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Open Access

Peer-reviewed

Research Article

Molecular epidemiology and risk factors of Anaplasma spp., Babesia spp. and Theileria spp. infection in cattle in Chongqing, China

  • Zuoyong Zhou ,

    Contributed equally to this work with: Zuoyong Zhou, Kai Li, Yingying Sun

    Roles Supervision, Writing – original draft, Writing – review & editing

    Affiliations College of Animal Science, Rongchang Campus of Southwest University, Chongqing, China, Veterinary Science Engineering Research Center of Chongqing, Chongqing, China

    ⨯
  • Kai Li ,

    Contributed equally to this work with: Zuoyong Zhou, Kai Li, Yingying Sun

    Roles Investigation, Writing – original draft

    Affiliation College of Animal Science, Rongchang Campus of Southwest University, Chongqing, China

    ⨯
  • Yingying Sun ,

    Contributed equally to this work with: Zuoyong Zhou, Kai Li, Yingying Sun

    Roles Investigation, Writing – original draft

    Affiliation College of Animal Science, Rongchang Campus of Southwest University, Chongqing, China

    ⨯
  • Junge Shi,

    Roles Writing – original draft

    Affiliation College of Animal Science, Rongchang Campus of Southwest University, Chongqing, China

    ⨯
  • Hexian Li,

    Roles Writing – original draft

    Affiliation College of Animal Science, Rongchang Campus of Southwest University, Chongqing, China

    ⨯
  • Yiwang Chen,

    Roles Writing – original draft

    Affiliation College of Animal Science, Rongchang Campus of Southwest University, Chongqing, China

    ⨯
  • Haoyue Yang,

    Roles Investigation

    Affiliation College of Animal Science, Rongchang Campus of Southwest University, Chongqing, China

    ⨯
  • Xiao Li,

    Roles Investigation

    Affiliation College of Animal Science, Rongchang Campus of Southwest University, Chongqing, China

    ⨯
  • Bi Wu,

    Roles Investigation

    Affiliation College of Animal Science, Rongchang Campus of Southwest University, Chongqing, China

    ⨯
  • Xiaoxia Li,

    Roles Investigation

    Affiliation College of Animal Science, Rongchang Campus of Southwest University, Chongqing, China

    ⨯
  • Zhiying Wang,

    Roles Writing – review & editing

    Affiliations College of Animal Science, Rongchang Campus of Southwest University, Chongqing, China, Veterinary Science Engineering Research Center of Chongqing, Chongqing, China

    ⨯
  • Fangjun Cheng ,

    Roles Supervision, Writing – review & editing

    2323005161@qq.com (FC); chhjj006@163.com (SH)

    Affiliations College of Animal Science, Rongchang Campus of Southwest University, Chongqing, China, Veterinary Science Engineering Research Center of Chongqing, Chongqing, China

    ⨯
  • Shijun Hu

    Roles Supervision, Writing – review & editing

    2323005161@qq.com (FC); chhjj006@163.com (SH)

    Affiliations College of Animal Science, Rongchang Campus of Southwest University, Chongqing, China, Veterinary Science Engineering Research Center of Chongqing, Chongqing, China

    ⨯

Correction

14 Aug 2019: The PLOS ONE Staff (2019) Correction: Molecular epidemiology and risk factors of Anaplasma spp., Babesia spp. and Theileria spp. infection in cattle in Chongqing, China. PLOS ONE 14(8): e0221359. https://doi.org/10.1371/journal.pone.0221359 View correction

Abstract

Tick-borne pathogens (TBPs) seriously affect cattle production and can be economically damaging. The epidemiology of these organisms in the Chongqing municipality of China is not well described. This study aimed to investigate the prevalence and risk factors of TBPs including Anaplasma spp., Babesia spp. and Theileria spp. in cattle in Chongqing municipality. The results showed that 43.48% (150/345) of cattle were infected with at least one TBP, of which single infections were detected in 104 (30.14%), double infections in 34 cattle (9.86%) and triple infections in 12 (3.48%) of the cattle. The overall prevalence of Anaplasma spp., Theileria spp. and B. bigemina were 22.32%, 23.19% and 7.24%, respectively. Among these, the prevalence of A. bovis, A. central, A. phagocytophilum, A. platys, A. marginale, T. sinensisi and T. orientalis were 8.41%, 7.83%, 4.93%, 4.35%, 2.61%, 22.32% and 2.60%, respectively. We could not detect B. bovis, T. annulata, T. luwenshuni or T. uilenbergi in cattle. Cattle ≥1-year-old were more likely to be infected with Theileria spp. [adjusted odd ratio (AOR) = 2.70, 95% CI = 1.12–6.56)] compared with younger cattle, while cattle ≥1-year-old had reduced susceptibility to B. bigemina (AOR = 0.14, 95% CI = 0.03–0.60). Cattle living at higher altitude (≥500 m) were more susceptible to B. bigemina (AOR = 6.97, 95% CI = 2.08–23.35) and Theileria spp. infection (AOR = 1.87, 95% CI = 1.06–3.32). The prevalence of Theileria spp. on farms with cats was significantly higher than that without cats (AOR = 2.56, 95% CI = 1.12–5.88). Infection with A. bovis and A. central were significantly associated with A. phagocytophilum infection. Furthermore, there were significant associations between A. bovis and A. central infection, T. sinensisi and A. marginale infection, and B. bigemina and T. orientalis infection. This study provides new data on the prevalence of Anaplasma spp., Babesia spp. and Theileria spp. in cattle in Chongqing, and for the first time we reveal a possible relationship between the afore-mentioned pathogens, which will help in formulating appropriate control strategies for these pathogens in this area.

Citation: Zhou Z, Li K, Sun Y, Shi J, Li H, Chen Y, et al. (2019) Molecular epidemiology and risk factors of Anaplasma spp., Babesia spp. and Theileria spp. infection in cattle in Chongqing, China. PLoS ONE 14(7): e0215585. https://doi.org/10.1371/journal.pone.0215585

Editor: Gordon Langsley, Institut national de la santé et de la recherche médicale - Institut Cochin, FRANCE

Received: October 16, 2018; Accepted: April 4, 2019; Published: July 15, 2019

Copyright: © 2019 Zhou et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Data Availability: All relevant data are within the manuscript and its Supporting Information files.

Funding: National Key Research and Development Program of China (2018YFD0501705), Chongqing Social Undertakings and Livelihood Security Technology Innovation Projects (CSTC2015SHMSZX80020) provided funding for this project. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Competing interests: The authors have declared that no competing interests exist.

Abbreviations: TBPs, tick-borne pathogens; AOR, adjusted odd ratio; 95% CI, 95% confidence intervals (CI); nPCR, nested PCR; ID, initial denaturation; FE, final extension; APH, A. phagocytophilum; AC, A. central; AB, A. bovis; AM, A. marginale; APL, A. platys; TS, T. sinensisi; TO, T. orientalis; BB, B. bigemina

Introduction

Tick-borne pathogens (TBPs) have always attracted the attention of researchers, not only for their damaging influence upon livestock production but also for their public health threat [1]. Among the tick-borne diseases, anaplasmosis, babesiosis and theileriosis are the most important and are distributed widely. These organisms affect cattle worldwide [2]. Till now, five Anaplasma pathogens (A. marginale, A. bovis, A. centrale, A. phagocytophilum, and A. platys) have been reported to cause bovine anaplasmosis, of which A. phagocytophilum has been shown to infect a variety of animals and humans [3–5]. Two mainly Babesia pathogens, Babesia bovis and B. bigemina, were found responsible for bovine babesiosis [6], and three species of Theileria including T. annulata, T. sinensis, and T. orientalis (also named T. sergenti) were the main causative agents of bovine theileriosis [7–10], and recently, T. luwenshuni has also been detected in blood samples from cattle and yaks [9].

Numerous studies have reported the infection and prevalence of Anaplasma spp., Babesia spp. and Theileria spp. in cattle across many countries [4,5,11–16]. In China, there have also been many studies [9,17–22]. However, these studies usually focus on single pathogen infections, and records on pathogen co-infections, the risk factors, and the mutual influence of each pathogen in cattle are absent. In addition, studies relating to the aforementioned pathogens in cattle in China have mainly been restricted to the northwest region, while the information is very limited for southwest China.

The total number of cattle approximated 300 million at the end of 2015 in Chongqing, and is one of the economic pillars of animal husbandry in this city. However, the prevalence of Anaplasma spp., Babesia spp., and Theileria spp. in cattle in this area is unclear. The objectives of this study were 1) to detect Anaplasma spp., Babesia spp., and Theileria spp. in cattle in Chongqing, 2) to analyze the risk factors for infection of Anaplasma spp., Babesia spp., and Theileria spp., and 3) to evaluate the associations of the aforementioned pathogens in cattle in Chongqing.

Materials and methods

Study area

Chongqing municipality is located in the southwest of China, between the northern latitudes of 28.10°–32.13°, and eastern longitudes of 105.11°–110.11°. Its altitude ranges between 73.1 m at the Yangtze River in Wushan and 2796.8 m at Liangshan peak in Wuxi. The climate tends to be subtropical, with a monsoon/humid climate and has an average annual temperature of 16–18°C.

Blood sample collection and DNA extraction

Three hundred and forty five sodium citrate anticoagulated blood samples were collected from 10 ranches located in Tongnan, Rongchang, Jiangjing, Changshou, Liangping, Kaizhou, Yunyang, Wushan, Fuling, and Qianjiang, from May 2016 to April 2017. The ranches were selected based on the number of cattle (≥50) and convenience of sampling. The sampled animals were randomly selected from apparently healthy cattle, and the information including gender and age of cattle, as well as the altitude and the existence of cats in ranches were recorded. The blood samples were sent back to the laboratory within an ice box. Whole blood genome was extracted using a Wizard Genomic extraction kit (Promega, Madison, WI, USA) according to the manufacturer's instructions. This study was approved by the Ethics Committee of Southwest. Consent was obtained from cattle owners before the collection of blood samples from their cattle by an experienced, practicing veterinarian.

PCR detection of Anaplasma spp., Babesia spp., and Theileria spp.

Anaplasma spp. (A. bovis, A. central, A. marginale, A. phagocytophilum and A. platys), Babesia spp.(B. bovis and B. bigemina) and Theileria spp. (T. annulata, T. sinensis, T. orientalis, T. luwenshuni and T. uilenbergi) infections were detected by PCR or nested PCR using the primers reported in previous studies [7,23–29], the detail of primers can be found in S1 Table. The primers were synthesized by Bioligo Biotechnology Co., Ltd (Shanghai, China). The PCRs were performed according to the amplification programs in Table 1, with a volume of 12.5 μL in the reaction system including: 6.25 μL Premix Taq (containing TaKaRa Taq, dNTP Mixture and Taq Buffer) (Takara Dalian, China), 0.5 μL of each forward and reverse primer (20 μmol/L), 1 μL whole blood genome and 4.25 μL ddH2O. The amplified PCR products were photographed after electrophoresis in 1% agarose gels. The PCR amplification product were randomly selected for sequencing to verify the reliability of test.

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Table 1. Overall prevalence of Anaplasma spp., B. bigeminaand Theileria spp. infection in cattle in Chongqing of southwest China.

https://doi.org/10.1371/journal.pone.0215585.t001

Risk factor analysis

Multivariable logistic regression was performed in SPSS for Windows (18.0 version, SPSS Inc., Chicago, IL, USA) to analyze factors associated with aforementioned infections. Adjusted odd ratios (AOR) and 95% confidence intervals (CI) were calculated. A p-value of <0.05 was considered statistical significant.

Results

Prevalence of Anaplasma spp., Babesia spp., and Theileria spp. infection

A total of 345 cattle in Chongqing were included in this study. Detailed information pertaining to infection is shown in Table 1 and Fig 1. The results showed that 43.48% (150/345) of cattle were infected with at least one TBP, of which single infections were detected in 104 (30.14%), double infections in 34 cattle (9.86%) and triple infections in 12 (3.48%) of the cattle. The overall prevalence of Anaplasma spp., Theileria spp., and B. bigemina in cattle were 22.32% (77/345), 23.19% (83/345), and 7.24% (25/345), respectively. Among the Anaplasma spp. detected, A. bovis (29/345, 8.41%) was the most prevalent species recorded, followed by A. central (27/345, 7.83%), A. phagocytophilum (17/345, 4.93%), and A. platys (15/345, 4.35%), while infection with A. marginale (9/345, 2.61%) was the lowest. Among the Theileria spp., T. sinensisi and T. orientalis infections in cattle were 22.32% (77/345) and 2.60% (9/345), respectively. In addition, we could not detect B. bovis, T. annulata, T. luwenshuni, or T. uilenbergi in this study.

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Fig 1. An UpSetR plot of Anaplasma spp., Babesia spp. and Theileria spp. infection in cattle from Chongqing of southwest China (n = 345).

The blue horizontal coordinate columns represent the number of positive samples infected by pathogens. The purple vertical coordinate columns represent the number of positive samples infected by a single pathogen or multiple pathogens. The black dots represent the corresponding pathogens, and the dots connected by lines indicate co-infection of pathogens.

https://doi.org/10.1371/journal.pone.0215585.g001

Risk factor analysis based on blood sample data

The prevalence of Anaplasma spp. (25.13%) and Theileria spp. (27.27%) in male cattle was significantly higher than that in females (Anaplasma spp.: AOR = 2.18, 95% CI = 1.05–4.52; Theileria spp.: AOR = 3.27, 95% CI = 1.47–7.25). Cattle ≥ 1-year were more likely to be infected with Theileria spp. than cattle aged below 1-year of age (25.00% vs. 20.99%), and the difference was statistically significant (AOR = 2.70, 95% CI = 1.12–6.56). In contrast, cattle ≥1-year had a lower risk of B. bigemina infection (AOR = 0.14, 95% CI = 0.03–0.60). Ranches at an altitude ≥500 m was found to be a risk factor for B. bigemina (AOR = 6.97, 95% CI = 2.08–23.35) and Theileria spp. infection (AOR = 1.87, 95% CI = 1.06–3.32). With exception of Theileria spp. (AOR = 2.56, 95% CI = 1.12–5.88), there were no significant associations between presence of cats and infection with Anaplasma spp., or B. bigemina, (Table 2).

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Table 2. Multivariate analysis of selected factors and their association with Anaplasma spp., B. bigeminaand Theileria spp. infection in cattle in Chongqing of southwest China.

https://doi.org/10.1371/journal.pone.0215585.t002

Risk factor analysis based on pathogen co-infection

In order to evaluate the effect of specific pathogen infections and how they influence other pathogen infections within the same host, we considered each tested pathogen species as a potential risk factor in the analysis. The results of correlation analyses between each species of pathogen in infected cattle in Chongqing, are shown in Table 3. Infection with A. bovis and A. central were significantly associated with A. phagocytophilum infection, and A. phagocytophilum was more likely to increase the risk of A. central infection (AOR = 3.80, 95% CI = 1.10–13.18, p = 0.035). However, A. central was less likely to impact upon infection with A. phagocytophilum (AOR = 3.50, 95% CI = 0.97–12.59, p = 0.055). Furthermore, there was a significant association between A. bovis and A. central infection, T. sinensisi and A. marginale infection, and B. bigemina and T. orientalis infection (p<0.05). There were no significant associations between other pathogens that we aimed to identify.

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Table 3. Correlation analysis based on species of Anaplasma spp., Babesia spp. and Theileria spp. infection in cattle in Chongqing, southwest China.

https://doi.org/10.1371/journal.pone.0215585.t003

Discussion

For the first time, this systematic study investigated the epidemiology of Anaplasma spp., Babesia spp. and Theileria spp. infection in cattle in Chongqing, China. The infection rate of Anaplasma spp. in our study was lower than that reported in Algeria [5] and in Tunisia [16], but higher than that reported in northwest China [30]. The prevalence of A. bovis (8.41%) in cattle in Chongqing was higher than that of cattle reported in other locations, where the prevalence varied from 3.9% to 6.2% [5,16,18,30]. In contrast, the prevalence of A. centrale (7.83%) was lower than that of cattle in previous studies (range between 12.1%-39.4%) [5,11,16,31,32]. Compared to the high prevalence of A. marginale in cattle in Madagascar (89.7%), north-eastern Uganda (82.9%) [11], South Africa (57%) [31], Thailand (39.1%) [13] and in China (31.6%)[33], we demonstrated a relatively low infection rate of A. marginale (2.61%) in Chongqing. In addition, 4.93% of cattle tested positive for A. phagocytophilum in this study, which was similar to the positivity rate (5.3%) of this pathogen in white yaks [30]. A. platys infection in cattle was first reported in Algeria [4], while Ben et al. reported a prevalence of A. platys-like species (3.5%, 13/367) in cattle in Tunisia [34]. In this study, we noted a prevalence of A. platys (4.35%, 15/345) in cattle for the first time in Chongqing.

The prevalence of B. bigemina in this study was similar to previous research by Liu et al [21], and is lower than that reported in other provinces of China [19–21,35], South Africa [36], and in Tanzania [37]. However, the prevalence in our study was higher than that recorded in the Philippines [38]. In this survey, only T. sinensisi and T. orientalis were detected, with the prevalence being lower than T. sinensisi and T. orientalis infection rates recorded elsewhere [13,14,17]. Similar to the previous report [17], we did not detected B. bovis infection in cattle. The reason may be that 1) B. bovis infection in tick is usually lower than B. bigemina, which result a lower transmission rates of B. bovis, and 2) B. bovis-infected red blood cells usually accumulate in the capillary bed and leading to low parasitemia in circulating blood [17]. For the reasons that T. annualata, transmitted by Hyalomma anatolicum anatolicum, is mainly distributed in Northern China [39], T. luwenshuni and T. uilenbergi, both transmitted by Haemaphysalis qinghaiensis and H. longicornis, usually infected sheep and goats in China [27], and there is no evidence of above ticks existence in Chongqing. It was not strange that we did not detected T. annulata, T. luwenshuni, or T. uilenbergi infection in cattle from Chongqing.

There were 117 described species in the Chinese tick, 38 of which carry multiple pathogens [40], and most of the ticks including H. anatolicum, H. qinghaiensis, H. longicornis, H. bispinosa, Rhipicephalus (Boophilus) microplus, R. sanguineus, Dermacentor abaensis, D. silvarum and D. nuttalli were founded in northwest, northeast or central of China [39,41–44], and these ticks are responsible for transmission of a large amount of TBPs. However, the only reported tick specie in Chongqing was R. microplus [45], which was recorded to be the vector of A. phagocytophilumin, A. marginale, B. bigemina and B. bovis in China [40,46,47]. The differences in the prevalence of some parasites in this study compared to that reported previously in other studies in China or other countries, might be associated with geographical difference and variation in tick species.

Risk factor analysis revealed a significant correlation of altitude and age with the prevalence of B. bigemina and Theileria spp., which supported a previous report that there was a trend in increased seropositivity for B. bigemina infection with age [37]. In addition, gender is a risk factor associated with prevalence of Anaplasma spp. and Theileria spp., in cattle, which showed that male cattle had higher risk for these two type of pathogens infection, and the presence of cats in farm had positive effect on Theileria spp. infection in cattle from Chongqing, and the reasons for these phenomenon are not clear.

This study first took a single infection as a risk factor in evaluating the impact on infection with other pathogens. We found that cattle infected with A. bovis or A. central were more likely to be infected with A. phagocytophilum, and there was also a strong association between A. bovis and A. central infection. In addition, a very close relationship was observed for co-infection with T. sinensisi and A. marginale, and B. bigemina and T. orientalis. Anaplasma spp., Babesia spp., and Theileria spp. are all tick borne pathogens (TBPs), and some ticks can harbor mixed TBPs [40,47,48]. For the reasons that one species of TBP can be spread by different types of ticks, and equally that the same type of tick may also be the transmission vector for many species of TBPs, the significant correlation of the aforementioned pathogens might be attributed to the fact that infected cattle were bitten by ticks carrying different pathogens. From current data, it is not possible to estimate the chronological order of the aforementioned pathogen infections but there does appear to be significant relationships among some of these pathogens during infection of cattle. Parasite-parasite interaction may modify the impact of the pathogenic species and affect the performance and survival of host [49,50]. It is a pity that this study failed to evaluated the effect of above TBPs on health of cattle, since all the sampled animals in this study were apparently healthy, and we did not track the outcome of these cattle and the causes of their death. Further research should be conducted to elucidate the type of ticks present in Chongqing and the proportion of ticks that carry TBPs. Furthermore, attempts should be made to confirm whether a pathogen significantly increases the incident infection of other pathogens and the effects on production performance of cattle.

Conclusions

The results of the present survey indicated that infection of cattle with Anaplasma spp., Babesia spp., and Theileria spp. is widespread in Chongqing. We provide a possible relationship between afore-mentioned pathogenic infections, which will help in formulating appropriate control strategies for these pathogens in this area.

Supporting information

S1 Table. Primers used for Anaplasma spp., Babesia spp. and Theileria spp. detection in cattle.

https://doi.org/10.1371/journal.pone.0215585.s001

(DOCX)

Acknowledgments

We really appreciate Prof. Kui Nie and Dr. Cailiang Fan for their practical support, Mrs. Chunxia Dong and Mr. Hongquan Lin for their assistant in sample collection. We also thank Prof. Jianjun Wen for English-language editing of this manuscript.

References

  1. 1. Vayssier-Taussat M, Cosson JF, Degeilh B, Eloit M, Fontanet A, Moutailler S, et al. How a multidisciplinary 'One Health' approach can combat the tick-borne pathogen threat in Europe. Future Microbiol. 2015, 10(5):809–818. pmid:26000651
  2. 2. Makala LH, Mangani P, Fujisaki K, Nagasawa H. The current status of major tick borne diseases in Zambia. Vet Res. 2003, 34(1):27–45. pmid:12588682
  3. 3. Battilani M, De Arcangeli S, Balboni A, Dondi F. Genetic diversity and molecular epidemiology of Anaplasma. Infect Genet Evol. 2017, 49:195–211. pmid:28122249
  4. 4. Dahmani M, Davoust B, Benterki MS, Fenollar F, Raoult D, Mediannikov O. Development of a new PCR-based assay to detect Anaplasmataceae and the first report of Anaplasma phagocytophilum and Anaplasma platys in cattle from Algeria. Comp Immunol Microbiol Infect Dis. 2015, 39:39–45. pmid:25748051
  5. 5. Rjeibi MR, Ayadi O, Rekik M, Gharbi M. Molecular survey and genetic characterization of Anaplasma centrale, A. marginale and A. bovis in cattle from Algeria. Transbound Emerg Dis. 2018, 65(2):456–464. pmid:29034616
  6. 6. Bock R, Jackson L, de Vos A, Jorgensen W. Babesiosis of cattle. Parasitology. 2004, 129 Suppl: S247–S269. pmid:15938514
  7. 7. Liu A, Guan G, Liu Z, Liu J, Leblanc N, Li Y, et al. Detecting and differentiating Theileria sergenti and Theileria sinensis in cattle and yaks by PCR based on major piroplasm surface protein (MPSP). Exp Parasitol. 2010, 126(4):476–481. pmid:20685208
  8. 8. Luo J, Lu W. Cattle theileriosis in China. Tropical Animal Health & Production. 1997, 29(4):4S–7S.
  9. 9. Qin G, Li Y, Liu J, Liu Z, Yang J, Zhang L, et al. Molecular detection and characterization of Theileria infection in cattle and yaks from Tibet Plateau Region, China. Parasitol Res. 2016, 115(7):2647–2652. pmid:27000088
  10. 10. Bai Q, Liu GY, Yin H, Zhao Q, Liu DK, Ren JX, et al. Theileria Sinensis sp nov: A New Species of Bovine Theileria—Classical Taxonomic Studies. Acta Veterinaria et Zootechnica Sinica 2002, 33(1):73–77.
  11. 11. Byaruhanga C, Collins NE, Knobel DL, Khumalo Z, Chaisi ME, Oosthuizen MC. Molecular detection and phylogenetic analysis of Anaplasma marginale and Anaplasma centrale amongst transhumant cattle in north-eastern Uganda. Ticks Tick Borne Dis. 2018, 9(3): 580–588. pmid:29422446
  12. 12. Han DG, Ryu JH, Chae JB, Kim DW, Kwon CH, Choi KS. First report of Anaplasma phagocytophilum infection in Holstein cattle in the Republic of Korea. Acta Trop. 2018, 183:110–113. pmid:29654732
  13. 13. Jirapattharasate C, Adjou MP, Cao S, Iguchi A, Liu M, Wang G, et al. Molecular detection and genetic diversity of bovine Babesia spp., Theileria orientalis, and Anaplasma marginale in beef cattle in Thailand. Parasitol Res. 2017, 116(2):751–762. pmid:28028631
  14. 14. Zhou M, Cao S, Sevinc F, Sevinc M, Ceylan O, Moumouni P, et al. Molecular detection and genetic identification of Babesia bigemina, Theileria annulata, Theileria orientalis and Anaplasma marginale in Turkey. Ticks Tick Borne Dis. 2016, 7(1):126–134. pmid:26492823
  15. 15. Adjou MP, Aboge GO, Terkawi MA, Masatani T, Cao S, Kamyingkird K, et al. Molecular detection and characterization of Babesia bovis, Babesia bigemina, Theileria species and Anaplasma marginale isolated from cattle in Kenya. Parasit Vectors. 2015, 8:496. pmid:26420543
  16. 16. Belkahia H, Ben SM, Alberti A, Abdi K, Issaoui Z, Hattab D, et al. First molecular survey and novel genetic variants' identification of Anaplasma marginale, A. centrale and A. bovis in cattle from Tunisia. Infect Genet Evol. 2015, 34:361–371. pmid:26117444
  17. 17. Li S, Liu J, Liu A, Li Y, Wang S, Wang S, et al. Molecular investigation of piroplasma infection in white yaks (Bos grunniens) in Gansu province, China. Acta Trop. 2017, 171:220–225. pmid:28427963
  18. 18. Yang J, Li Y, Liu Z, Liu J, Niu Q, Ren Q, et al. Molecular detection and characterization of Anaplasma spp. in sheep and cattle from Xinjiang, northwest China. Parasit Vectors. 2015, 8:108. pmid:25889906
  19. 19. Li Y, Liu P, Wang C, Chen G, Kang M, Liu D, et al. Serologic Evidence for Babesia bigemina Infection in Wild Yak (Bos mutus) in Qinghai Province, China. J Wildl Dis. 2015, 51(4):872–875. pmid:26267460
  20. 20. Niu Q, Liu Z, Yu P, Yang J, Abdallah MO, Guan G, et al. Genetic characterization and molecular survey of Babesia bovis, Babesia bigemina and Babesia ovata in cattle, dairy cattle and yaks in China. Parasit Vectors. 2015, 8:518. pmid:26452623
  21. 21. Liu J, Guan G, Liu A, Li Y, Yin H, Luo J. A PCR method targeting internal transcribed spacers: the simultaneous detection of Babesia bigemina and Babesia bovis in cattle. Acta Parasitol. 2014, 59(1):132–138. pmid:24570060
  22. 22. Liu A, Guan G, Du P, Gou H, Zhang J, Liu Z, et al. Rapid identification and differentiation of Theileria sergenti and Theileria sinensis using a loop-mediated isothermal amplification (LAMP) assay. Vet Parasitol. 2013, 191(1–2):15–22. pmid:22947247
  23. 23. D'Oliveira C, Van DWM, Habela MA, Jacquiet P, Jongejan F. Detection of Theileria annulata in blood samples of carrier cattle by PCR. J Clin Microbiol. 1995, 33(10):2665. pmid:8567902
  24. 24. Barlough JE, Madigan JE, DeRock E, Bigornia L. Nested polymerase chain reaction for detection of Ehrlichia equi genomic DNA in horses and ticks (Ixodes pacificus). Vet Parasitol. 1996, 63(3–4):319–329. pmid:8966998
  25. 25. Inokuma H, Ohno K, Onishi T, Raoult D, Brouqui P. Detection of ehrlichial infection by PCR in dogs from Yamaguchi and Okinawa Prefectures, Japan. J Vet Med Sci. 2001, 63(7):815–817.
  26. 26. Kawahara M, Rikihisa Y, Lin Q, Isogai E, Tahara K, Itagaki A, et al. Novel genetic variants of Anaplasma phagocytophilum, Anaplasma bovis, Anaplasma centrale, and a novel Ehrlichia sp. in wild deer and ticks on two major islands in Japan. Appl Environ Microbiol. 2006, 72(2):1102–1109. pmid:16461655
  27. 27. Yin H, Liu Z, Guan G, Liu A, Ma M, Ren Q, et al. Detection and differentiation of Theileria luwenshuni and T. uilenbergi infection in small ruminants by PCR. Transboundary & Emerging Diseases 2008, 55(5‐6):233–237.
  28. 28. Terkawi MA, Huyen NX, Shinuo C, Inpankaew T, Maklon K, Aboulaila M, et al. Molecular and serological prevalence of Babesia bovis and Babesia bigemina in water buffaloes in the northeast region of Thailand. Vet Parasitol. 2011, 178(3–4):201–207.
  29. 29. Zhang L, Wang Y, Cai D, He G, Cheng Z, Liu J, et al. Detection of Anaplasma marginale in Hyalomma asiaticum ticks by PCR assay. Parasitol Res. 2013, 112(7):2697–2702. pmid:23636309
  30. 30. Yang J, Liu Z, Niu Q, Liu J, Guan G, Xie J, et al. First molecular survey and identification of Anaplasma spp. in white yaks (Bos grunniens) in China. Parasitology. 2016, 143(6):686–691. pmid:27003378
  31. 31. Hove P, Chaisi ME, Brayton KA, Ganesan H, Catanese HN, Mtshali MS, et al. Co-infections with multiple genotypes of Anaplasma marginale in cattle indicate pathogen diversity. Parasit Vectors. 2018, 11(1):5. pmid:29298712
  32. 32. Sisson D, Hufschmid J, Jolles A, Beechler B, Jabbar A. Molecular characterisation of Anaplasma species from African buffalo (Syncerus caffer) in Kruger National Park, South Africa. Ticks Tick Borne Dis. 2017, 8(3):400–406. pmid:28169172
  33. 33. Yang J, Han R, Liu Z, Niu Q, Guan G, Liu G, et al. Insight into the genetic diversity of Anaplasma marginale in cattle from ten provinces of China. Parasit Vectors. 2017, 10(1):565. pmid:29132409
  34. 34. Ben SM, Belkahia H, El MN, Saidani M, Alberti A, Zobba R, et al. Anaplasma platys-like strains in ruminants from Tunisia. Infect Genet Evol. 2017, 49:226–233. pmid:28130168
  35. 35. Qin SY, Wang JL, Ning HR, Tan QD, Yin MY, Zhang XX, et al. First report of Babesia bigemina infection in white yaks in China. Acta Trop. 2015, 145:52–54. pmid:25700713
  36. 36. Mtshali PS, Tsotetsi AM, Thekisoe MM, Mtshali MS. Nested PCR detection and phylogenetic analysis of Babesia bovis and Babesia bigemina in cattle from Periurban localities in Gauteng Province, South Africa. J Vet Med Sci. 2014, 76(1):145–150. pmid:24065081
  37. 37. Swai ES, Karimuribo ED, French NP, Fitzpatrick JL, Bryant MJ, Kambarage DM, et al. Seroprevalence of Babesia bigemina in smallholder dairy cattle in Tanzania and associated risk factors. J S Afr Vet Assoc. 2007, 78(1):15–20. pmid:17665760
  38. 38. Yu L, Terkawi MA, Cruzflores MJ, Claveria FG, Aboge GO, Yamagishi J, et al. Epidemiological survey of Babesia bovis and Babesia bigemina infections of cattle in Philippines. J Vet Med Sci. 2013, 75(7):995–998. pmid:23470278
  39. 39. Luo JX, Yin H, Guan GQ, Ma ML, Sun CQ, Lu BY, et al. Experiments on Transmission of Babesia U sp.and Theileria annulata with Hyalomma anatolicum anatolicum. Acta Veterinaria et Zootechnica S inica. 2005, 36(11): 1197–1201. (in Chinese).
  40. 40. Yu Z, Wang H, Wang T, Sun W, Yang X, Liu J. Tick-borne pathogens and the vector potential of ticks in China. Parasit Vectors. 2015, 8:24. pmid:25586007
  41. 41. Han R, Yang J, Niu Q, Liu Z, Chen Z, Kan W, et al. Molecular prevalence of spotted fever group rickettsiae in ticks from Qinghai Province, northwestern China. Infect Genet Evol. 2018, 57: 1–7. pmid:29107656
  42. 42. Yu P, Liu Z, Niu Q, Yang J, Abdallah MO, Chen Z, et al. Molecular evidence of tick-borne pathogens in Hyalomma anatolicum ticks infesting cattle in Xinjiang Uygur Autonomous Region, Northwestern China. Exp Appl Acarol. 2017, 73: 269–281. pmid:28875270
  43. 43. Niu Q, Liu Z, Yang J, Gao S, Pan Y, Guan G, et al. Genetic characterization and molecular survey of Babesia sp. Xinjiang infection in small ruminants and ixodid ticks in China. Infect Genet Evol 49: 330–335. pmid:28131866
  44. 44. Chen Z, Liu Q, Liu JQ, Xu BL, Lv S, Xia S, et al. Tick-borne pathogens and associated co-infections in ticks collected from domestic animals in central China. Parasit Vectors. 2014, 7: 237. pmid:24886497
  45. 45. Tang M, Nie K, Wu Q, Jin A, Wen C, Sun T, et al.Investigation of livestock and poultry parasites in Chongqing. Chinese Journal of Veterinary Parasitology.2003, 11(1): 25–30. (in Chinese).
  46. 46. Tian Z, Du J, Yang J, Liu A, Liu X, Liu G, et al. A PCR-RFLP Assay targeting RPS8 gene for the discrimination between bovine Babesia and Theileria species in China. Parasit Vectors. 2015, 8: 475. pmid:26382041
  47. 47. Zhang L, Liu H, Xu B, Lu Q, Li L, Chang L, et al. Anaplasma phagocytophilum infection in domestic animals in ten provinces/cities of China. Am J Trop Med Hyg. 2012, 87(1):185–189. pmid:22764312
  48. 48. Palomar AM, Portillo A, Santibanez P, Mazuelas D, Roncero L, Garcia-Alvarez L, et al. Detection of tick-borne Anaplasma bovis, Anaplasma phagocytophilum and Anaplasma centrale in Spain. Med Vet Entomo. 2015, 29(3):349–353.
  49. 49. Thumbi SM, Bronsvoort BM, Poole EJ, Kiara H, Toye PG, Mbole-Kariuki MN, et al. Parasite co-infections and their impact on survival of indigenous cattle. PLoS One. 2014, 9: e76324. pmid:24586220
  50. 50. Craig BH, Tempest LJ, Pilkington JG, Pemberton JM. Metazoan-protozoan parasite co-infections and host body weight in St Kilda Soay sheep. Parasitology. 2008. 135: 433–441. pmid:18215336