Microbiome variation between male and female Hyalomma dromedarii ticks from camels in the UAE

Abstract

Hyalomma dromedarii, a predominant camel tick species in the Arabian Peninsula, plays a significant role in pathogen transmission, yet its sex-specific microbiome composition remains poorly understood. This study analyzed the bacterial communities of male and female H. dromedarii ticks collected from camels across three locations in the United Arab Emirates: Al Khazna, Al Ja’ae, and Al Jabeeb. High throughput 16S rRNA gene sequencing revealed a total of 432,495 reads clustering into 848 operational taxonomic units, encompassing 20 phyla and 261 genera. Male ticks consistently exhibited higher microbial diversity and genus richness compared to females. Notably, Francisella dominated the female microbiome, accounting for over 90% of sequences in Al Ja’ae and Al Jabeeb, while males displayed a more balanced microbial profile, with genera such as Corynebacterium, Staphylococcus, and Trueperella being prevalent. Pearson’s correlation and principal coordinate analyses (PCoA) indicated distinct sex-based microbial associations and clustering patterns. These findings underscore sex-associated disparities in the microbiome of H. dromedarii, suggesting potential implications for tick physiology and vector competence within the One Health framework.

Introduction

Ticks are obligate hematophagous chelicerate arachnids and are considered disease vectors transmitting the widest variety of diseases in the phylum Arthropoda¹. During feeding, ticks facilitate the transfer of infectious zoonotic diseases to humans and animals². Additionally, ticks and tick-borne diseases pose a global economic burden as they reduce livestock productivity and are costly to control. In Uganda alone, losses in animal production due to ticks and the cost of applying acaricides are estimated at 1.1 billion USD yearly³. Likewise, the annual cost to suppress the black-legged tick (Ixodes scapularis Say) ranges between 508 and 3,102 USD per household⁴.

Moreover, ticks host a variety of symbiotic microorganisms that play a vital role in the symbiont-host-microorganism complex interactions⁵. Tick endosymbionts could play a role in immunity, response to environmental stressors, nutritional biology, reproduction, fitness, and the transmissibility of certain tick-borne pathogens⁶. The complexity of the tick microbiome also includes microbiome-microbiome interactions, where microorganisms surviving in the same environment could express an exclusive, competitive, facilitative, or neutral relationships⁷. The microbiome of Hyalomma dromedarii, for instance, showed a positive correlation between Lysinibacillus and Bacillus, whereas a negative correlation was recorded between Francisella and Corynebacterium as well as Acinetobacter and Escherichia⁸. Although endosymbionts have been less studied than tick-borne pathogens, some endosymbiotic species have been identified as common in almost all ticks, such as Coxiella-like and Francisella-like endosymbionts inferring that the latter two are essential for tick survival⁹. Consequently, studying the tick microbiome and the factors affecting microbial diversity is essential.

Variation in tick microbial communities has been recorded as a function of other variables such as tick species, local environment, life stage¹⁰ blood meal host¹¹ time, seasonality¹² and tick gender. In male Dermacentor reticulatus, the genera Acidiphilium, Actinomycetospora, Cutibacterium, Pseudomonas, and Staphylococcus were significantly more abundant, whereas Brevundimonas, Candidatus, Aquirestis, Collinsella, Mycobacterium, Nakamurella, Nocardioides, and Rickettsia were significantly more abundant in females¹³. Gender-based disparities in tick microbiomes have also been recorded in Ixodes sp. where females I. ricinus and I. persulcatus were associated with Candidatus Midichlorii and Candidatus Lariskella, respectively. Moreover, Rickettsia presence was negatively correlated with endosymbiont abundance in female I. ricinus only¹⁴.

Studies on tick microbiomes have successfully employed next-generation sequencing (NGS) of bacterial 16S rRNA genes¹⁵. The reliability of 16S rRNA gene sequencing lies in its adequate size for proper identification, wide availability as a housekeeping gene in all bacteria, stability over time and feasibility of identifying bacteria to the genus level more than 90% of the time¹⁶.

With the tick microbiome playing an important role in the spread of tick-borne diseases⁶ it is important to study these microbiomes and the factors affecting them, especially in environmental conditions favorable for tick survival. Temperature has been shown to affect the life cycle of ticks including the camel tick (H. dromedarii). At higher temperatures, H. dromedarii ticks have shorter periods of adult oviposition and egg incubation as well as a faster life cycle¹⁷. In addition to temperature, host availability positively correlates with the abundance of some Hyalomma ticks¹⁸.

The United Arab Emirates (UAE), located on the Arabian Peninsula, is characterized by harsh climatological conditions including high summer temperatures¹⁹. Host-wise, camels are quite abundant and continuously increasing in number in the UAE. An average of 2.99 camels/ were recorded in the UAE in 2009 ²⁰ with the number of camels reaching 593,294 in 2023²¹. The abundance of livestock hosts and high summer temperature could create the ideal survivability conditions for H. dromedarii ticks. This is an additional incentive to better understand the microbial communities of H. dromedarii and the factors affecting them in the UAE.

Hyalomma dromedarii is considered a disease vector and associated with a variety of tick-borne diseases. For instance, in Riyadh, Saudi Arabia, both Rickettsia and Borrelia were recorded with an infection rate of 31% and 37%, respectively²². In the UAE, Candidatus, Rickettsia andeanae and Rickettsia endosymbionts were reported in camel ticks sampled in Al Ain in 2011 with a frequency of 1.12%, while Theileria annulata was recorded in samples taken from the same locality in 2010 and 2011 with a relative frequency of 2.3% and 1.6%, respectively²³. Furthermore, both Rickettsia sp. and Francisella sp. were found in camel ticks collected in Abu Dhabi with a relative abundance of 1.36% and 5.81%, respectively²⁴.

The microbiome of H. dromedarii has also been previously investigated in several regions. In Al Hfouf city of Saudi Arabia, the microbiome of the camel tick was dominated by Francisella endosymbionts (94.37%) with the main phyla being Proteobacteria (98%), Firmicutes (1.38%), Actinobacteria (0.36%) then Bacteroidetes (0.17%)²⁵. Moreover, the microbiome of camel ticks in Al Khotha and Al Gayed regions of Saudi Arabia were dominated by the phyla Proteobacteria (61.3% and 81.2% for Al Khotha and Al Gayed, respectively) and Firmicutes (31.2% and 9.2% for Al Khotha and Al Gayed, respectively)¹⁵. In the UAE, the microbiome of H. dromedarii has been studied in ticks collected in 2010 and 2019 from Al Ain city. The most dominant phyla were Proteobacteria, Bacteroidetes, Firmicutes, Actinobacteria, Cyanobacteria, Verrucomicrobia in and Planctomycetes in 2010 and Proteobacteria, Actinobacteria, Bacteroidetes, Firmicutes, Cynobacteria, Verrucomicrobia and Planctomycetesin in 2019⁸. Even though multiple studies investigated sexual dimorphism in tick microbiomes , limited data exists on the sex-based microbiome variation in H. dromedarii in the UAE with current published work limited to only Al Ain city²⁶. This paper utilizes 16S rRNA next-generation sequencing to provide a pioneering insight into sexual dimorphism in the microbiome of the camel tick H. dromedarii collected from three emirates in the UAE.

Results

Across all sampled ticks, a total of 432,495 reads were recorded, clustering into 848 operational taxonomic units (OTUs) at 97% similarity. In total, 20 phyla, 36 classes, 89 families, and 261 genera were identified. The diversity and abundance of taxa varied among the analyzed pools. In addition, rarefaction curves plateaued for all samples, indicating sufficient sequencing depth to capture most of the bacterial diversity present.

Al Khazna

In male ticks, 98,336 reads clustered into 11 phyla, 16 classes, 45 families, and 80 genera, whereas in females, 48,881 reads clustered into 5 phyla, 10 classes, 43 families, and 58 genera (Table 1). Taxonomic profiling revealed that Proteobacteria, Actinobacteriota, and Firmicutes were the most abundant phyla in both sexes, whereas Deinococcota, Gemmatimonadota, and Planctomycetota were the least abundant in males, and Fusobacteriota was the least abundant in females (Fig. 1).

Table 1.

Number of reads, OTUs, and taxonomic groups identified in Hyalomma dromedarii tick pools.

Pool name	Tick sex	Reads before denoising	Reads after denoising	Phyla	Classes	Families	Genera
Al Khazna	♂	128,238	98,336	11	16	45	80
	♀	61,796	48,881	5	10	43	58
Al Jabeeb	♂	68,346	56,347	19	35	127	219
	♀	97,421	84,213	7	8	28	38
Al Ja’ae	♂	77,342	63,484	14	25	90	141
	♀	89,914	81,234	11	16	50	61

Fig. 1.

Bacterial phyla identified in Hyalomma dromedarii tick pools collected from three locations: Al Khazna, Al Ja’ae, and Al Jabeeb.

At the class level, the most dominant taxa in males were Actinobacteria (66.92%), Clostridia (21.33%), and Bacilli (5.97%), while in females, Actinobacteria (55.75%), Gammaproteobacteria (38.70%), and Bacilli (4.20%) prevailed (Fig. 2). The predominant bacterial genera in males were Corynebacterium (46.38%), Trueperella (20.46%), and Anaerococcus (8.94%), whereas in females, Corynebacterium (54.11%), Francisella (38.31%), and Staphylococcus (3.56%) dominated (Fig. 3).

Fig. 2.

Bacterial classes identified in Hyalomma dromedarii tick pools collected from three locations: Al Khazna, Al Ja’ae, and Al Jabeeb.

Fig. 3.

Bacterial genera identified in Hyalomma dromedarii tick pools collected from three locations: Al Khazna, Al Ja’ae, and Al Jabeeb.

Al Ja’ae

In Al Ja’ae, male ticks yielded 63,484 reads, clustering into 14 phyla, 25 classes, 90 families, and 141 genera, while female ticks had 81,234 reads, clustering into 11 phyla, 16 classes, 50 families, and 61 genera (Table 1).

The most dominant phyla in both sexes were Actinobacteriota, Firmicutes, and Proteobacteria, while the least abundant phyla in males were Sumerlaeota, Euryarchaeota, and Cyanobacteria, and in females, Patescibacteria, Gemmatimonadota, and Planctomycetota (Fig. 1).

At the class level, Bacilli (40.77%), Actinobacteria (30.54%), and Gammaproteobacteria (22.71%) were dominant in males, whereas females were overwhelmingly dominated by Gammaproteobacteria (93.77%) (Fig. 2). At the genus level, the most abundant bacteria in males were Staphylococcus (25.48%), Corynebacterium (23.85%), and Francisella (19.28%), whereas in females, Francisella (93.10%) was the most prevalent, followed by Staphylococcus (2.14%) and Corynebacterium (0.88%) (Fig. 3).

Al Jabeeb

Similarly, male and female ticks from Al Jabeeb exhibited distinct microbial compositions. In males, 56,347 reads clustered into 19 phyla, 35 classes, 127 families, and 219 genera, whereas in females, 84,213 reads clustered into 7 phyla, 8 classes, 28 families, and 38 genera (Table 1).

The dominant phyla in both sexes were Firmicutes, Actinobacteriota, and Proteobacteria, whereas the least abundant phyla in males were Patescibacteria, Retaria, and Acidobacteriota, and in females, Deinococcota, Cyanobacteria, and Myxococcota (Fig. 1).

At the class level, Bacilli (38.92%), Gammaproteobacteria (22.46%), and Actinobacteria (20.49%) were most prevalent in males, while Gammaproteobacteria (98.28%) dominated females (Fig. 2). Further analysis at the genus level showed that in males, Staphylococcus (23.96%), Francisella (17.84%), and Corynebacterium (15.12%) were predominant, whereas in females, Francisella (98.07%) was overwhelmingly dominant (Fig. 3).

Prevalence of the main bacterial genera

Corynebacterium

Corynebacterium was the most prevalent genus in both male (46.38%) and female (54.11%) ticks from Al Khazna. It ranked second in males from Al Ja’ae (23.85%) and in females from Al Jabeeb (0.38%), while being the third most common genus in females from Al Ja’ae (0.88%) and in males from Al Jabeeb (15.12%) (Fig. 4).

Fig. 4.

Relative abundance of Corynebacterium in H. dromedarii tick pools collected from three locations: Al Khazna, Al Ja’ae, and Al Jabeeb.

Francisella

Francisella was the most dominant genus in female ticks from Al Ja’ae (93.1%) and Al Jabeeb (98.07%). It ranked second in males from Al Jabeeb (17.84%) and in females from Al Khazna (38.31%). Across all locations, Francisella was consistently more abundant in females than in males (Fig. 5).

Fig. 5.

Relative abundance of Francisella in H. dromedarii tick pools collected from three locations: Al Khazna, Al Ja’ae, and Al Jabeeb.

Staphylococcus

Staphylococcus was the second most abundant genus in males from Al Ja’ae (25.48%) and Al Jabeeb (23.96%). In females, it ranked second in Al Ja’ae (2.14%) and third in Al Khazna (3.56%) and Al Jabeeb (0.36%). Overall, Staphylococcus was more prevalent in males than in females across all locations (Fig. 6).

Fig. 6.

Relative abundance of Staphylococcus in Hyalomma dromedarii tick pools collected from three locations: Al Khazna, Al Ja’ae, and Al Jabeeb.

Trueperella

Trueperella was the second most abundant genus in males from Al Khazna (20.46%) but was detected at a much lower percentage in females from the same location (1.33%). It was entirely absent in both sexes from Al Ja’ae and in female ticks from Al Jabeeb, with only a minor presence in males from Al Jabeeb (Table 2).

Table 2.

Relative abundance of Trueperella in all H. dromedarii tick pools.

Pool name	Al Khazna		Al Ja’ae		Al Jabeeb
Tick sex	♀	♂	♀	♂	♀	♂
%Trueperella	1.33	20.46	0	0	0	0.08

Diversity indices genus

Diversity indices were compared between male and female H. dromedarii pools collected from Al Khazna, Al Jabeeb, and Al Ja’ae. Although none of the differences reached statistical significance at the 0.05 level, male pools consistently exhibited higher microbial diversity across all metrics (Table 3). The mean richness was notably greater in males (146.67 ± 40.23) compared to females (52.33 ± 7.22), and Shannon diversity followed a similar pattern (2.47 ± 0.36 vs. 0.55 ± 0.27). Dominance was higher in females (0.757 ± 0.160) than in males (0.185 ± 0.046), indicating a less even microbial community in female ticks. Evenness was also greater in males (0.0876 ± 0.0061) compared to females (0.0353 ± 0.0074). All datasets passed the Shapiro–Wilk normality test (p > 0.05), validating the use of paired t-tests for statistical comparison.

Table 3.

Comparison of diversity indices between male and female Hyalomma dromedarii pools collected from three locations in Abu dhabi, UAE (Al khazna, al jabeeb, and al Ja’ae).

Indices	Male	Female	Statistic (Paired t-test)	p Value	Normality test (Shapiro-Wilk, p Value)
Richness	146.67 ± 40.23	52.33 ± 7.22	2.03	0.179	0.704
Shannon	2.47 ± 0.36	0.55 ± 0.27	3.06	0.092	0.784
Dominance	0.185 ± 0.046	0.757 ± 0.160	-2.78	0.109	0.379
Evenness	0.0876 ± 0.0061	0.0353 ± 0.0074	4.23	0.052	0.327

Values represent means ± standard errors. Statistical comparisons were made using paired t-tests. Normality of the data was assessed using the Shapiro–Wilk test*.

* For the Shapiro–Wilk test, a high p-value (> 0.05) indicates that the data are normally distributed.

Association between the main detected bacterial genera

Pearson’s correlation analysis revealed several significant positive associations between bacterial genera (Fig. 7). Corynebacterium exhibited strong correlations with Trueperella, Anaerococcus, Peptoniphilus, and Fusobacterium. Similarly, Trueperella showed significant correlations with Anaerococcus and Fusobacterium. Staphylococcus was positively correlated with Bacillus, while Anaerococcus demonstrated correlations with Peptooniphilus and Fusobacterium. These associations suggest potential microbial interactions within the tick microbiome.

Fig. 7.

Correlation matrix of major bacterial genera in Hyalomma dromedarii tick pools. Blue circles indicate positive correlations, red circles indicate negative correlations, and boxes denote statistically significant correlations.

PCoA analysis

Overall, this analysis showed central clustering of Anaerococcus, Bacillus, Fusobacterium, Peptoniphilus, Romoutsia, Rothia, Staphylococcus and Trueperella (Fig. 8). Specifically, the clustering patterns revealed Corynebacterium and Trueperella grouped in the second quadrant, Anaerococcus, Bacillus, Fusobacterium, Peptoniphilus, Romoutsia, Rothia, and Staphylococcus in the third quadrant, and Francisella occupying the fourth quadrant alone. The analysis also suggested overall clustering patterns highlighting sex-based disparities (Fig. 9). It revealed distinct clustering by sex and location: female and male pools from Al Jabeeb and Al Ja’ae form separate clusters (first quadrant and third quadrant, respectively), indicating sex-based differences in microbial composition. In contrast, male and female pools from Al Khazna cluster together (second quadrant).

Fig. 8.

Principal coordinates analysis (PCoA) of correlated bacterial genera in Hyalomma dromedarii tick pools. The plot shows distinct clustering patterns among bacterial genera. These spatial groupings suggest that genera within the same quadrant may share ecological niches or functional roles within the tick microenvironment, reflecting co-occurrence shaped by similar physiological conditions or transmission pathways.

Fig. 9.

Principal coordinates analysis (PCoA) of microbial communities in Hyalomma dromedarii tick pools. The plot reveals distinct clustering by sex and location: female and male pools from Al Jabeeb and Al Ja’ae form separate clusters (Q1 and Q3, respectively), indicating sex-based differences in microbial composition. In contrast, male and female pools from Al Khazna cluster together (Q2), suggesting a stronger geographic influence on microbiota at that site.

Discussion

In this study, 16S rRNA gene sequencing was used to investigate the sex-based disparities in the microbiome of male and female ticks sampled from three different localities in the UAE. We noticed that male ticks had higher richness, Shannon diversity, and evenness indices, along with lower dominance values, indicating greater microbial diversity and community balance in males when compared to their female counterparts sampled from the same region. This work demonstrates compositional differences in the microbiome of male and female H. dromedarii ticks although the observed differences in diversity indices were not statistically significant (p > 0.05). However, the consistent trends across all indices suggest potential biological relevance. In addition, certain genera were completely absent or more prevalent in one sex compared to the other, supporting sex-specific variation in microbial community composition.

The high relative abundance of Corynebacterium has been previously recorded in ticks of the genus Hyalomma. Corynebacterium is the most abundant genus in Hyalomma truncatum in South Africa (35.6%)²⁷, and the second most abundant in Hyalomma anatolicum in the UAE (41.5%)²⁸. In H. dromedarii, Corynebacterium was one of the main genera found in the microbiome of ticks from the Al Gayed area in Saudi Arabia with a relative abundance of 0.6%²⁶. In the UAE, Corynebacterium was also detected with high relative abundance in ticks sampled from Al Ain (38.2%)²⁹ and was the second most common genus (36.62%) in ticks sampled from the eastern UAE⁸. Corynebacterium was more abundant in males in Al Ja’ae and Al Jabeeb and females in Al Khazna. This could be linked to variation in feeding duration and host species³⁰ especially since H. dromedarii can function as a two-host or three-host tick³¹. In general, genus Corynebacterium encompasses species normally existing on animal skin and opportunistic pathogens that cause zoonotic diseases³².

Likewise, Staphylococcus was more common in males across all locations. This genus promotes the reproductive fitness of ticks³³ which could explain its abundance in males as the sampling coincided with the reproductive season of ticks. Furthermore, the previous detection of the pathogenic Staphylococcus lentus and Staphylococcus saprophyticus in Hyalomma ticks³⁴ suggests that a part of the detected Staphylococcus in this study could be pathogenic. Staphylococcus has been previously documented in Hyalomma ticks making up 90% of the microbiome of H. analoticum in South India³⁵ and 57.6% of the microbiome of H. analoticum in the UAE²⁸. In H. dromedarii, Staphylococcus was among the most common genera detected in Al Gayed (3.85%), Al Khotha (22.83%)¹⁵ and Al Hofuf (0.51%) in the Kingdom of Saudi Arabia²⁵. Moreover, Staphylococcus was common in camel ticks sampled from Al Ain city, UAE throughout the year reaching its peak relative abundance of 53.1% in July 2019 ³¹.

Unlike the previous two genera, Francisella was found at higher percentages in females across all sampling locations. This could be explained by the transovarial or vertical transmission of Francisella³⁶ and preference of Francisella for infecting female gonads over male gonads in certain hard ticks³⁷. Nevertheless, Francisella has been found in all tick species suggesting that it is essential for tick survivability⁹. The high relative abundance of Francisella was consistent with the literature. Ticks of the genus Hyalomma have been recorded to have a high relative abundance of Francisella sp. In Caceres province, southwestern Spain, the microbial community of H. lusitanicum was dominated by Francisella sp. with a relative abundance ranging from 52 to 99% in the five tested pools³⁸. In Hyalomma anatolicum sampled from Sistan and Baluchestan, Iran, Francisella made up more than 92% of all OTU reads³⁹. As for the camel tick H. dromedarii, the family Francisellaceae was common in ticks sampled from Al Gayed region (2.52%) and Al Khotha region (46%)¹⁵. Moreover, Francisella was the second most common genus in ticks sampled from Al Ain, UAE throughout the year reaching its highest relative abundance of 79.4% in September²⁹.

On the contrary, Trueperella was not more prevalent in one gender compared to the other. Trueperella sp. are opportunistic pathogens naturally found in the urogenital tract, upper respiratory tract and skin of animals. Nevertheless, little is known about the pathology of this genus⁴⁰. The limited detection of Trueperella in this study aligns with previously published work. This genus was found in lower abundance in genus Hyalomma. For instance, in H. analoticum sampled from the UAE, Trueperella made up 6 to 22% of the tick’s microbiome depending on seasonality²⁸ and 3.7% of the microbiome of H. truncatum in South Africa²⁷. As for H. dromedarii, Trueperella was detected in the UAE in 2019 with a relative abundance of 0.23%, 19.81% and 7% in ticks collected in June, October and December, respectively²⁹.

In all three locations, the microbiome of males showed higher genus richness. The observed richness in males’ microbiomes is consistent with the literature. In Ixodes scapularis ticks collected from Massachusetts and Texas, USA, males had higher microbial diversity than females⁴¹. Likewise, Haemaphysalis longicornis from Mengyin county, Shandong Province, China showed higher species richness in males when compared to female ticks³⁰. In contrast, our findings on H. dromedarii, differ from earlier research in the UAE, which reported richer microbiomes in female ticks²⁶. These differences may stem from variation in sampling design as the previous study pooled ticks from various time points and locations, potentially introducing spatial and temporal variability into the microbial profiles.

As for the statistical correlation, all significant correlations recorded in this study were positive. Despite previous reports on negative correlations between Francisella and other genera in the microbiome of H. dromedarii in the UAE in previous work, the majority of correlations reported were positive⁸. The Principal Coordinate Analysis (PCoA) showed central clustering of Anaerococcus, Bacillus, Fusobacterium, Peptoniphilus, Romoutsia, Rothia, Staphylococcus and Trueperella indicating higher similarity in their compositional profiles indicating that their positive correlation could be due to their commensal relationship with the tick, mutualistic relationship to one another or both. On the other hand, Francisella and Corynebacterium are plotted distinctly from each other. This is due to their inversely proportional dominance in the microbiome of tested ticks. Despite the detection of slight overlap in the microbiome of males and females, the performed PCoA shows a clustering pattern highlighting sex-based compositional differences in the microbiome of the camel tick.

Our investigation reveals sex-based differences in the microbiome of H. dromedarii ticks collected from camels in the UAE. Male ticks consistently exhibited higher microbial diversity and genus richness across all sampled locations, whereas female ticks were predominantly dominated by Francisella species. Understanding these differences is crucial for developing targeted strategies for tick-borne disease control and highlights the importance of considering sex as a biological variable in vector microbiome studies.

Methods

Sampling

A total of 300 ticks were collected from 30 camels, with 10 ticks sampled per camel. Sampling was conducted on farms across three locations—Al Khazna, Al Je’ea, and Al Jabeeb—between June and September 2022 (Fig. 10). The three locations were selected because they are major camel-rearing areas in Abu Dhabi, each with a high concentration of camel farms. Together, they provide a representative overview of camel husbandry practices across the region, making them suitable for studying the microbiome of H. dromedarii ticks associated with camels in different management settings. To assess the effect of tick gender on the microbiome, only farms rearing camels were selected, and sampling at each location was conducted on the same day to minimize temporal variation. Ticks were collected from five camels per farm, with two farms sampled per location. The ticks were placed in 50-mL sterile tubes and transported to the entomology laboratory at the College of Sciences, United Arab Emirates University (UAEU), Al Ain City, Emirate of Abu Dhabi, UAE. Sampling followed the guidelines of the Animal Research Ethics Committee at UAEU, and samples were stored at − 80 °C until processing. One semi-engorged female tick was identified and selected from each camel for DNA extraction and pooling. Tick collection was carried out in strict accordance with the recommendations of the Animal Research Ethics Committee (A-REC) of the UAEU (ethical approval no. ERA_2022_1629). In addition, the experimental protocol was approved by the Research Office at UAEU.

Fig. 10.

Map of sampling locations for Hyalomma dromedarii ticks collected from camels in the United Arab Emirates. Map produced using QGIS Desktop 3.36.2 (https://qgis.org) (Zürich) and Basemap imagery © Esri, Maxar, Earthstar Geographics, and the GIS User Community (https://www.arcgis.com/home/webmap/viewer.html).

Tick identification

Hyalomma dromedarii ticks were identified both morphologically and molecularly. For morphological identification, ticks were examined under a stereoscope following guidelines from the literature^31,42. Hyalomma dromedarii ticks are primarily characterized by pale rings on their legs, with males having parallel and strongly curved margins on the adanal plates, while females possess a V-shaped posterior lip on their genital pores.

Molecular confirmation was performed using DNA sequencing of the cytochrome oxidase subunit I (COI) gene²³. The gene was amplified via PCR (Polymerase Chain Reaction) using the primer pair Fish1F: 5′-TCAACCAACCACAAAGACATTGGCAC-3′ and Fish1R: 5′-TAGACTTCTGGGTGGCCAAAGAATCA-3′⁴³. The PCR was conducted under the following thermal cycling conditions: initial denaturation at 95 °C for 2 min, followed by 30 cycles of 94 °C for 1 min, 54 °C for 1 min, and 72 °C for 90 s.

DNA extraction, quality control and pooling

Ticks were individually washed with distilled water and 70% ethanol to minimize external contamination. They were then placed in 1.5-mL centrifuge tubes and cut into four equal pieces using sterile scissors. Samples were homogenized, and DNA was extracted using the QIAamp Tissue Kit (Qiagen, Hilden, Germany) following the manufacturer’s protocol. The quality of the extracted DNA was assessed using a spectrophotometer (NanoDrop ND-2000, Erlangen, Germany). Extracted DNA was pooled into six groups based on location and gender: Al Khazna – males, Al Khazna – females, Al Jabeeb – males, Al Jabeeb – females, Al Je’ea – males, and Al Je’ea – females (Table 4).

Table 4.

Pooling of Hyalomma dromedarii ticks.

Pools	Pool 1	Pool 2	Pool 3	Pool 4	Pool 5	Pool 6
Components	5 male ticks	5 female ticks	5 male ticks	5 female ticks	5 male ticks	5 female ticks
Sampling location	Al Khazna	Al Khazna	Al je’ea	Al je’ea	Al Jabeeb	Al Jabeeb

Next generation sequencing of the 16 S rRNA and bioinformatic computations

All six pools were sent to Macrogen Inc. (Seoul, South Korea) for 16S rRNA gene sequencing using the Illumina MiSeq platform. For this purpose, the V3-V4 region of the 16S rRNA gene was amplified using the primers Bakt_341F: CCTACGGGNGGCWGCAG and Bakt_805R: GACTACHVGGGTATCTAATCC70⁴⁴ along with Herculase II Fusion DNA polymerase Nextera XT Index Kit V2. Microbiome data were processed using QIIME 2 (version 2024.2). Raw paired-end sequence files were first imported into QIIME 2. Adapter trimming was performed using the cutadapt trim-paired plugin. The trimmed output was saved as paired-end-demux-trimmed.qza and quality-checked using qiime demux summarize. Denoising and error correction were carried out using the dada2 denoise-paired method. The output included feature tables and representative sequences, saved to a specified output directory. Taxonomic classification was performed using a pre-trained Naive Bayes classifier trained on the SILVA 138 99% OTUs reference database. The classifier was applied to the representative sequences to assign taxonomy, and the results were tabulated for inspection. Taxonomic composition was visualized using bar plots generated via the qiime taxa barplot plugin, incorporating sample metadata for grouping. In all performed analyses we used Phred quality score 20. Following this, taxonomic data from phylum to genus levels were downloaded as Excel spreadsheets from the QIIME 2 barplot visualization files (.qzv) using the “Export” feature. These data were further analyzed using the software PAST version 5.2.2 (2025) for statistical analysis and diversity comparisons. Rarefaction analysis was performed to assess sequencing depth adequacy and to compare species richness across male and female H. dromedarii tick pools from three locations in Abu Dhabi, UAE. Curves were generated using the number of observed taxa (S) plotted against the number of sequencing reads (specimens) for each sample. This approach allowed evaluation of whether sampling captured the majority of microbial diversity within each pool and enabled comparisons of richness independent of sequencing depth. Raw 16S rRNA sequencing reads (FASTQ format) were deposited in NCBI Sequence Read Archive as a BioProject (PRJNA1273452).

Statistical calculations and visualizations

The t-tests, Shapiro-Wilk, and biodiversity indices were calculated using PAST version 4.17 (Paleontological statistics software package). As for data visualization, the sampling site map was produced using QGIS, correlation matrix and PCoA were plotted using PAST version 4.17, stacked column charts and bar charts were created using Microsoft PowerPoint.

Author contributions

Conceptualization: MAA. Methodology: MAA, SBM, RV. Investigation: MTA, ASA. Formal analysis: MTA, MAA, ASA, RV. Visualization: MTA. Supervision: MAA. Project administration: MAA. Funding acquisition: MAA and SBM. Writing—original draft: MTA (lead) and MAA. Writing—review & editing: MAA and MTA.

Funding

The funding for this study was provided by the UAE University through UPAR Grant no. G00003718.

Data availability

All data generated or analyzed during the current study are available in the paper and the GenBank under the following BioProject: PRJNA1273452 and BioSample accessions SAMN48946900, SAMN48946901, SAMN48946902, SAMN48946903, SAMN48946904, SAMN48946905.

Declarations

Competing interests

The authors declare no competing interests.