Multilocus sequence typing of Cryptosporidium hominis from northern India
Pooja Yadav1, Bijay Ranjan Mirdha1, Govind K Makharia2
1Department of Microbiology, All India Institute of Medical Sciences, New Delhi, India.
Background & Objectives:
Human cryptosporidiosis is endemic worldwide, and at least eight species have been reported in humans; the most common being Cryptosporidium hominis and C. parvum. Detailed understanding of the epidemiology of Cryptosporidium is increasingly facilitated using standardized universal technique for species differentiation and subtyping. In this study micro- and minisatellite targets in chromosome 6 were used to assess genetic diversity of C. hominis by sequence length polymorphisms along with single nucleotide polymorphisms (SNPs).
Methods:
A total of 84 Cryptosporidium positive stool specimens were subjected to speciation and genotyping using small subunit (SSU) ribosomal RNA (rRNA) as the target gene. Genetic heterogeneity amongst C. hominis isolates was assessed by sequencing minisatellites, microsatellites and polymorphic markers including genes encoding the 60 kDa glycoprotein (GP60), a 47 kDa protein (CP47), a mucin-like protein (Mucin-1), a serine repeat antigen (MSC6-7) and a 56 kDa transmembrane protein (CP56).
Results:
Of the 84 Cryptosporidium positive stool specimens, 77 (92%) were positive by SSU rRNA gene polymerase chain reaction (PCR) assay. Of these 77 isolates, 54 were identified as C. hominis and 23 as C. parvum. Of all the loci studied by multilocus sequence typing (MLST), GP60 gene could reveal the highest genetic diversity. Population substructure analysis of C. hominis performed by combined sequence length and nucleotide polymorphism showed nine multilocus subtypes, all of which were distinct groups in the study population.
Interpretation & Conclusions:
MLST, a powerful discriminatory test, demonstrated both variations and distribution pattern of Cryptosporidium species and its subtypes.
Insights
Multilocus sequence typing (MLST) revealed nine distinct subtypes of Cryptosporidium hominis, enhancing our understanding of this common parasitic infection. This genetic diversity analysis is crucial for tracking Cryptosporidium epidemiology.
Area of Science:
- * Molecular epidemiology
- * Parasitology
- * Infectious disease genetics
Background:
- * Human cryptosporidiosis is a globally prevalent parasitic disease caused by various Cryptosporidium species, primarily Cryptosporidium hominis and Cryptosporidium parvum.
- * Accurate species differentiation and subtyping are essential for understanding the epidemiology of Cryptosporidium infections.
- * Genetic markers, including microsatellites and single nucleotide polymorphisms (SNPs), are valuable tools for assessing genetic diversity.
Purpose of the Study:
- * To assess the genetic diversity of Cryptosporidium hominis using microsatellite and minisatellite targets on chromosome 6.
- * To differentiate and subtype Cryptosporidium isolates from human stool specimens.
- * To evaluate the utility of multilocus sequence typing (MLST) for Cryptosporidium genotyping.
Main Methods:
- * Speciation and genotyping of 84 Cryptosporidium-positive stool specimens using small subunit ribosomal RNA (SSU rRNA) gene sequencing.
- * Analysis of genetic heterogeneity in Cryptosporidium hominis isolates via sequencing of minisatellites, microsatellites, and polymorphic markers (GP60, CP47, Mucin-1, MSC6-7, CP56).
- * Application of multilocus sequence typing (MLST) incorporating sequence length and nucleotide polymorphisms.
Main Results:
- * Of 84 specimens, 77 were positive by SSU rRNA PCR, with 54 identified as Cryptosporidium hominis and 23 as Cryptosporidium parvum.
- * The GP60 gene exhibited the highest genetic diversity among the studied loci.
- * MLST analysis revealed nine distinct multilocus subtypes within the Cryptosporidium hominis population.
Conclusions:
- * Multilocus sequence typing (MLST) is a robust method for discriminating Cryptosporidium species and subtypes.
- * The study identified significant genetic variation and distinct population substructures within Cryptosporidium hominis.
- * Understanding Cryptosporidium genetic diversity is key to effective epidemiological surveillance and control strategies.
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