Molecular Genotyping of Trypanosoma cruzi for Lineage Assignment and Population Genetics

Messenger, Louisa A.; Yeo, Matthew; Lewis, Michael D.; Llewellyn, Martin S.; Miles, Michael A.

doi:10.1007/978-1-4939-1438-8_19

Cited by 22 publications

(18 citation statements)

References 77 publications

(31 reference statements)

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“…Accordingly, further long-term studies are necessary to investigate this issue, especially those monitoring the treatment responses of patients with Chagas disease whose parasite DTU has been identified. Routine T. cruzi genotyping can be feasible at medical centers with scientific research support, since presently, there are several available molecular techniques for its characterization 8,14,17,72 , and the classical MLEE analysis can reveal the main genetic groups, mixed stocks, and other trypanosome species 14,25,[30][31][32][33] . Despite some controversies and exceptions, nowadays there are recommendations for treating all patients with chronic Chagas disease using new dosing strategies and drug combinations for preventing side effects, thus increasing the chances of treatment completion and monitoring of patients for a longer time [73][74][75] .…”

mentioning

confidence: 99%

Trypanosoma cruzi I genotype among isolates from patients with chronic Chagas disease followed at the Evandro Chagas National Institute of Infectious Diseases (FIOCRUZ, Brazil)

Oliveira¹,

Santos²,

Galdino³

et al. 2017

Rev. Soc. Bras. Med. Trop.

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confidence: 99%

Trypanosoma cruzi I genotype among isolates from patients with chronic Chagas disease followed at the Evandro Chagas National Institute of Infectious Diseases (FIOCRUZ, Brazil)

Oliveira¹,

Santos²,

Galdino³

et al. 2017

Rev. Soc. Bras. Med. Trop.

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“…Intraspecific variation in other species of Kudoa and myxozoa have been analysed by sequencing the small subunit, large subunit and internal transcribed spacer of nuclear rRNA gene and the heatshock protein 70 gene (Whipps & Kent 2006;Atkinson & Bartholomew 2010;Atkinson et al 2011;Heiniger, Cribb & Adlard 2013). Mitochondrial genome is known to be well conserved but to a lesser degree than the nuclear rRNA (Hwang & Kim 1999) and has been used, for example, in lineage typing of Trypanosoma cruzi (Messenger et al 2015) and in evolutionary study of human races (Cann, Stoneking & Wilson 1987). The advantages of mitochondrial genes for intraspecific genotyping are the higher DNA copy number compared with nuclear DNA, haploidy and moderate sequence conservation.…”

Section: Introductionmentioning

confidence: 99%

Genetic variants of Kudoa septempunctata (Myxozoa: Multivalvulida), a flounder parasite causing foodborne disease

Takeuchi

Ogasawara

Kato

et al. 2015

Journal of Fish Diseases

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Foodborne disease outbreaks caused by raw olive flounders (Paralichthys olivaceus) parasitized with Kudoa septempunctata have been reported in Japan. Origins of olive flounders consumed in Japan vary, being either domestic or imported, and aquaculture-raised or natural. Although it is unknown whether different sources are associated with different outcomes, it is desirable to identify whether this is the case by determining whether unique K. septempunctata strains occur and if so, whether some are associated with foodborne illness. We here developed an intraspecific genotyping method, using the sequence variation of mitochondrial genes. We collected olive flounder samples from foodborne disease outbreaks, domestic fish farms or quarantine offices and investigated whether K. septempunctata genotype is associated with pathogenicity or geographic origin. The 104 samples were classified into three genotypes, ST1, ST2 and ST3. Frequency of symptomatic cases differed by genotypes, but the association was not statistically significant. Whereas K. septempunctata detected from aquaculture-raised and natural fish from Japan were either ST1 or ST2, those from fish inspected at quarantine from Korea to Japan were ST3. Our method can be applied to phylogeographic analysis of K. septempunctata and contribute to containing the foodborne disease. The genotype database is hosted in the PubMLST website (http://pubmlst.org/kseptempunctata/).

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“…3 (TcVI), Chaco9 col. 15 (TcVI) and CL Brener (TcVI) was obtained from the London School of Hygiene & Tropical Medicine (LSHTM). DNA extractions at LSHTM followed Messenger et al 2015 40 .…”

Section: Methodsmentioning

confidence: 99%

Genome-wide locus sequence typing (GLST) of eukaryotic pathogens

Schwabl¹,

et al. 2020

Preprint

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20Analysis of genetic polymorphism is a powerful tool for epidemiological surveillance and research. Powerful 21 inference from pathogen genetic variation, however, is often restrained by limited access to representative 22 target DNA, especially in the study of obligate parasitic species for which ex vivo culture is resource-intensive 23 or bias-prone. Modern sequence capture methods enable pathogen genetic variation to be analyzed directly 24 from vector/host material but are often too complex and expensive for resource-poor settings where infectious 25 diseases prevail. This study proposes a simple, cost-effective 'genome-wide locus sequence typing' (GLST) 26 tool based on massive parallel amplification of information hotspots throughout the target pathogen genome. 27 The multiplexed polymerase chain reaction amplifies hundreds of different, user-defined genetic targets in a 28 single reaction tube, and subsequent agarose gel-based clean-up and barcoding completes library preparation 29 at under 4 USD per sample. Approximately 100 libraries can be sequenced together in one Illumina MiSeq 30 run. Our study generates a flexible GLST primer panel design workflow for Trypanosoma cruzi, the parasitic 31 agent of Chagas disease. We successfully apply our 203-target GLST panel to direct, culture-free 32 metagenomic extracts from triatomine vectors containing a minimum of 3.69 pg/µl T. cruzi DNA and further 33 elaborate on method performance by sequencing GLST libraries from T. cruzi reference clones representing 34 discrete typing units (DTUs) TcI, TcIII, TcIV, and TcVI. The 780 SNP sites we identify in the sample set 35 repeatably distinguish parasites infecting sympatric vectors and detect correlations between genetic and 36 geographic distances at regional (< 150 km) as well as continental scales. The markers also clearly separate 37 DTUs. We discuss the advantages, limitations and prospects of our method across a spectrum of 38 epidemiological research. 40Genome-wide single nucleotide polymorphism (SNP) analysis is a powerful and increasingly common 41 approach in the study and surveillance of infectious disease. Understanding patterns of SNP diversity within 42 pathogen genomes and across pathogen populations can resolve fundamental biological questions (e.g., 43 reproductive mechanisms in T. cruzi 1 , reconstruct past 2 and present transmission networks (e.g., 44 Staphylococcus infections within hospitals) 3 or identify the genetic bases of virulence 4,5 and resistance to drugs 45 (see examples from Plasmodium spp. 6,7 ). A number of obstacles, however, complicate access to 46 representative, genome-wide SNP information using modern sequencing tools. Micro-pathogens are often 47 sampled in low quantities and together with large amounts of host/vector tissue, microbiota, or environmental 48 DNA. Sequencing is rarely viable directly from the infection source and studies have often found it necessary 49 to isolate and culture the target organism to higher densities before extracting D...

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Molecular Genotyping of Trypanosoma cruzi for Lineage Assignment and Population Genetics

Cited by 22 publications

References 77 publications

Trypanosoma cruzi I genotype among isolates from patients with chronic Chagas disease followed at the Evandro Chagas National Institute of Infectious Diseases (FIOCRUZ, Brazil)

Trypanosoma cruzi I genotype among isolates from patients with chronic Chagas disease followed at the Evandro Chagas National Institute of Infectious Diseases (FIOCRUZ, Brazil)

Genetic variants of Kudoa septempunctata (Myxozoa: Multivalvulida), a flounder parasite causing foodborne disease

Genome-wide locus sequence typing (GLST) of eukaryotic pathogens

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