besides the risk of borreliosis and co-infections with different Borrelia species, physicians should also be aware of B. miyamotoi infections among patients.

Molecular Prevalence of Selected Tick-Borne Pathogens in Dermacentor reticulatus Collected in a Natural Park in Italy.

Dermacentor reticulatus is one of the most important vectors of tick-borne pathogens (TBPs) in Europe causing diseases in animals and humans. A longitudinal study was planned, aimed to detect the molecular prevalence of tick-borne pathogens, i.e., Babesia spp. and the spotted fever group Rickettsiae, and its seasonal variation in D. reticulatus questing ticks to define the temporal infection risk. Ticks were collected monthly over a period of 15 months in a peri-urban park in Lombardy, Italy. DNA extraction and molecular analyses were performed. Statistical analysis was carried out. Out of 488, 53 (P = 10.9%) adult questing ticks were positive for Babesia DNA. A higher prevalence was revealed in male (32/241, P = 13.3%) than in female (21/247, P = 8.5%) ticks. Positive ticks were mostly collected in winter months (P = 13.3%) compared to early (P = 7.9) and late (P = 12.8) spring months. A similar percentage of positive ticks was evidenced in transects 1 and 3 (5.8% and 6.5%, respectively); instead, a significant higher prevalence was recorded in transect 2 (P = 16.0%). Obtained sequences confirmed a homology of 100% with B. canis sequences deposited in GenBank. No ticks tested positive for Rickettsia spp. DNA (0/488, P = 0%). The conspicuous circulation of B. canis infection in D. reticulatus adult questing ticks confirms their role in the epidemiology of canine babesiosis and requires preventive measures for dogs in this recreational area. Even if no tick was positive for the spotted fever group Rickettsia, its capacity as a vector of zoonotic pathogens should not be neglected.

Exploring the Individual Bacterial Microbiota of Questing Ixodes ricinus Nymphs.

Ixodes ricinus is the most common hard tick species in Europe and an important vector of pathogens of human and animal health concerns. The rise of high-throughput sequencing has facilitated the identification of many tick-borne pathogens and, more globally, of various microbiota members depending on the scale of concern. In this study, we aimed to assess the bacterial diversity of individual I. ricinus questing nymphs collected in France using high-throughput 16S gene metabarcoding. From 180 dragging-collected nymphs, we identified more than 700 bacterial genera, of which about 20 are abundantly represented (>1% of total reads). Together with 136 other genera assigned, they constitute a core internal microbiota in this study. We also identified 20 individuals carrying Borreliella. The most abundant species is B. afzelii, known to be one of the bacteria responsible for Lyme disease in Europe. Co-detection of up to four Borreliella genospecies within the same individual has also been retrieved. The detection and co-detection rate of Borreliella in I. ricinus nymphs is high and raises the question of interactions between these bacteria and the communities constituting the internal microbiota.

Quantitative Parameters of the Body Composition Influencing Host Seeking Behavior of Ixodes ricinus Adults.

Ixodes ricinus, a hematophagous arthropod species with great medical importance in the northern hemisphere, is characterized by an ability to survive prolonged periods of starvation, a wide host spectrum, and high vector competence. The aim of the present study was to determine the quantitative parameters of questing I. ricinus ticks collected in eastern Poland during the spring peak of their activity. The study consisted in the determination of quantitative parameters characterizing I. ricinus females and males, i.e., fresh body mass, reduced body mass, lipid-free body mass, water mass, and lipid mass and calculation of the lipid index. A statistically significant difference was observed between the mean values of the lipid index in females collected during the first and last ten days of May, which indicates the progressive utilization of reserve materials in the activity period. Higher activity of I. ricinus female ticks was observed during the last ten days of May despite the less favorable weather conditions, indicating their strong determination in host-seeking behaviors accompanying a decline in the lipid content and the use of the "now or never" strategy.

The Bacterial Community in Questing Ticks From Khao Yai National Park in Thailand.

Ticks are known vectors for a variety of pathogens including bacteria, viruses, fungi, and parasites. In this study, bacterial communities were investigated in active life stages of three tick genera (Haemaphysalis, Dermacentor, and Amblyomma) collected from Khao Yai National Park in Thailand. Four hundred and thirty-three questing ticks were selected for pathogen detection individually using real-time PCR assays, and 58 of these were subjected to further metagenomics analysis. A total of 62 ticks were found to be infected with pathogenic bacteria, for a 14.3% prevalence rate, with Amblyomma spp. exhibiting the highest infection rate (20.5%), followed by Haemaphysalis spp. (14.5%) and Dermacentor spp. (8.6%). Rickettsia spp. were the most prevalent bacteria (7.9%) found, followed by Ehrlichia spp. (3.2%), and Anaplasma spp. and Borrelia spp. each with a similar prevalence of 1.6%. Co-infection between pathogenic bacteria was only detected in three Haemaphysalis females, and all co-infections were between Rickettsia spp. and Anaplasmataceae (Ehrlichia spp. or Anaplasma spp.), accounting for 4.6% of infected ticks or 0.7% of all examined questing ticks. The prevalence of the Coxiella-like endosymbiont was also investigated. Of ticks tested, 65.8% were positive for the Coxiella-like endosymbiont, with the highest infection rate in nymphs (86.7%), followed by females (83.4%). Among tick genera, Haemaphysalis exhibited the highest prevalence of infection with the Coxiella-like endosymbiont. Ticks harboring the Coxiella-like endosymbiont were more likely to be infected with Ehrlichia spp. or Rickettsia spp. than those without, with statistical significance for Ehrlichia spp. infection in particular (p-values = 0.003 and 0.917 for Ehrlichia spp. and Rickettsia spp., respectively). Profiling the bacterial community in ticks using metagenomics revealed distinct, predominant bacterial taxa in tick genera. Alpha and beta diversities analyses showed that the bacterial community diversity and composition in Haemaphysalis spp. was significantly different from Amblyomma spp. However, when examining bacterial diversity among tick life stages (larva, nymph, and adult) in Haemaphysalis spp., no significant difference among life stages was detected. These results provide valuable information on the bacterial community composition and co-infection rates in questing ticks in Thailand, with implications for animal and human health.

Tick species diversity and population dynamics of Ixodes ricinus in Galicia (north-western Spain).

Ticks are the major arthropod vectors of animal and human pathogens in Europe; thus, information of tick species and their phenology allows identifying those areas and periods involving a high risk of exposure to particular tick-borne pathogens. In order to assess the diversity of exophilic tick species in north-western Spain, questing ticks were collected by flagging in 17 different locations; ticks were microscopically and molecularly identified at the 16S rDNA loci. In addition, phenology of questing ticks in the same region was studied in representative sampling areas of three different ecological regions, where ticks were monthly collected in 300-m transects during two complete years. The possible influence of different climate variables on the normalized accumulated fraction of nymphal and adult Ixodes ricinus was assessed using a generalized linear mixed model (GLMN). I. ricinus was the most frequently and widely distributed tick species; Dermacentor marginatus, Dermacentor reticulatus, Ixodes frontalis and Ixodes acuminatus were sporadically found. Nymphs were the most commonly collected stage. Both nymphs and adults of I. ricinus presented a unimodal activity pattern; density showed a peak in late spring-early summer whereas the lowest levels were found at late summer-early autumn. GLMN showed that both nymphal and adult I. ricinus phenology was directly associated with the accumulated temperature, photoperiod and average daily maximum temperature and inversely associated with average daily maximum relative humidity. This information could be used to predict the annual periods associated with a higher exposure and risk of transmission of pathogens and to complete the global map of distribution of tick species.

Borrelia Diversity and Co-infection with Other Tick Borne Pathogens in Ticks.

Identifying Borrelia burgdorferi as the causative agent of Lyme disease in 1981 was a watershed moment in understanding the major impact that tick-borne zoonoses can have on public health worldwide, particularly in Europe and the USA. The medical importance of tick-borne diseases has long since been acknowledged, yet little is known regarding the occurrence of emerging tick-borne pathogens such as Borrelia spp., Anaplasma phagocytophilum, Rickettsia spp., Bartonella spp., "Candidatus Neoehrlichia mikurensis", and tick-borne encephalitis virus in questing ticks in Romania, a gateway into Europe. The objective of our study was to identify the infection and co-infection rates of different Borrelia genospecies along with other tick-borne pathogens in questing ticks collected from three geographically distinct areas in eastern Romania. We collected 557 questing adult and nymph ticks of three different species (534 Ixodes ricinus, 19 Haemaphysalis punctata, and 4 Dermacentor reticulatus) from three areas in Romania. We analyzed ticks individually for the presence of eight different Borrelia genospecies with high-throughput real-time PCR. Ticks with Borrelia were then tested for possible co-infections with A. phagocytophilum, Rickettsia spp., Bartonella spp., "Candidatus Neoehrlichia mikurensis", and tick-borne encephalitis virus. Borrelia spp. was detected in I. ricinus ticks from all sampling areas, with global prevalence rates of 25.8%. All eight Borrelia genospecies were detected in I. ricinus ticks: Borrelia garinii (14.8%), B. afzelii (8.8%), B. valaisiana (5.1%), B. lusitaniae (4.9%), B. miyamotoi (0.9%), B. burgdorferi s.s (0.4%), and B. bissettii (0.2%). Regarding pathogen co-infection 64.5% of infected I. ricinus were positive for more than one pathogen. Associations between different Borrelia genospecies were detected in 9.7% of ticks, and 6.9% of I. ricinus ticks tested positive for co-infection of Borrelia spp. with other tick-borne pathogens. The most common association was between B. garinii and B. afzelii (4.3%), followed by B. garinii and B. lusitaniae (3.0%). The most frequent dual co-infections were between Borrelia spp. and Rickettsia spp., (1.3%), and between Borrelia spp. and "Candidatus Neoehrlichia mikurensis" (1.3%). The diversity of tick-borne pathogens detected in this study and the frequency of co-infections should influence all infection risk evaluations following a tick bite.

Seasonal dynamics of adult Dermacentor reticulatus in a peri-urban park in southern Europe.

Studies on the human-biting pathogen vectors Dermacentor reticulatus and Ixodes ricinus have been scarce in southern Europe. The aims of the present study were to determine the abundance of these ticks in a peri-urban park in northern Italy, describe the seasonal activity of D. reticulatus and examine the correlation between tick occurrence and environmental factors. Ticks were collected monthly from April 2015 to May 2016 using both dragging and flagging techniques. Various climatic variables (mean temperature, relative humidity and evapotranspiration for the collection dates; and 30-day moving averages preceding each collection date were calculated for rainfall, temperature, relative humidity and saturation deficit) also were recorded. Overall, 444 adults of D. reticulatus and 10 adults of I. ricinus were collected. Males of D. reticulatus appeared earlier in the year than females, but overall females were collected more frequently than males (1:1.25). Statistical analysis showed significant differences in the density of D. reticulatus among sampling transects and among months. The seasonal dynamic of D. reticulatus was characterized by a single peak of activity in the early spring. Tick density was associated with climatic variables: the 30-day moving average saturation deficit was particularly significant as most ticks (83%) were collected at values below 5.2mmHg. At the level of individual sampling transects, seasonal dynamics could be influenced by habitat type and host availability. We found D. reticulatus to be most abundant in mixed forests dominated by oaks and rich in ponds. As to I. ricinus, though found in a small number, its presence can be confirmed by our investigation. In light of the results of this study, the risk for encounters with D. reticulatus and I. ricinus may be higher than previously thought in northern Italy.

Diversity of ticks (Acari: Ixodidae) infesting cheetahs (Acinoyx jubatus) at three breeding centres in South Africa and activity patterns of questing ticks.

Ticks were collected from 191 cheetahs at three breeding centres in North West and Limpopo Provinces, South Africa. Haemaphysalis elliptica, a common tick of large felids, was the most abundant species collected, while Amblyomma hebraeum and Rhipicephalus simus occurred in lower numbers. In addition to these three species, drag-sampling of the vegetation revealed the presence of Amblyomma marmoreum, Rhipicephalus (B.) decoloratus and Rhipicephalus zambeziensis. The presence of free-ranging antelopes, murid rodents and tortoises at the breeding centres probably contributed to the availability of immature tick stages on the vegetation. Diurnal and seasonal questing patterns of ixodid ticks were investigated at monthly intervals at the largest cheetah-breeding centre. Questing ticks were most abundant on the vegetation during the warm summer months. Most questing H. elliptica larvae and nymphs were collected from the vegetation in the early morning and late afternoon and fewest during the middle of the day.

Questing Amblyomma mixtum and Haemaphysalis juxtakochi (Acari: Ixodidae) Infected with Candidatus "Rickettsia amblyommii" from the Natural Environment in Panama Canal Basin, Panama.

This work emphasizes the detection of Candidatus "Rickettsia amblyommii" in questing Haemaphysalis juxtakochi and Amblyomma mixtum. From February 2009 to December 2012, questing ticks were collected from the vegetation and leaf-litter of four protected forests and two grassy areas around the Panama Canal basin. DNA was extracted from Amblyomma mixtum, Amblyomma naponense, Amblyomma oblongoguttatum, Amblyomma pecarium, Amblyomma tapirellum, Haemaphysalis juxtakochi, and unidentified immature Amblyomma. Specific primers of citrate synthase gene gltA were used to detect and identify the rickettsiae. Amplicons with the expected band size were purified and sequenced. DNA of C. "R. amblyommii" was found in A. mixtum, H. juxtakochi and Amblyomma immatures. To our knowledge, these finding represent the first report of C. "R. amblyommii" in free-living ticks in the wilderness of Central America.