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Article

Tick Infestation and Molecular Detection of Tick-Borne Pathogens from Indian Long-Eared Hedgehogs (Hemiechinus collaris) in Pakistan

by
Shahzad Ali
1,*,
Michael E. von Fricken
2,*,
Asima Azam
3,
Ahmad Hassan
1,
Nora G. Cleary
2,
Kiran Iftikhar
4,
Muhammad Imran Rashid
5 and
Abdul Razzaq
6
1
Wildlife Epidemiology and Molecular Microbiology Laboratory (One Health Research Group), Discipline of Zoology, Department of Wildlife and Ecology, University of Veterinary and Animal Sciences, Ravi Campus, Lahore 54000, Pakistan
2
Department of Environmental and Global Health, University of Florida, Gainesville, FL 32610, USA
3
Department of Zoology, Shaheed Benazir Bhutto Women University, Peshawar 25000, Pakistan
4
Department of Mathematics and Statistics, University of Agriculture Faisalabad, Faisalabad 38000, Pakistan
5
Department of Parasitology, University of Veterinary and Animal Sciences, Lahore 54200, Pakistan
6
Animal Sciences Division, Pakistan Agricultural Research Council, Islamabad 44000, Pakistan
*
Authors to whom correspondence should be addressed.
Animals 2024, 14(22), 3185; https://doi.org/10.3390/ani14223185
Submission received: 23 September 2024 / Revised: 30 October 2024 / Accepted: 4 November 2024 / Published: 6 November 2024
(This article belongs to the Special Issue Disease and Health in Free-Ranging and Captive Wildlife: Second Edition)
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Simple Summary

Hedgehogs play a significant role as a vertebrate host for ticks and the tick-borne pathogens (TBPs) they carry, which include bacterial, viral, and parasitic diseases of concern for both humans and livestock. Not only are TBPs a major health concern, but they also directly impact the economy and livelihood of farmers and pastoral systems that rely on livestock. This study was conducted to molecularly confirm tick species infesting hedgehogs and to detect TBPs in both ticks and hedgehogs. The DNA of ticks and the blood of the hedgehogs were extracted and analyzed. Based on the sequence results, Rhiphicephalus turanicus was identified as the only tick species in this study. Anaplasma marginale, Babesia bigemina, and Theileria lestoquardi were detected in the ticks, and A. marginale was detected in the blood of one hedgehog. Phylogenetic analysis revealed that these pathogens were highly similar to sequences from cattle blood reported in Thailand, the United States, India, South Africa, and Uganda. By understanding genetic differences and similarities, improved measures to monitor for TBPs as well as interventions that aim to protect human and animal populations can be implemented.

Abstract

Hedgehogs can act as reservoirs for the transmission of tick-borne pathogens (TBPs) to domestic livestock, wild animals, and humans. Understanding host–tick dynamics is essential to evaluate the impact of TBPs. This study was conducted in Pakistan and aimed to determine the prevalence and species of TBPs in the blood and ticks of Indian long-eared hedgehogs captured from various environments. A total of 64 hedgehogs were captured to check for tick infestation. Tick species were identified morphologically and molecularly including ITS-2 region amplification by PCR and subsequent Sanger sequencing. Moreover, TBPs were identified in both ticks and the blood of hedgehogs through conventional PCR and sequencing, targeting the regions msp1b, 18S rRNA, and cytb for Anaplasma spp., Babesia spp., and Theileria spp., respectively. Out of 64 hedgehogs, 16 (25%) were found to be infested with ticks. Morphological and molecular analysis identified all 109 collected ticks as Rhipicephalus turanicus. Only one hedgehog (6.2%) was infected with A. marginale. From the tick samples, 3.7% tested positive for Theileria lestoquardi, 2.8% for Anaplasma marginale, and another 2.8% for Babesia bigemina. This study provides critical insights into circulating TBPs in this region and what possible role hedgehogs might play in disease maintenance for Anaplasma marginale while identifying multiple pathogens that are of concern to human and animal health.

1. Introduction

Ticks are well-known vectors of disease and have been recorded on numerous domestic and wild animals, [1,2] transmitting pathogens including bacteria (Anaplasma spp., Borrelia spp., Coxiella spp., Ehrlichia spp., and Francisella spp.), viruses (Crimean-Congo Haemorrhagic Fever virus, and Tick-Borne Encephalitis virus), and parasites (Babesia spp. and Theileria spp.) [3,4]. In Pakistan, tick genera, Haemaphysalis, Hyalomma, Ornithodoros, and Rhipicephalus, have been reported to infest humans and animals [5], with Hyalomma, Rhipicephalus, and Haemaphysalis genera most commonly found in Pakistan’s Punjab province [6].
There are three major tick-borne diseases (TBDs), anaplasmosis, babesiosis, and theileriosis, which pose a significant risk to livestock in Pakistan [7]. Anaplasmosis is distributed worldwide in tropical and subtropical areas often in bovines and is mainly caused by Anaplasma marginale followed by A. centrale. Babesiosis, a disease of veterinary and human importance, can be caused by Babesia bovis and B. bigemina in livestock. Theileriosis, which is found throughout Africa and Asia, is responsible for severe economic losses in cattle, water buffaloes, sheep, goats, and horses [8,9]. The main vectors of these diseases in Pakistan are Rhipicephalus spp. and Hyalomma spp. All three diseases can result in reduced animal production and decreased meat and milk yield, ultimately affecting the livelihood of farmers with a mortality rate upwards of 90% from babesiosis and theileriosis. The surveillance of tick-infested animals in Pakistan is vital to monitor the threat of TBDs for veterinary health [6].
The Indian long-eared hedgehog (Hemiechinus collaris), a member of the Chordata and Mammalia phyla, the Eulipotyphla order, and the Erinaceidae family, is native to Central Asia and parts of the Middle East. Hedgehogs play an important role in the transmission and maintenance of zoonotic pathogens as they serve as reservoirs and ectoparasite hosts to a range of diseases and arthropod vectors [10]. Tick species from the genera Hyalomma, Amblyomma, Haemaphysalis, Dermacentor, Ornithodoros, and Rhipicephalus have been recorded on hedgehogs [11,12]. Tick-infested hedgehogs are known to harbor Borrelia spp., Rickettsia felis, A. marginale, and A. phagocytophilum [12,13,14]. The domestication of hedgehogs has become more frequent in recent decades, resulting in greater animal–human contact, therefore increasing the threat of TBP spillover to people from hedgehogs. While hedgehogs are widely distributed in Pakistan, especially in the South, there is limited information regarding ticks and TBPs of hedgehogs. The characterization of these pathogens and their vectors is critical to improving our understanding of their reservoir hosts, distribution, and the potential threat they pose to human and animal health. The present study aims to determine the prevalence and molecular confirmation of tick species and TBPs associated with Indian long-eared hedgehogs in Pakistan.

2. Materials and Methods

2.1. Study Site

This study was conducted in the Sahiwal division (which contains three districts: Sahiwal, Okara, and Pakpattan) of Punjab province in Pakistan from December 2019 to December 2020 (Figure 1). Sahiwal has a total area of 10,302 km2. This region has extreme weather conditions due to substantial temperature variation, a low of 5 °C during winter and a high of 50 °C in the summer, along with an annual average rainfall of 349 mm [15]. Sahiwal division is recognized for its livestock production, with a focus on rearing Sahiwal cattle breeds and Nili-Ravi buffaloes. In addition to these breeds, sheep and goats are also raised in this area. Unfortunately, these livestock species are susceptible to various ticks and TBPs [16,17].

2.2. Sampling and Data Collection

Live cage traps and hand nets were used to collect Indian long-eared hedgehogs (Hemiechinus collaris). Hedgehog traps were set at dusk, baited with peanut butter and apples, and examined for captured hedgehogs at dawn. The entire body of the hedgehog was examined for ticks, which were removed using a delicate tweezer without damaging tick mouth parts. The ticks were then preserved in sealed bottles containing 70% ethanol following collection, as previously described [18]. Blood samples were collected from the jugular vein of each tick-infested hedgehog using a 3 mL syringe fitted with a 22-gauge needle [19]. The field data, including sex, age, urbanicity, habitat, and district, were recorded. The hedgehogs were released back to their natural habitat after the collection of samples and data.

2.3. Tick Identification

A total of 109 ticks were morphologically identified using taxonomic keys under a stereomicroscope (Euromex, Duiven, The Netherlands) as to the sex, developmental stage (nymph/adult), and species level [20].

2.4. DNA Extraction from Ticks and Blood

The ticks were homogenized in liquid nitrogen using a mortar and pestle and then placed into a 1.5 mL microcentrifuge tube. DNA was extracted from the homogenized ticks (20 mg) and hedgehog blood (200 µL) using the Thermo Scientific GeneJET genomic DNA purification kit (Thermo Fisher Scientific, Vilnius, Lithuania) according to manufacturer instructions. DNA concentration was assessed with the Nanodrop 2000 spectrophotometer (Thermo Fisher Scientific, Waltham, MA, USA). Purified DNA was kept at −20 °C till further use.

2.5. PCR Confirmation of Tick Species

Tick DNA was detected by the amplification of the intergenic spacer 2 (ITS2) region (710 bp) using the following primers: ITS-2F (5′-AGGACACACTGAGCACTGATTC-‘3) and ITS-2R (5′-ACTGCGAAGCACTTRGACCG-‘3) for the molecular identification of tick species [21]. The PCR mixture was made of 10 µL master mix (WizBio, Seongnam-si, Republic of Korea), 2.5 µL of template DNA, 1 µL of forward primer, 1 µL of reverse primer (Macrogen, Seoul, Republic of Korea), and 10.5 µL of DEPC water (Thermo Fisher Scientific, Waltham, MA, USA) for a total volume of 25 µL. The PCR was run for 35 cycles with the first denaturation for 5 min at 95 °C, the second for 30 s at 95 °C, annealing for 30 s at 57 °C, extension for 30 s at 72 °C, and the final extension for 5 min at 72 °C [21].

2.6. PCR Screening for Microbes

The genus-specific primers corresponding to msp1b, 18S rRNA, and cytb regions were used for the detection of Anaplasma spp., Babesia spp., and Theileria spp. in ticks and blood, respectively (Table 1). The PCR master mix consisted of 12 µL master mix (WizBio, Seongnam-si, Republic of Korea), 5 µL of template DNA, 1 µL of forward primer, 1 µL of reverse primer (Macrogen, Seoul, Republic of Korea), and 6 µL of DEPC water (Thermo Fisher Scientific, Waltham, MA, USA) for a total volume of 25 µL. The amplification of target regions was confirmed on a 1.5% Agarose gel (Bio World, Visalia, CA, USA) using electrophoresis. The PCR results were visualized with Bio-Rad gel visualization equipment (Bio-Rad, Berkeley, CA, USA).

2.7. Sequencing and Statistical Analysis

The PCR-positive samples were sequenced by Microgen, Seoul, Korea. All sequences were trimmed with a biological sequence alignment editor (BioEdit 7.2). To compare the identity of each tick and TBPs, reference sequences were obtained from GenBank. The sequences were aligned in Geneious Prime 2022.11.0.14.1 using the MAFFT algorithm. Maximum likelihood phylogenetic trees were created in MEGA 10.2.6 using the best match substitution model for each corresponding pathogen with bootstrap repetitions set at 1000 [24,25]. The bootstrap values under 50% were excluded from the phylogenetic trees. Statistical analysis was performed using the SPSS version 21 statistical tool (IBM Armonk, New York, NY, USA). A host with a single tick present was classified as infested, while a host with no ticks detected was considered non-infested. The statistical significance of tick and tick-borne pathogens prevalence was determined using a Chi-square test with significance set at 5% [26].

3. Results

3.1. Tick Infestation

From December 2019 to December 2020, 64 hedgehogs were captured from the Sahiwal division and 16 (25%) were infested with ticks. Female hedgehogs had a higher tick infestation (29.6%) compared to male hedgehogs (21.6%). A higher number of adult hedgehogs were captured than juvenile hedgehogs, but juvenile hedgehogs had a higher prevalence of ticks (31.2%) than adults (22.9%). A similar number of hedgehogs captured in rural areas (26.3%) and urban areas (23.1%) were found to be infested with ticks. The prevalence of ticks was also similar on hedgehogs found in agricultural land (25.7%) and animal farms (24.1%). Tick prevalence was comparable in all three districts: Okara (26.1%), Pakpattan (25%), and Sahiwal (23.8%). No significant difference was found among these factors for the prevalence of tick infestation (p > 0.05) (Table 2). A total of 109 R. turanicus ticks were collected from long-eared hedgehogs. Male ticks were more prevalent (67.9%; n = 74) compared to female ticks (32.1%; n = 35). The ticks were in two life stages: adult (91.7%; n = 100) and nymph (8.3%; n = 9) (Table 3).

3.2. Prevalence of TBPs in Hedgehog Blood and Ticks

Anaplasma marginale was detected in an adult male hedgehog captured from a rural area of district Sahiwal (Table 4). Out of 109 ticks, 10 (9.2%) were positive for TBPs such as T. lestoquardi (3.7%), A. marginale (2.8%), and B. bigemina (2.8%). A higher number of male ticks were infected with T. lestoquardi (2.7%) and A. marginale (2.7%) compared to B. bigemina (1.4%). However, female ticks had higher infection rates of T. lestoquardi (5.7%) and B. bigemina (5.7%) than A. marginale (2.9%). Adult ticks were comparatively more infected with T. lestoquardi (4.0%) compared to B. bigemina (3.0%) and A. marginale (3.0%). All nymphs tested negative for any pathogen (Table 5).

3.3. Phylogenetic Analysis

The sequences of the ticks in the present study were 100% identical to those of R. turanicus reported from Israel (KF958426). We identified three unique sequences of A. marginale from three samples, three identical sequences of B. bigemina from three samples, and three unique sequences of T. lestoquardi from four samples in this study. Phylogenetic analysis revealed A. marginale (PP798377) was highly similar to sequences from cattle blood in Thailand, MT796683. Other A. marginale sequences (PP798376 and PP798375) were identical to two sequences from cattle ticks in the United States, AF348137, and the Florida strain, AF221693, respectively (Figure 2). Babesia bigemina sequences were highly similar to many sequences from cattle blood in Bolivia (LC645222), India (MT322431), Pakistan (KY656456), South Africa (MH257723), and Uganda (KU206297). Lastly, T. lestoquardi sequences were highly similar to Iraqi sheep blood, OQ304461, OQ304462, and OQ304458, and Indian sheep blood, MZ665958 (Figure 3).
The sequences obtained in this study were submitted to GenBank under the following accession numbers: PP798375.1, PP798376.1, and PP798377.1 (A. marginale), PP809859.1, PP809860.1, and PP809861.1 (B. bigemina), and PP798371.1, PP798372.1, PP798373.1 and PP798374.1 (T. lestoquardi). All sequences were from tick samples aside from PP798375.1 which was obtained from Indian long-eared hedgehog blood.

4. Discussion

Our study only detected the presence of a single tick species, R. turanicus, which was found on 25% of collected hedgehogs. Rhipicephalus species are potential vectors and reservoirs for several diseases [27,28,29]. Tick prevalence was similar across all three districts: Okara, Pakpattan, and Sahiwal, with no significant differences in the number of ticks collected. Contrarily, research on ectoparasite infestations in European hedgehogs in Iran differed by city, with Urmia City reporting a tick infestation rate of 67.70%, whereas Tabriz City reported an infestation rate of 5.26% [30,31]. This study also identified all ticks as R. turanicus. Additionally, a study conducted in Tokat City, Central Anatolia, Turkey, found a higher tick infestation rate of R. turanicus at 77.80% [32]. Another study in Pakistan identified R. turanicus as the most prevalent tick species, with a 50% infestation rate in free-ranging wild animals [33]. These variations in tick infestation among studies can likely be attributed to differences in geography, temperature, and climate conditions across the studied regions [34]. The two most common tick species observed in European hedgehogs (Erinaceus europaeus) in northern Iran were the sheep tick (Ixodes ricinus) and the hedgehog tick (Ixodes hexagonus), which differ from the ticks, R. turanicus, found on the Indian long-eared hedgehog. The geographical spread of I. ricinus includes Europe and Northern Africa where I. hexagonus includes Europe and central Asia, corresponding with the distribution of their European hedgehog host [34,35]. The exclusion of European hedgehogs in Pakistan may explain the lack of detection of I. ricinus and I. hexagonus in this study. In the Ninevah district in Iraq, R. sanguineus, Haemaphysalis erinacei, Boophilus annulatus, and Hyalomma detritum were found on the long-eared hedgehog [36]. Furthermore, Shubber et al. [37] discovered that Hyalomma spp., R. leparis, and R. turanicus had high infestation rates (76.19%) in long-eared hedgehogs. Additional surveillance on long-eared hedgehogs in Pakistan is needed across a wider geographic range to better understand the diversity of ticks, TBPs, and differences in tick infestation rates. The tick prevalence in livestock in the region of Pakistani Punjab is increasing rapidly, highlighting the need for public health awareness about TBPs and quantifying their distribution in animals [38,39,40].
In this study, female hedgehogs were observed to have a higher tick infestation compared to males; however, no significant association was detected. This finding is consistent with a study conducted in Iraq, where female long-eared hedgehogs (H. auritus) exhibited a higher infestation rate (50%) compared to males (44.44%) [41]. Similarly, research conducted by Egyed et al. [42] in Hungary on the northern white-breasted hedgehog (Erinaceus roumanicus) in an urban park found that tick prevalence was higher in females (84.21%) than in males (59.65%), whereas no significant differences in sex were found in the hedgehogs of Britain and Germany [43,44]. Variations in sex could likely be explained by behavior variations in male hedgehogs compared to females [45]. In this investigation, young Indian long-eared hedgehogs did not have a statistically significant higher infestation rate compared to adult Indian long-eared hedgehogs. This aligns with no significant difference in tick infestation detected in hedgehogs in Belgium [46]. In contrast, a study on European hedgehogs (Erinaceus europaeus) in Germany found subadult hedgehogs had a higher prevalence of tick infestations compared to adults [47]. Variations in tick infestation in hedgehogs of different life stages could be linked to intensified activity levels in younger animals, who may be at a higher risk of pathogen infection due to undeveloped immune systems [48].
From our findings, tick prevalence was similar in Indian long-eared hedgehogs captured from rural areas and urban areas. Hedgehogs naturally inhabit rural areas, where previous research observed European hedgehogs captured from golf courses having higher tick prevalence than those from peri-urban areas [45]. They tend to travel across various agricultural parcels, avoiding areas with significant human populations. The fragmented nature of livestock farms, characterized by multiple walls and fences, forces hedgehogs to cover greater distances with fewer restrictions than in agricultural lands [49,50]. The proximity of agricultural land to natural environments provides a richer variety of habitats compared to non-agricultural areas, often leading to higher species diversity and, consequently, a larger number of ticks [51,52]. We did not detect significant variability in tick infestations between agricultural land and animal farms, which could be explained by our limited sample size.
Detected pathogens belong to the genera of the three most important economic TBDs globally: Anaplasma, Babesia, and Theileria. Among R. turanicus, the positive detection rates for A. marginale and Babesia bigemina were both 2.8%. Both A. marginale and B. bigemina are endemic in Pakistan [53]. Anaplasma marginale has previously been detected from ticks collected from cattle and water buffalo in Pakistan including R. turanicus, R. microplus, and H. anatolicum in Punjab province [53]. In the current study, A. marginale was detected in the blood of one hedgehog, with a prevalence rate of 6.3%. Two hedgehogs collected from the Zabol and Bonjar areas of Iran were found to be infected with Anaplasma (3.8%), and three R. turanicus pools from these infected hedgehogs tested positive for Anaplasma spp. [12]. In Punjab province, A. marginale and B. bigemina have been detected in cattle in a nearby district, Sargodha, Pakistan [54]. The detection of these pathogens in cattle as well as hedgehogs in Pakistan indicates their reservoir potential for tick-borne diseases in this region [55]. This enables ticks to transmit the pathogen from reservoir hosts, such as hedgehogs, to other animals, facilitating the spread of tick-borne pathogen infection. A. marginale and B. bigemina can be transmitted by multiple tick species, increasing the risk of their spread to livestock. Theileria lestoquardi in this study had a prevalence of 3.7% in R. turanicus and is the most common Theileria spp. found in small ruminants in Africa and Asia [8]. In western Iran, T. lestoquardi has been recorded in R. turanicus, while in Pakistan and India, it has been detected in goats and sheep [56]. Our study serves as a foundation for future epidemiological studies into ticks, TBPs, and the role hedgehogs play in disease maintenance and potential spillover to livestock and humans.
A limitation of this study is the focus on three specific pathogen taxa, Anaplasma, Babesia, and Theileria. Therefore, we were not able to assess for the presence of other pathogen species that may be transmitted by R. turanicus ticks in this region.

5. Conclusions

A high prevalence of R. turanicus in Indian long-eared hedgehogs was detected in the three study districts in Pakistan. Rhipicephalus turanicus harbors numerous pathogens that can cause clinical illness in humans and animals. Due to changes in tick habitat, paired with human, animal, and wildlife behavior changes related to climate change and anthropogenic activities, continued surveillance is necessary to monitor disease dynamics over time. Further epidemiological research is needed in Pakistan outside of Punjab that expands to include other small mammal hosts and tick species of concern, which in time can be used to guide effective prevention and control measures.

Author Contributions

Conceptualization, S.A.; data curation, A.A., A.H. and N.G.C.; formal analysis, M.E.v.F., N.G.C. and K.I.; funding acquisition, S.A. and M.E.v.F.; investigation, S.A., M.I.R. and A.R.; methodology, M.E.v.F., A.H., N.G.C., M.I.R. and A.R.; software, A.H. and K.I.; validation, A.A.; writing—original draft, S.A., M.E.v.F., A.A., A.H., N.G.C., K.I., M.I.R. and A.R.; writing—review and editing, S.A., M.E.v.F., A.A., A.H., N.G.C., K.I., M.I.R. and A.R. All authors have read and agreed to the published version of the manuscript.

Funding

This research was funded by the Agricultural Linkages Program (ALP)-PARC, Pakistan, grant number AS 141.

Institutional Review Board Statement

The use of animals for research purposes was performed according to the Institutional Ethical Review Committee for Animal Care and Use, the University of Veterinary and Animal Sciences, Lahore, Pakistan, via approval number DR/750, Dated 15 July 2019.

Informed Consent Statement

Not applicable.

Data Availability Statement

The data presented in this study are available within the article.

Conflicts of Interest

The authors declare no conflicts of interest.

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Animals 14 03185 g001
Figure 1. A map showing the area for hedgehog collections in the Sahiwal division, containing three districts: Sahiwal, Okara, and Pakpattan.
Figure 1. A map showing the area for hedgehog collections in the Sahiwal division, containing three districts: Sahiwal, Okara, and Pakpattan.
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Animals 14 03185 g002
Figure 2. Maximum likelihood tree using the K2 + G model from the msp1b region of Anaplasma. Sequences from this study are bolded and outlined by the black rectangles. Corresponding country information of GenBank samples is provided to the left of the tree and pathogen species are color coded.
Figure 2. Maximum likelihood tree using the K2 + G model from the msp1b region of Anaplasma. Sequences from this study are bolded and outlined by the black rectangles. Corresponding country information of GenBank samples is provided to the left of the tree and pathogen species are color coded.
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Animals 14 03185 g003
Figure 3. Maximum likelihood tree using the T92 model from the cytb region of Theileria. Sequences from this study are bolded and outlined by the black rectangles. Corresponding country information of GenBank samples is provided to the left of the tree and pathogen species are color coded.
Figure 3. Maximum likelihood tree using the T92 model from the cytb region of Theileria. Sequences from this study are bolded and outlined by the black rectangles. Corresponding country information of GenBank samples is provided to the left of the tree and pathogen species are color coded.
Animals 14 03185 g003
Table 1. List of genus-specific primers used for the detection of TBPs in ticks and blood.
Table 1. List of genus-specific primers used for the detection of TBPs in ticks and blood.
PathogenSequenceTarget RegionSize (bp)Reference
Anaplasma spp.5′-CAGAGCATTGACGCACTACC-3′msp1b245[22]
5′-TCCAGACCTTCCCTAACTA-3′
Babesia spp.5′-AGAGGGACTCCTGTGCTTCA-‘318s rRNA321[23]
5′-GACGAATCGGAAAAGCCACG-‘3
Theileria spp.5′-AAGTATAGCAACTGCTTTTGTT-3′Cyt-b517[23]
5′-TCCTGCCATTGCCAAAAGTC-3′
Table 2. Risk factors associated with tick infestation in the Indian long-eared hedgehog population based on Chi-square analysis from Sahiwal division, Punjab province, Pakistan.
Table 2. Risk factors associated with tick infestation in the Indian long-eared hedgehog population based on Chi-square analysis from Sahiwal division, Punjab province, Pakistan.
VariablesCategoryTotal CapturedInfestedPrevalence (%)Chi-Squarep-Value
SexFemale27829.60.4650.563
Male37821.6
AgeAdult481122.90.5050.519
Young16531.2
UrbanicityRural381026.30.7690.506
Urban26623.1
HabitatAgriculture land35925.70.8850.559
Animal farm29724.1
DistrictOkara23626.10.0300.985
Pakpattan20525.0
Sahiwal21523.8
Table 3. Sex and life stage prevalence of R. turanicus.
Table 3. Sex and life stage prevalence of R. turanicus.
VariablesCategoryTotalPrevalence (%)
SexMale7467.9
Female3532.1
Life stageAdult10091.7
Nymph98.3
Table 4. Infection rates of tick-borne pathogens in hedgehog (n = 16) blood using conventional PCR.
Table 4. Infection rates of tick-borne pathogens in hedgehog (n = 16) blood using conventional PCR.
VariablesAnaplasma marginale
Sex
Male (n = 8)1 (12.5%)
Female (n = 8)0
Age
Adult (n = 11)1 (9.1%)
Young (n = 5)0
Urbanicity
Rural (n = 10)1 (10.0%)
Urban (n = 6)0
Location
Okara (n = 6)0
Pakpattan (n = 5)0
Sahiwal (n = 5)1 (20.0%)
Table 5. Infection rate of tick-borne pathogens in ticks (n = 109) using conventional PCR.
Table 5. Infection rate of tick-borne pathogens in ticks (n = 109) using conventional PCR.
VariablesCategoryAnaplasma
marginale		Babesia
bigemina		Theileria lestoquardiTotal
Sex Male2 (2.7%)1 (1.4%)2 (2.7%)74
Female1 (2.9%)2 (5.7%)2 (5.7%)35
Life stageAdult3 (3.0%)3 (3.0%)4 (4.0%)100
Nymph0009
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Ali, S.; von Fricken, M.E.; Azam, A.; Hassan, A.; Cleary, N.G.; Iftikhar, K.; Rashid, M.I.; Razzaq, A. Tick Infestation and Molecular Detection of Tick-Borne Pathogens from Indian Long-Eared Hedgehogs (Hemiechinus collaris) in Pakistan. Animals 2024, 14, 3185. https://doi.org/10.3390/ani14223185

AMA Style

Ali S, von Fricken ME, Azam A, Hassan A, Cleary NG, Iftikhar K, Rashid MI, Razzaq A. Tick Infestation and Molecular Detection of Tick-Borne Pathogens from Indian Long-Eared Hedgehogs (Hemiechinus collaris) in Pakistan. Animals. 2024; 14(22):3185. https://doi.org/10.3390/ani14223185

Chicago/Turabian Style

Ali, Shahzad, Michael E. von Fricken, Asima Azam, Ahmad Hassan, Nora G. Cleary, Kiran Iftikhar, Muhammad Imran Rashid, and Abdul Razzaq. 2024. "Tick Infestation and Molecular Detection of Tick-Borne Pathogens from Indian Long-Eared Hedgehogs (Hemiechinus collaris) in Pakistan" Animals 14, no. 22: 3185. https://doi.org/10.3390/ani14223185

APA Style

Ali, S., von Fricken, M. E., Azam, A., Hassan, A., Cleary, N. G., Iftikhar, K., Rashid, M. I., & Razzaq, A. (2024). Tick Infestation and Molecular Detection of Tick-Borne Pathogens from Indian Long-Eared Hedgehogs (Hemiechinus collaris) in Pakistan. Animals, 14(22), 3185. https://doi.org/10.3390/ani14223185

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