Dataset Information

Using the Plasmodium mitochondrial genome for classifying mixed-species infections and inferring the geographical origin of P. falciparum parasites imported to the U.S.

ABSTRACT: The ability to identify mixed-species infections and track the origin of Plasmodium parasites can further enhance the development of treatment and prevention recommendations as well as outbreak investigations. Here, we explore the utility of using the full Plasmodium mitochondrial genome to classify Plasmodium species, detect mixed infections, and infer the geographical origin of imported P. falciparum parasites to the United States (U.S.). Using the recently developed standardized, high-throughput Malaria Resistance Surveillance (MaRS) protocol, the full Plasmodium mitochondrial genomes of 265 malaria cases imported to the U.S. from 2014-2017 were sequenced and analyzed. P. falciparum infections were found in 94.7% (251/265) of samples. Five percent (14/265) of samples were identified as mixed- Plasmodium species or non-P. falciparum, including P. vivax, P. malariae, P. ovale curtisi, and P. ovale wallikeri. P. falciparum mitochondrial haplotypes analysis revealed greater than eighteen percent of samples to have at least two P. falciparum mitochondrial genome haplotypes, indicating either heteroplasmy or multi-clonal infections. Maximum-likelihood phylogenies of 912 P. falciparum mitochondrial genomes with known country origin were used to infer the geographical origin of thirteen samples from persons with unknown travel histories as: Africa (country unspecified) (n = 10), Ghana (n = 1), Southeast Asia (n = 1), and the Philippines (n = 1). We demonstrate the utility and current limitations of using the Plasmodium mitochondrial genome to classify samples with mixed-infections and infer the geographical origin of imported P. falciparum malaria cases to the U.S. with unknown travel history.

SUBMITTER: Schmedes SE

PROVIDER: S-EPMC6490880 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Using the Plasmodium mitochondrial genome for classifying mixed-species infections and inferring the geographical origin of P. falciparum parasites imported to the U.S.

Schmedes Sarah E SE Patel Dhruviben D Kelley Julia J Udhayakumar Venkatachalam V Talundzic Eldin E

PloS one 20190430 4

The ability to identify mixed-species infections and track the origin of Plasmodium parasites can further enhance the development of treatment and prevention recommendations as well as outbreak investigations. Here, we explore the utility of using the full Plasmodium mitochondrial genome to classify Plasmodium species, detect mixed infections, and infer the geographical origin of imported P. falciparum parasites to the United States (U.S.). Using the recently developed standardized, high-through ...[more]

PMID: 31039178

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Project description:BackgroundMalaria cases in non-endemic zero-indigenous case areas are most likely to have been imported whatever of the route of importation. In countries recently declared malaria-free and now without local transmission, imported cases remain a threat to re-introduction of the disease and a burden on the health system.Case presentationThree days after returning from a long trip to malaria- endemic countries; Abyei-Sudan, Chad and Uganda, a 41-year-old male resident from Jericho, Palestine, suffered paroxysms of fever, general fatigue, myalgia, arthralgia, headache, and a strong desire to vomit. Thin and thick Giemsa-stained blood smears were prepared and examined microscopically using oil immersion. Immature trophozoites (ring forms) were seen to parasitize approximately 10% of the erythrocytes revealing hyperparasitemia equivalent to > 100,000 parasites/ µl indicating severe malaria [1, 2]. The double chromatin configuration (headphones) and accolé (applique) position are both indicative of Plasmodium falciparum infection. The 18S rRNA- PCR targeting the rPLU6-rPLU5 region was used to confirm the diagnosis. The next-generation sequencing (NGS) method was carried out according to the manufacturer's instructions (Illumina® DNA Prep, (M) Tagmentation kit (20060060), Illumina) to identify Plasmodium spp. Furthermore, NGS produced a whole-genome sequence of 22.8Mbp of the 14 chromosomes and 25Kbp of the apicoplast. A BLAST search of the apicoplast DNA and selected chromosomal DNA revealed that P. falciparum was the causative agent. The merozoite surface protein-1 (msp-1) was used to construct a phylogenetic tree of 26 P. falciparum, including the one isolated from the patient from Jericho, which clustered with the Sudanese isolate indicating genetic relatedness between the two.ConclusionThe travel history together with signs and symptoms of malaria, followed by prompt diagnosis using conventional microscopic inspection of Giemsa-stained films together with molecular DNA tracking tools like msp-1 were key means in tracking the place of origin of infection in the case of travel to multiple destination.

Project description:BackgroundImported malaria parasites with anti-malarial drug resistance (ADR) from Africa is a serious public health challenge in non-malarial regions, including Wuhan, China. It is crucial to assess the ADR status in African Plasmodium falciparum isolates from imported malaria cases, as this will provide valuable information for rational medication and malaria control.MethodsDuring 2017-2019, a cross-sectional study was carried out in Wuhan, China. Peripheral blood 3 ml of returned migrant workers from Africa was collected. The target fragments from pfcrt, pfmdr1, and k13 propeller (pfk13) genes were amplified, sequenced, and analysed.ResultsIn total, 106 samples were collected. Subsequently, 98.11% (104/106), 100% (106/106), and 86.79% (92/106) of these samples were successfully amplified and sequenced for the pfcrt (72-76), pfmdr1, and pfk13 genes, respectively. The prevalence of the pfcrt 76 T, pfmdr1 86Y, and pfmdr1 184F mutations was 9.62, 4.72, and 47.17%, respectively. At codons 72-76, the pfcrt locus displayed three haplotypes, CVMNK (wild-type), CVIET (mutation type), CV M/I N/E K/T (mixed type), with 87.50%, 9.62%, and 2.88% prevalence, respectively. For the pfmdr1 gene, NY (wild type), NF and YF (mutant type), N Y/F, Y Y/F, and N/Y Y/F (mixed type) accounted for 34.91, 43.40, 3.77, 15.09, 0.94, and 1.89% of the haplotypes, respectively. A total of 83 isolates with six unique haplotypes were found in pfcrt and pfmdr1 combined haplotypes, of which NY-CVMNK and NF-CVMNK accounted for 40.96% (34/83) and 43.37% (36/83), respectively. Furthermore, 90 cases were successfully sequenced (84.91%, 90/106) at loci 93, 97, 101, and 145, and 78 cases were successfully sequenced (73.58%, 78/106) at loci 343, 353, and 356 for pfcrt. However, the mutation was observed only in locus 356 with 6.41%. For pfk13, mutations reported in Southeast Asia (at loci 474, 476, 493, 508, 527, 533, 537, 539, 543, 553, 568, 574, 578, and 580) and Africa (at loci 550, 561, 575, 579, and 589) were not observed.ConclusionsThe present data from pfcrt and pfmdr1 demonstrate that anti-malarial drugs including chloroquine, amodiaquine, and mefloquine, remain effective against malaria treatment in Africa. The new mutations in pfcrt related to piperaquine resistance remain at relatively low levels. Another source of concern is the artemether-lumefantrine resistance-related profiles of N86 and 184F of pfmdr1. Although no mutation in pfk13 is detected, molecular surveillance must continue.

Project description:Background: The cytoadherence of Plasmodium falciparum is thought to be mediated by variant surface antigens (VSA), encoded by var, rif, stevor and pfmc-2tm genes. The last three families have rarely been studied in the context of cytoadherence. As most VSA genes are unique, the variability among sequences has impeded the functional study of VSA across different P. falciparum strains. However, many P. falciparum genomes have recently been sequenced, allowing the development of specific microarray probes to each VSA gene. Methods: All VSA sequences from the HB3, Dd2 and IT/FCR3 genomes were extracted using HMMer. Oligonucleotide probes were designed with OligoRankPick and added to the 3D7-based microarray chip. As a proof of concept, IT/R29 parasites were selected for and against rosette formation and the transcriptomes of isogenic rosetting and non-rosetting parasites were compared by microarray. Results: From each parasite strain 50-56 var genes, 125-132 rif genes, 26-33 stevor genes and 3-8 pfmc-2tm genes were identified. The ability of the VSA-supplemented microarray chip to detect cytoadherence-related genes was assessed using P. falciparum clone IT/R29, in which rosetting is known to be mediated by PfEMP1 encoded by ITvar9. Whole transcriptome analysis showed that the most highly upregulated gene in rosetting parasites was ITvar9 (19 to 429-fold upregulated over six time points). Only one rif gene (IT4rifA_042) was upregulated by more than 4-fold (5-fold at 12 hours post-invasion), and no stevor or pfmc-2tm genes were upregulated by more than 2-fold. 49 non-VSA genes were upregulated in rosetting parasites by more than 3-fold in at least two time-points, although none as markedly as ITvar9. Conclusions: We demonstrate that the VSA of newly sequenced P. falciparum strains can be added to the 3D7-based microarray chip, allowing the analysis of the entire transcriptome of multiple strains. For the rosetting clone IT/R29, the striking transcriptional upregulation of ITvar9 was confirmed, and the data did not support the involvement of other VSA families in rosette formation.

Dataset Information

Using the Plasmodium mitochondrial genome for classifying mixed-species infections and inferring the geographical origin of P. falciparum parasites imported to the U.S.

Publications

Using the Plasmodium mitochondrial genome for classifying mixed-species infections and inferring the geographical origin of P. falciparum parasites imported to the U.S.

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