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Grijalva, M. J.

Publications and source records attributed to Grijalva, M. J..

5 recordsLinked to original sources

The genomic basis of domestic colonisation and dispersal in Chagas disease vectors

The biology of vector adaptation to the human habitat remains poorly understood for many arthropod-borne diseases but underpins effective and sustainable disease control. We adopted a landscape genomics approach to investigate gene flow, signatures of local adaptation, and drivers of population structure among multiple linked wild and domestic population pairs in Rhodnius ecuadoriensis, an important vector of Chagas Disease. Evidence of high triatomine gene flow (FST) between wild and domestic ecotopes at sites throughout the study area indicate insecticide-based control will be hindered by constant re-infestation of houses. Genome scans revealed genetic loci with strong signal of local adaptation to the domestic setting, which we mapped to annotated regions in the Rhodnius prolixus genome. Our landscape genomic mixed effects models showed Rhodnius ecuadoriensis population structure and connectivity is driven by landscape elevation at a regional scale. Our ecologically- and spatially-explicit vector dispersal model enables targeted vector control and recommends spatially discrete, periodic interventions to local authorities as more efficacious than current, haphazard approaches. In tandem, evidence for parallel genomic adaptation to colonisation of the domestic environment at multiple sites sheds new light on the evolutionary basis of adaptation to the human host in arthropod vectors.

genomics↗

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

Analysis of genetic polymorphism is a powerful tool for epidemiological surveillance and research. Powerful inference from pathogen genetic variation, however, is often restrained by limited access to representative target DNA, especially in the study of obligate parasitic species for which ex vivo culture is resource-intensive or bias-prone. Modern sequence capture methods enable pathogen genetic variation to be analyzed directly from vector/host material but are often too complex and expensive for resource-poor settings where infectious diseases prevail. This study proposes a simple, cost-effective genome-wide locus sequence typing (GLST) tool based on massive parallel amplification of information hotspots throughout the target pathogen genome. The multiplexed polymerase chain reaction amplifies hundreds of different, user-defined genetic targets in a single reaction tube, and subsequent agarose gel-based clean-up and barcoding completes library preparation at under 4 USD per sample. Approximately 100 libraries can be sequenced together in one Illumina MiSeq run. Our study generates a flexible GLST primer panel design workflow for Trypanosoma cruzi, the parasitic agent of Chagas disease. We successfully apply our 203-target GLST panel to direct, culture-free metagenomic extracts from triatomine vectors containing a minimum of 3.69 pg/l T. cruzi DNA and further elaborate on method performance by sequencing GLST libraries from T. cruzi reference clones representing discrete typing units (DTUs) TcI, TcIII, TcIV, and TcVI. The 780 SNP sites we identify in the sample set repeatably distinguish parasites infecting sympatric vectors and detect correlations between genetic and geographic distances at regional (< 150 km) as well as continental scales. The markers also clearly separate DTUs. We discuss the advantages, limitations and prospects of our method across a spectrum of epidemiological research.

genetics↗

Remarkable genetic diversity of Trypanosoma cruzi and Trypanosoma rangeli in two localities of southern Ecuador identified via deep sequencing of mini-exon gene amplicons

Trypanosoma cruzi, causative agent of Chagas disease, and T. rangeli are kinetoplastid parasites endemic to Latin America. Although closely-related to T. cruzi and capable of infecting humans, T. rangeli is non-pathogenic. Both parasite species are transmitted by triatomine bugs, and the presence of T. rangeli constitutes a confounding factor in the study of Chagas prevalence and transmission dynamics. T. cruzi possesses high molecular heterogeneity: six discrete typing units (DTUs) are currently recognized. In Ecuador, TcI predominates while other DTUs are seldom reported. Here, infection by T. cruzi and/or T. rangeli in triatomine bugs from two communities of southern Ecuador was evaluated via PCR product size polymorphism of kinetoplast-minicircle sequences and the non-transcribed spacer region of the mini-exon gene. Additionally, mini-exon amplicons were deep-sequenced to analyze single nucleotide polymorphisms and mixed infections. As a control, mini-exon products from several monoclonal reference strains were included. T. cruzi genetic diversity was significantly greater in adult vectors than in nymphal stage V vectors. Among infected triatomines, deep sequencing revealed one T. rangeli infection (3%), 9 T. cruzi infections (27.3%) and 23 T. cruzi / T. rangeli mixed infections (69.7%), suggesting that T. rangeli prevalence has been largely underestimated in the region. Furthermore, deep-sequencing detected TcIV sequences in six samples (first TcIV record in southern Ecuador). Our data indicate that amplicon size analysis alone is not reliable for parasite identification/typing in mixed infections containing both T. cruzi and T. rangeli, or when multiple T. cruzi DTUs are present. Additionally, our analysis showed extensive overlap among the parasite populations present in the two studied localities (ca. 28 km apart); suggesting active parasite dispersal over the study area. Our results highlight the value of amplicon sequencing methodologies to clarify the population dynamics of kinetoplastid parasites in endemic regions and inform control campaigns in southern Ecuador.

genetics↗

Parallel sexual and parasexual population genomic structure in Trypanosoma cruzi

Genetic exchange and hybridization in parasitic organisms is fundamental to the exploitation of new hosts and host populations. Variable mating frequency often coincides with strong metapopulation structure, where patchy selection or demography may favor different reproductive modes. Evidence for genetic exchange in Trypanosoma cruzi over the last 30 years has been limited and inconclusive. The reproductive modes of other medically important trypanosomatids are better established, although little is known about their variability on a spatio-temporal scale. Targeting a contemporary focus of T. cruzi transmission in southern Ecuador, we present compelling evidence from 45 sequenced genomes that T. cruzi (discrete typing unit I) maintains sexual populations alongside others that represent clonal bursts of parasexual origin. Strains from one site exhibit genome-wide Hardy-Weinberg equilibrium and intra-chromosomal linkage decay consistent with meiotic reproduction. Strains collected from adjacent areas (>6 km) show excess heterozygosity, near-identical haplo-segments, common mitochondrial sequences and levels of aneuploidy incompatible with Mendelian sex. Certain individuals exhibit trisomy in as many as fifteen chromosomes. Others present fewer, yet shared, aneuploidies reminiscent of mitotic genome erosion and parasexual genetic exchange. Genomic and intra-genomic phylogenetics as well as haplotype co-ancestry analyses indicate a clear break in gene-flow between these distinct populations, despite the fact that they occasionally co-occur in vectors and hosts. We propose biological explanations for the fine-scale disconnectivity we observe and discuss the epidemiological consequences of flexible reproductive modes and their genomic architecture for this medically important parasite.

evolutionary biology↗

Repeat-driven generation of antigenic diversity in a major human pathogen, Trypanosoma cruzi

Trypanosoma cruzi, a zoonotic kinetoplastid protozoan with a complex genome, is the causative agent of American trypanosomiasis (Chagas disease). The parasite uses a highly diverse repertoire of surface molecules, with roles in cell invasion, immune evasion and pathogenesis. Thus far, the genomic regions containing these genes have been impossible to resolve and it has been impossible to study the structure and function of the several thousand repetitive genes encoding the surface molecules of the parasite. We here present an improved genome assembly of a T. cruzi clade I (TcI) strain using high coverage PacBio single molecule sequencing, together with Illumina sequencing of 34 T. cruzi TcI isolates and clones from different geographic locations, sample sources and clinical outcomes. Resolution of the surface molecule gene structure reveals an unusual duality in the organisation of the parasite genome, a core genomic region syntenous with related protozoa flanked by unique and highly plastic subtelomeric regions encoding surface antigens. The presence of abundant interspersed retrotransposons in the subtelomeres suggests that these elements are involved in a recombination mechanism for the generation of antigenic variation and evasion of the host immune response. The comparative genomic analysis of the cohort of TcI strains revealed multiple cases of such recombination events involving surface molecule genes and has provided new insights into T. cruzi population structure.

genomics↗