Molecular survey of piroplasm species from selected areas of China and Pakistan

Hassan, Mohammad Farooque; Liu, Junlong; Rashid, Imran; Iqbal, Naveed; Guan, Guiquan; Yin, Hong; Luo, Jianxun

doi:10.1186/s13071-018-3035-x

Cited by 26 publications

(24 citation statements)

References 29 publications

(37 reference statements)

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“…Babesia ovis was also found in one tick sample using species-specific primers. All of these pathogens have been previously reported from Pakistan [18, 30, 31, 60]. Theileriosis is an economically important disease affecting domestic and wild ruminants in tropical and subtropical regions of the world [61].…”

Section: Discussionmentioning

confidence: 99%

“…Furthermore, previous studies were confined to the surrounding areas of major metropolitan cities without considering diverse climatic conditions and bovine production systems in Pakistan which are important factors in the design of epidemiological studies [7]. Recently, several studies have utilized molecular methods for the detection of pathogens in bovines [30, 31] and their ticks [18]. However, these studies only targeted the main TBPs pathogens (e.g.…”

Section: Introductionmentioning

confidence: 99%

“…In the last decade, various TBPs using molecular methods have been reported from bovines in Pakistan [17, 18, 30, 31]. However, these methods are time-consuming as they can detect only a few pathogens at a time and require large volumes of DNA for the detection of multiple pathogens.…”

Section: Introductionmentioning

confidence: 99%

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Bovine ticks harbour a diverse array of microorganisms in Pakistan

et al. 2020

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BackgroundTicks and tick-borne pathogens (TTBP) are a major constraint to livestock production in Pakistan; despite a high prevalence of TTBPs, knowledge on the capacity of Pakistani ticks to carry pathogens and endosymbionts is limited. Furthermore, mixed infections with multiple microorganisms further complicate and limit the detection potential of traditional diagnostic methods. The present study investigated the tick-borne microorganisms in bovine ticks in Pakistan, employing a high-throughput microfluidic real-time PCR based technique.MethodsTicks were collected from clinically healthy cattle (n = 116) and water buffaloes (n = 88) from 30 villages across six districts located in five agro-ecological zones (AEZs) of Pakistan from September to November 2017. The microfluidic real-time PCR was used to test the genomic DNA of individual ticks for the presence of 27 bacterial and eight parasitic microorganisms. Phylogenetic methods were used to assess the genetic relationship of DNA sequences determined herein.ResultsPCR detected DNA of at least one microorganism in each of 221 ticks tested (94.4%, 221/234). DNA-based detection inferred that single pathogens/endosymbionts were the most common (43.4%, 96/221) followed by double (38.9%, 86/221), triple (14.5%, 32/221), quadruple (2.3%, 5/221) and quintuple (0.9%, 2/221) mixed infections. Piroplasms (Babesia/Theileria spp.) were the most prevalent (31.6%, 74/234), followed by Ehrlichia spp. (20%, 47/234) and Anaplasma marginale (7.7%, 18/234). Anaplasma phagocytophilum, A. ovis, A. centrale, Babesia ovis, Borrelia spp., Rickettsia spp., R. massiliae, Bartonella spp. and Hepatozoon spp. were also detected. Endosymbionts such as Francisella-like (91.5%, 214/234) and Coxiella-like (1.3%, 3/234) organisms were also detected in ticks. The highest diversity of microorganisms was detected in Hyalomma anatolicum ticks (test-positive for 14/14 microorganisms), followed by Rhipicephalus microplus (4/14), Hy. hussaini (3/14) and Rh. annulatus (2/14). Ticks collected from cattle carried significantly more frequently piroplasms (41.2%, 54/131; P < 0.05) than those from buffaloes (19.4%, 20/103). However, the overall prevalence of microorganisms did not vary significantly among ticks from the two host species as well as across different AEZs.ConclusionsTo our knowledge, this is the first study to investigate a wide range of tick-borne microorganisms in bovine ticks using a high-throughput diagnostic method from different AEZs in Pakistan. These findings will aid in establishing the distribution patterns and the control of tick-borne pathogens of bovines in Pakistan.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Bovine ticks harbour a diverse array of microorganisms in Pakistan

et al. 2020

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“…Bovine theileriosis affects domesticated cattle and is associated with significant reduction in cattle productivity [24]. Theileriosis caused by T. annulata (tropical theileriosis) and T. parva (East coast fever) are of high pathogenicity [25], compared to the benign bovine theileriosis caused by the T.…”

Section: Bovine Hemoplasmosis Is Caused By the Hemotropic Mycoplasmasmentioning

confidence: 99%

First report of bovine anaemia associated Theileria sinensis infection and phylogenetic analyses of partial gene sequences of Theileria and Anaplasma species detected in naturally infected Malaysian cattle

Agina¹,

Shaari²,

Isa³

et al. 2020

Preprint

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Background: Serious disease outbreaks in cattle are usually associated with blood pathogens. This study aims to detect blood pathogens namely Trypanosoma evansi, Theileria, Anaplasma and Mycoplasma species, and studied their phylogenetic relationships, haemato-biochemical abnormalities and erythrocyte osmotic fragility (EOF) in Malaysian cattle. Methods: Molecular analysis was achieved by PCR amplification and sequencing of PCR amplicons of 18SrRNA gene of Theileria species, 16SrRNA genes of Anaplasma and Mycoplasma species, 16SrRNA gene of Candidatus Mycoplasma haemobos, MPSP gene of T. orientalis and RoTaT1.2 VSG gene of Trypanosoma evansi, in sixty-one (61) Kedah-Kelantan X Brahman cattle from Pahang, Malaysia. Haemato-biochemical analyses were performed using automated analysers while EOF was determined with the aid of saline solutions. Results: PCR amplification produced the expected fragment sizes for MPSP gene of T. orientalis, msp4 gene of Anaplasma marginale, 16S rRNA gene of C. M. haemobos, RoTaT1.2VSG gene of T. evansi. Nucleotide blast demonstrated that sequences of the PCR amplicons showed a high degree of molecular similarity in comparison with reference sequences. Analysis of 18SrRNA gene sequences of Theileria species and 16S rRNA gene sequences of Anaplasma species revealed Theileria sinensis and Anaplasma platys as additional species detected in these cattle. Theileria species was the most detected blood pathogen in the sampled cattle. The blood picture of all cattle group revealed poikilocytosis, anisocytosis, rouleaux formation and degenerative left shift. Erythrocyte fragility values of all the cattle groups were above the reference range. Anaemia of the macrocytic normochromic type was observed in the Trypanosoma evansi; and Anaplasma platys + Theileria sinensis double species co-infected cattle. Normocytic normochromic anaemia was observed in the T. sinensis infected cattle group. Significant (p<0.05) increases in serum liver and kidney parameters, total protein, globulin, total and unconjugated bilirubin and decreased albumin was observed in the Trypanosoma evansi infected cattle.Conclusion: We present the first report of anaemia associated with Theileria sinensis infection in Malaysian cattle. Because of the high occurrence of bovine theileriosis and detection of Anaplasma platys, there is an urgent need for appropriate preventive and control measures, as Theileria species and A. platys are of great economic and zoonotic importance respectively.

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“…Furthermore, blood from pathogen-free cattle was processed for cell isolation and used as a control (∆Ct as calibrator for nBCs-LPS and TaBC), and nBCs were stimulated with LPS to induce cytokine production and compared with TaBCs. Piroplasm-free and piroplasm-infected cattle were confirmed by thin blood smear microscopic examination and PCR amplification of the target 18S rRNA gene by previously used and optimized genus-specific primers (primers details are shown in the Results section) [25].…”

Section: Methodsmentioning

confidence: 99%

Establishment and Expression of Cytokines in a Theileria annulata-Infected Bovine B Cell Line

Rashid

Guan

Luo

et al. 2019

Genes

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This study aimed to establish a pure single-cell Theileria annulata-infected B cell line for the assessment of cytokine production in transformed and lipopolysaccharide (LPS)-stimulated cells. Several studies have aimed to identify cell surface markers in T. annulata-transformed cells; however, no information on cytokine production in these cells is available. To investigate the potential of the transformed cells to produce cytokines and their potential responses to antigen-stimulation, we purified mature B cells (CD21) from the whole blood of cattle experimentally infected with the T. annulata Kashi strain by magnetic separation. The purity and specificity of the established cell line was assessed by the identification of specific cell surface markers (CD21, IgM, and WC4) by flow cytometry analysis. The transcript levels of the cytokines IL1A, IL1B, IL2, IL4, IL6, IL8, IL10, IL16, LTA, TGFB1, TNFA, IFNA, and IFNB in transformed, buparvaquone (BW720c)-treated cells, and antigen-stimulated cells were analyzed by quantitative polymerase chain reaction (qPCR) using cDNA from these cells. A T. annulata-infected bovine B cell line was successfully established with a purity of ~98.8% (CD21). IL4 and IL12A were significantly (p < 0.01) upregulated in the transformed cells. In BW720c-treated transformed cells, IL12B, TGFB1, and IFNB were significantly (p < 0.01) upregulated. Notably, no significant (p > 0.05) upregulation of cytokines was observed in LPS-stimulated transformed cells. Moreover, IL1A, IL1B, IL8, and IL16 were significantly (p < 0.01) upregulated in LPS-stimulated B cells. Our data signify the potential use of this cell line for cytokine production, observance of immunoglobulins, and production of an attenuated vaccine against tropical theileriosis.

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Molecular survey of piroplasm species from selected areas of China and Pakistan

Cited by 26 publications

References 29 publications

Bovine ticks harbour a diverse array of microorganisms in Pakistan

Bovine ticks harbour a diverse array of microorganisms in Pakistan

First report of bovine anaemia associated Theileria sinensis infection and phylogenetic analyses of partial gene sequences of Theileria and Anaplasma species detected in naturally infected Malaysian cattle

Establishment and Expression of Cytokines in a Theileria annulata-Infected Bovine B Cell Line

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