ABSTRACT
Background: fasciola species are parasitic trematode with worldwide distribution that infects wild and domesticated herbivores, particularly ruminants
Objectives: the aim of the present study was to investigate the intra species variations of F. gigantica, from goats and buffalo isolates in two common geographic climates of Iran
Methods: fasciola species were collected from goat, buffalo, sheep, and cattle in different regions. Cytochrome c oxidase I [COX1] of mitochondrial DNA [mt-DNA] was amplified from individual trematodes by polymerase chain reaction [PCR], using universal primers, and the amplicons were consequently sequenced and sequencing data were analyzed, using Clutal W software against the GenBank database
Results: a monomorphic DNA segment of approximately 499bp was seen in Fasciola isolates. The results of the amino acid sequence alignment defined strictly conserved amino acid residues in buffalo isolates of F. gigantica and partially conserved residues for goat isolates of F. gigantica. There are four tandem amino-acid replacements in the goat isolates at the position of 135-138, where Leucine [L], F [Phenylalanine], T [Threonine], and D [Aspartate] sequences changed into S [Serine], L [Leucine], H [Histidine], and L [Leucine], respectively. Furthermore, a replacement in the sequence of amino acid was found in isolates from buffalo at the position of 154, where Serine [S] was transformed into Leucine [L]
Conclusions: the findings of our study indicate that the variants of goat and buffalo can be responsible for persistence of Fasciola infection in the endemic areas of Iran. It seems that biological differences could occur by considering a variety of F. gigantica-hosts in Iran. Thus, suitable approaches are required for effective treatments and useful control strategies
ABSTRACT
Because of the strong immunologic responses of surface protein TaSp in Theileria annulata infected host, we tried to characterize this protein in a T. annulata isolate from Iran. The RNA prepared from T. annulata infected cells was used to produce SMART-DS-cDNA. The Double strand cDNA was then amplified with primers derived from TaSp mRNA sequences. The PCR product was cloned in pTZ57R/T vector, sequenced and registered under accession no. JQ003240 in GenBank. The sequence analysis showed 90%-94% nucleotide sequence identity and 68%-94% amino acid homology to the corresponding sequences of TaSp gene by T. annulata, T. sp. china I, T. sp. china and T. lestoquardi and three T. annulata reported from Iran respectively. Interestingly, the sequence analysis also showed small nucleotide sequence region near the 5` end in which the presented TaSp protein differed very strongly from the other known TaSp sequences. For the preparation of the recombinant protein, the cDNA was cloned in pQE-32 vector, the recombinant protein was prepared and assayed by Theileria infected bovine serum. The polymorphism in TaSp gene could be detected in intra- as well as inter species. The different characterized TaSp proteins had a common identic region, which may be helpful for development of broad band vaccine based on the recombinant proteins. The polymorphism in this gene, make this protein also interesting for the diagnostic purposes
Subject(s)
Cloning, Molecular , Protozoan Proteins , Theileria , Polymorphism, GeneticABSTRACT
We used the PCR technique based on the abovementioned primer pair and sequencing to demonstrate the Theileria infection in the sheep samples collected from Sultanate of Oman. According to the frame work of "integrated control of ticks and tick borne diseases in globalized world managed by EU-ICTTD-3 project, the samples from blood, liver, spleen, lymph node and lung were sent to the laboratory of Iranian Research Center for Ticks and Tick-borne Diseases [IRCTTD]. Samples from blood smear and impression smears from liver, spleen, lymph node, and lung were analyzed by Geimsa staining. The DNA was extracted from the abovementioned samples and analyzed by PCR technique using specific primers derived from the nucleotide sequences of 18S rRNA gene of T. lestoquardi, which can amplify the common region in other Theileria and Babesia spp. Subsequently the amplified DNA was sequenced. The analysis of blood smears of the sheep was negative for piroplasmosis performed through the Giemsa staining. The impression smears prepared from liver, spleen, lymph node, and lung showed suspicious structures mimicking Theileria schizonts in some cells. The results showed an expected PCR product of 428 bp in length, which is specific for Theileria spp. The PCR products were subsequently sequenced. The corresponding nucleotide sequence is registered under accession number JF309152 in GenBank. The sequence alignment in Gen Bank showed that the PCR products had 99% homology to the known T. lestoquardi registered under accession number AF081135 in the GenBank. Oman sheep are highly susceptible for Theileria infection and the infected sheep mostly die before the microschizonts or erythrocytic form of Theileria appears in the nucleated or erytrocytic cells respectively