Simultaneous detection of piroplasma infections in field Haemaphysalis qinghaiensis ticks by reverse line blotting

Abstract

Piroplasmosis is caused by tick-borne haemoprotozoan parasites in the genera of Theileria and Babesia, in which numbers of agents are highly pathogenic for cattle, sheep and goats. We developed a reverse line blot (RLB) assay for detection and differentiation of four different parasites, in which 18S ribosomal RNA gene sequence was amplified with a set of universal primers specific for all members in the genera of Theileria and Babesia; and the probes were designed on the basis of hypervariable region 4 (V4 region) of 18S rRNA gene. Three Theileria and one Babesia can be detected simultaneously on this system and it was sensitive to detect a parasitemia level between 10(-5) and 10(-8)%. A total of 149 Haemaphysalis qinghaiensis ticks collected from Lintan County of Gannan Tibetan Autonomous Region was tested by RLB. Among these, 136 tick samples were also tested by a nested PCR assay developed previously. After comparison of these results, it showed that more T. luwenshuni was detected in RLB assay, while more T. uilenbergi was detected in H. qinghaiensis ticks by nested PCR. The RLB has shown capability for simultaneous detection of four species of piroplasm in H. qinghaiensis ticks, indicating its usefulness for epidemiological studies of piroplasmosis.

Identification of 12 Piroplasms Infecting Ten Tick Species in China Using Reverse Line Blot Hybridization

“…A previous study reported a high infection rate of T. luwenshuni in ticks by RLB, or in small ruminants via PCR and FRET-qPCR (Niu et al, 2012;Li et al, 2014a;Yang et al, 2014). In our study, T. luwenshuni was mostly prevalent in Linzhou, Henan province, corresponding with a previous study, which reported T. luwenshuni infecting sheep in the same province (Chen et al, 2014b).…”

“…motasi isolates, Babesia sp. Xinjiang, B. major, B. ovata, B. bigemina, B. bovis, T. uilenbergi, T. luwenshuni, T. ovis, T. annulata, T. sinensis, and T. sergenti) containing a N-(trifluoroacetamidohexyl-cyanoethyl, N, N-diisopropyl phosphoramidite [TFA])-C6 amino linker were synthetized (Gubbels et al, 1999;Schnittger et al, 2004;Niu et al, 2009Niu et al, , 2012Abdallah et al, 2016). Positive plasmids from Babesia or Theileria species genomic DNA, 2 species of bacteria (Anaplasma marginale and Borrelia burgdorferi sensu stricto), and water were used as positive, negative, and blank control, respectively, to perform the specificity and sensitivity of the RLB assay (Abdallah et al, 2016).…”

Detection of piroplasms infection in sheep, dogs and hedgehogs in Central China

“…It was reported that T. lunwenshuni was transmitted by Haemaphysalis qinghaiensis [31], and small ruminant animals were infected with it in the northern and central parts of China [32], with high pathogenicity causing great economic losses. Animals that survived acute infection could become low-level carriers of the parasite, and would remain persistently infected for years without apparent clinical signs [33].…”

Malignant Ovine Theileriosis ( Theileria Lestoquardi ) : A Review

“…It also requires sophisticated laboratory equipment and, due to a complex protocol with a need for controlled hybridization conditions, it may be subject to reproducibility problems in different laboratories. However, the small subunit ribosomal RNA gene (18S RNA gene) sequence data have been successfully used to improve the classification of previously known data and identify several novel Theileria and Babesia species (Altay et al, 2007;Niu et al, 2009;Oosthuizen et al, 2008Oosthuizen et al, , 2009Niu et al, 2012;Ranjbar et al, 2012).…”