Molecular tools for the classification and identification of members of the Leishmania donovani 'complex'

B. Jamjoon, Manal (2003) Molecular tools for the classification and identification of members of the Leishmania donovani 'complex'. PhD thesis, University of Liverpool.

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Abstract

Visceral leishmaniasis is a potentially lethal disease. It is clinically presented by viscerotropic dissemination to internal organs. The agents causing the disease are all grouped within the L. donovani "complex" (Lainson and Shaw, 1987). Many issues in the taxonomy within this complex are still controversial, such as the agents causing visceral leishmaniasis in Sudan and the validity and the identity of L. "archibaldi". The agents causing VL in Sudan are at present defined on the basis of an isoenzyme classification as Leishmania donovani, L. infantum and L. "archibaldi". L. "archibaldi" only differs from L. donovan; in the mobility of one enzyme (GOT). The presence of all three species in Sudan has been contested by many authors who suggested grouping the zymodemes belonging to these two taxa in a single group L. donovan; sensu lato (Ashford et al. 1992). In this study we have obtained the sequence of the chitinase gene of 37 stocks of these parasites from the Sudan and elsewhere to construct a phylogenetic tree. A panel of microsatellite markers suitable for classifying these species was also developed. Two strategies were used to develop the panel of microsatellites. Firstly, the Leishmania major genome sequence was used to identify microsatellite markers. Twenty-seven independent microsatellite loci were identified by BLAST search of the L. major genome. 13 out of 27 primers designed against the L. major sequences also amplified a single product of approximately the expected size from L. donovani. These 13 microsatellites were tested for variation in a panel of the L. donovan; "complex". Only two out of 13'loci that were polymorphic in L. major were also polymorphic in L. donovani. Almost all microsatellite peR products were significantly smaller in all the test strains than they were in L. major. Therefore, the use of the L. major genome sequence to identify microsatellite loci is unlikely to be an efficient method of identifying microsatellite loci in other Leishmania strains. Secondly, forty microsatellites were identified in L. donovani by an enrichment method (http://WWW.liv.ac.uk/-kempsj/genomics.htmI.).Primers for twenty of these amplified a single sharp band from L. donovani DNA and were found to be polymorphic between L. donovani stocks. Phylogenetic trees were constructed using microsatellite data. Both chitinase and microsatellite phylogenies showed that stocks of all three species isolated from Sudan form a single monophyletic group within the Leishmania donovani, / L. infantum clade. We conclude that L. "archibaldi" is not a valid species and the definitions of L. donovani and L. infantum may have to be revised in the light of this data.

Item Type:	Thesis (PhD)
Depositing User:	Symplectic Admin
Date Deposited:	23 Oct 2023 10:19
Last Modified:	23 Oct 2023 10:27
DOI:	10.17638/03176017
Copyright Statement:	Copyright © and Moral Rights for this thesis and any accompanying data (where applicable) are retained by the author and/or other copyright owners. A copy can be downloaded for personal non-commercial research or study, without prior permission or charge.
URI:	https://livrepository.liverpool.ac.uk/id/eprint/3176017