Variable Number Tandem Repeats for Global Strain Typing and Strain Differentiation of Mycobacterium leprae

Leprosy is chronic granulomatous disease caused by Mycobacterium leprae and associated with the disability, stigma and discrimination to the affected individuals. Though often considered a disease of antiquity, it is found most commonly today in tropical and sub-tropical regions. Global efforts to eradicate leprosy have been largely successful in controlling its spread. Despite these efforts, the disease remains endemic countries emphasizing the need for greater scrutiny of its epidemiology. Strain typing and strain differentiation by Variable Number Tandem repeats (VNTR) could be useful in tracing origins and routes of infection, general leprosy surveillance and prevalence. Strain typing of Mycobacterium leprae by VNTR has been successfully carried out and predominance of leprosy bacilli in different geographical region also done by VNTR. In the present review, we reported global distribution M. leprae by Variable Number tandem repeats in systematic way to easy understanding of leprosy distribution.


Review
Leprosy is chronic granulomatous disease caused by Mycobacterium leprae and associated with the disability, stigma and discrimination to the affected individuals [1]. Though often considered a disease of antiquity, it is found most commonly today in tropical and sub-tropical regions [2]. Global efforts to eradicate leprosy have been largely successful in controlling its spread. Despite these efforts, the disease remains endemic countries, with 232,857 cases reported cases globally in 2012 [3]. India accounted for 1.27 of cases reported in 2011-2012, emphasizing the need for greater scrutiny of its epidemiology. strain typing and strain differentiation are very helpful to identify the source of infection, transmission of infection and spreading of disease, differentiating cases of relapse from re infection, and for unravelling possible links between human and nonreservoir sources [4,5]. In recent years molecular typing methodologies have complemented conventional infectious disease.
Strain tying and strain differentiation is a method which useful to distinguishing members of the same microbial species from one another on the basis of genotype. Strain typing explains that one isolate is same or it's different. Several molecular typing methods have been employed to distinguish the M. leprae strains over the years.
Initially methods like surface antigen typing [6], multi-locus enzyme electrophoresis (MLEE) [7] and phage typing [8] were used for strain typing. DNA based strain typing methods were started from 1990. The methods includes fragment length polymorphism (RFLP) typing at locus such as rRNA operons [9] fingerprinting by randomly primed PCR [10] pulsed-field gel electrophoresis (PFGE), multi-locus sequence typing (MLST) [11], variable umber tandem repeats (VNTR) and SNP. If genetic diversity less, VNTR is good source for the identification of strain variation [12,13]. VNTRs have short, tandemly repeated motifs typically ranging from 1 to 60 bp in length. Polymorphisms at those sites consist of differences in the number of repeat sequences contained by a VNTR and are the consequence of mutations which occur during DNA replication, when repeats are inserted or deleted by DNA polymerase due to slipped-strand mispairing [14]. There is considerably more variation in repeat numbers at VNTR locus than in non-repetitive DNA sequences, because length-altering mutations due to slipped-strand mispairing occur at a much higher rate than the inherent DNA substitution or mutation frequency of DNA polymerase [15].
Surprisingly little is known about the biology and epidemiology M. leprae. Two key properties of the organism explain both much of our ignorance about the organism and the fairly primitive state of the molecular epidemiology of M. leprae. In most of the bacteria small sequence of few housekeeping genes as in MLST is enough for inferring relationships among strains. But genetically monomorphic species reveals that no genetic polymorphism in these species [16,17]. M. leprae has associated with the less genetic diversity. This less genetic diversity due to high content of pesudogenes and several DNA repair genes [18]. Sever notable issues have been identified which limit the molecular epidemiological study of leprosy. Good efforts were made to strain typing even within less genetic polymorphism genome by restriction fragment length polymorphism (RFLP) [19,20], examination of dispersed repeats [21] and sequence analysis of variable genomic regions [22] proved universally unsuccessful but last decades powerful methods have been made to strain typing of leprosy.
The actual strain typing methods were started after complete genome sequence of M. leprae TN strain and initially 50 VNTR loci have identified. Continue efforts on this VNTR studies showed the path for utility in molecular epidemiology studies for strain typing [23][24][25][26]. Robust PCR amplification technique have involved in the development and validation of VNTR markers. To date, based on several techniques, several VNTR markers were addressed and in future several VNTR loci may add. Initial studies of VNTR have associated with several controversies which were later solved by subsequent VNTR studies [27]. Whole genome sequencing methods were dropped as it is cost effective so that alternative studies have made for strain typing in which SNP and VNTRs are most effective for leprosy. Although several polymorphic sites were available, only small fraction regions were associated with variation and those were involved in the global strain typing of M.leprae. The information provided by VNTR neither was nor did similar to the SNP result and resolution of strain relationship with SNP quite limit. So VNTR study was suitable for strain typing of microorganisms.
The selection of VNTR was mainly dependent on timescale of the event which can easily tracing strain variation within short time. These VNTR markers were also more suitable for probing historical distribution of strains over hundreds to thousands of years. Molecular epidemiology studies of M. leprae strains collected within individual nations have revealed several general properties of leprosy and leprosy transmission at that population level. After standardization VNTR PCR study in proper way by Gillis in the year 2009, several studies were used those primers for study of strain variation globally [26,[28][29][30][31]. These studies indicated that, the strain possess same allelic distribution with the country and those strains have some alleles which were alike of the other countries in allelic distribution which indicated that those are genetically similar and migration was there in these populations. Some studies have strongly suggested that leprosy bacilli strains were migrated from India to Philippines with highest genetic diversity and identified 3 distinct groups. Interestingly the first sequenced Indian strain TN was genetically similar with Philippines leprosy strains which suggested that TN is not an Indian strain and which may emigrate from some other countries. The detection rate of new and relapse cases in each year and the national prevalence reflects the continuing spread of leprosy. In such areas the strain typing and strain differentiation are very helpful to identify the source of infection, transmission of infection and spreading of disease, differentiating cases of relapse from re infection, and for unravelling possible links between human and non-reservoir sources.