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In this study, we used PCR to detect and characterize B. canis from naturally infected dogs in Poland with subclinical babesiosis by amplifying and sequencing a portion of the 18S ribosomal RNA (rRNA) gene. Venous blood samples were collected from ten dogs with subclinical babesiosis. A 559-bp fragment of the B. canis 18S rRNA gene was amplified by PCR. Sequencing of the PCR products led to the identification of a new variant of Babesia canis, differing from the previously detected protozoa genotypes (18S rRNA-A and 18S rRNA-B) with nucleotide substitutions in positions 150 and 151 of the tested gene fragment. The results indicate the emergence within the Polish territory of a new, previously unencountered Babesia canis genotype responsible for the development of subclinical babesiosis.
The aim of the study was the molecular analysis of B. canis strains isolated from dogs from Poland based on 18S RNA and Bc28 genes. The study involved 140 protozoan strains derived from clinical disease cases. All DNA samples of parasites were analyzed in two ways (amplification and restriction digestion) which made it possible to demonstrate the polymorphism of the Bc28 gene, and to show 18S RNA gene polymorphism (amplification and restriction digestion). Amplification of the Bc28 gene fragment and digestion of the resulting PCR products allowed for the classification of 104 isolates of B. canis to the Bc28-A group, and 36 strains of protozoa to the Bc28-B group. Amplification of the Bc28 gene fragment and digestion of the resulting PCR products allowed for the classification of 33 isolates to 18S RNA-A group, while to 18S RNA-B – 107 parasite isolates. Comparison of both groups of protozoa among themselves allowed partial but not complete correlation of polymorphisms in Bc28 and 18S RNA genes.
The aim of this paper was to analyse the amino acid sequences of the 18S rRNA gene of Babesia canis strains and the proteomic analysis of the serum of dogs infected with three various genotypes: 18S rRNA B. canis. Material for the research was DNA B. canis obtained from dogs with babesiosis. In total, 60 DNA tested samples were divided into three groups (20 samples each). The groups were formed by DNA samples of the sequences marked as 18S RNA-A (group 1), 18S RNA-B (group 2), and 18S RNA-C (group 3). The basis for the classification of protozoa to a specific group was the location of relevant nucleotides (GA, AG, or TT) in position 150-151 of the tested nucleotide sequence 18S rRNA. Nucleotide sequences were transcribed into amino acid sequences and then analysed using DNASTAR software. From all 60 infected and ten healthy dogs (control group), the serum was taken to make proteomic tests using MALDI-TOF mass spectrometer. It was demonstrated that the mutations found in position 150 and 151 of the nucleotide sequence, result in a change of amino acid sequences. Moreover, it was also demonstrated that the disease course in dogs infected with different strains of protozoa is different. Each of the analysed strains of protozoa induced in the serum of infected animals the appearance of a protein fraction of mass 51 kDa, which may then be treated as a nonspecific disease marker used for the diagnosis of this disease but not to differentiate the protozoa strains.
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