Toxocara canis (Ascaridida: Nematoda): Mitochondrial gene content, arrangement and composition compared with other Toxocara species

Introduction: The complete mitochondrial (mt) genomes of nematodes sequenced thus far are circular and small in length, containing 37 genes, including 12–13 protein-coding genes: namely, cytochrome oxidase subunits 1–3 (cox1–3), nicotinamide adenine dinucleotide dehydrogenase subunits 1–6 and 4L (nad1–6 and nad4L), ATP synthase subunit 6 and 8 (atp6 and 8), cytochrome b (cytb), small subunit ribosomal RNA gene (rrnS), large subunit ribosomal RNA. The aims of the present study are to determine the gene order, composition, codon usage pattern, and translation initiation/termination codons of the mt genome of Toxocara canis found in Sri Lanka and to compare those findings with other Toxocara species.

Methodology: Toxocara canis worms used in the present study were obtained from Sri Lanka. The total genomic DNA was extracted using a part of a single worm using an Easy-DNATM Kit. PCR reactions were performed as described previously by Wickramasinghe et al., (2009). Nucleotide sequences were determined with an ABI PRISM 3100 Avant DNA sequencer using a Big Dye Terminator v3.1 Cycle Sequencing Kit. The open-reading frames and codon usage profiles of protein-coding genes were analyzed using the program DNAsis and GENETYXMAC (ver. 6.0).

Results: The partial mt genome of T. canis is 9611 bp in length. This partial mt genome contains 9 protein-coding genes (cox1-3, nad2-5, atp6 and cytb), 14 transfer RNA (tRNA) genes and one ribosomal RNA gene. All genes are transcribed in the same direction as found in other members of the genus Toxocara. The nucleotide compositions of the obtained partial mtDNA sequences for T. canis is biased toward A and T, with T being the most favored nucleotide and C the least favored, in accordance with mt genomes of other nematodes. The content of A+T is 67.48% for T. canis (19.9% A, 47.5% T, 23.3% G and 9.3% C). The start codons we inferred in T. canis were ATT, GTG, ATA, GTT, TTG and ATG. Stop codons were TAG, TA and T. Six of the 8 protein genes used TAG as a translation termination codon and the other two genes (atp6 and nad2) used TA and T respectively. The length of 14 tRNAs is ranging from 52 bp to 68 bp and 8 more tRNAs remain to be sequenced and identified. The rrnL gene of T. canis was identified by sequence comparison with other Toxocara species. The rrnL is located between tRNA-His and nad3. The rrnL gene is 958 bp in length. A non-coding region is located between nad4 and cox1. The length of this region is 112 bp in T. canis.

Conclusion: The present study determined the partial mt genome sequence of T. canis obtained from a dog in Sri Lanka. This study provides molecular markers for studying the systematics, population genetics and ecology of the nematodes of socioeconomic importance. Furthermore, we observed a significant difference in nucleotide length of the mitochondrial protein-coding genes, tRNAs, rrnL and non-coding regions of T. canis from three different geographical locations, Sri Lanka, China and Australia. These variations should be further investigated in future studies.

Note: Nucleotide data reported in this paper is available in the GenBank^TM, EMBL and DDBJ databases under the accession number JN593098.

DOI: http://dx.doi.org/10.4038/sljid.v4i2.6946

Sri Lankan Journal of Infectious Diseases 2014; Vol.4(2):90-98