Implementation of a novel PCR based method for detecting malaria parasites from naturally infected mosquitoes in Papua New Guinea

Hasan, Azeem; Suguri, Setsuo; Sattabongkot, Jetsumon; Fujimoto, Chigusa; Amakawa, Masao; Harada, Masakazu; Ohmae, Hiroshi

doi:10.1186/1475-2875-8-182

Cited by 23 publications

(29 citation statements)

References 46 publications

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“…Others have also found the cytochrome b method to be more sensitive than 18S rDNA nested PCR (13,30), likely because there are more copies of the gene in each parasite (15,25). It also performed better than two other single-round PCR methods, including one real-time method.…”

Section: Discussionmentioning

confidence: 99%

PCR-Based Pooling of Dried Blood Spots for Detection of Malaria Parasites: Optimization and Application to a Cohort of Ugandan Children

Hsiang¹,

Lin²,

Dokomajilar

et al. 2010

J Clin Microbiol

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Sensitive, high-throughput methods to detect malaria parasites in low-transmission settings are needed. PCR-based pooling strategies may offer a solution. We first used laboratory-prepared samples to compare 2 DNA extraction and 4 PCR detection methods across a range of pool sizes and parasite densities. Pooled Chelex extraction of DNA, followed by nested PCR of cytochrome b, was the optimal strategy, allowing reliable detection of a single low-parasitemic sample (100 parasites/l) in pool sizes up to 50. This PCR-based pooling strategy was then compared with microscopy using 891 dried blood spots from a cohort of 77 Ugandan children followed for 2 years in an urban setting of low endemicity. Among 419 febrile episodes, 35 cases of malaria were detected using the PCR-based pooling strategy and 40 cases using microscopy. All five cases of malaria not detected by PCR were from samples stored for >2 years with parasitemia of <6,000/l, highlighting the issue of possible DNA degradation with long-term storage of samples. Among 472 samples collected from asymptomatic children as part of routine surveillance, 15 (3.2%) were positive by PCR-based pooling compared to 4 (0.8%) by microscopy (P ‫؍‬ 0.01). Thus, this PCR-based pooling strategy for detection of malaria parasites using dried blood spots offers a sensitive and efficient approach for malaria surveillance in low-transmission settings, enabling improved detection of asymptomatic submicroscopic infections and dramatic savings in labor and costs.Due to large-scale implementation of effective control measures, many countries where malaria is endemic are experiencing dramatic declines in disease burden. With this success has come a shift in the end goal from control to elimination (9, 10). However, when the goal is elimination, accurate detection of persons infected with malaria parasites becomes essential (11). Standard surveillance systems depend on diagnosis by microscopy, a method that is technically challenging, labor-intensive, and often inaccurate in operational settings. More recently available rapid diagnostic tests (RDTs) provide convenience and ease of use, but they have limitations in specificity, sensitivity, species identification, and cost (22).PCR-based methods for malaria parasite detection are relatively simple and provide improved sensitivity compared to microscopy and RDTs, especially in settings where infections have low parasitemia or contain mixed species (22). PCR can also be performed on dried blood spots, which are convenient for collection, storage, and transport. Nonetheless, PCR has a long turnaround time, making it an impractical tool for clinical care of individual patients. One exception would be real-time PCR, which has a short turnaround time but carries with it cost and capacity constraints that make it unavailable for rapid diagnosis in most settings. However, there is a potential role for PCR in situations where large numbers of samples are being screened: a high-throughput system could allow accurate, rapid, and cost-effective ass...

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Section: Discussionmentioning

confidence: 99%

PCR-Based Pooling of Dried Blood Spots for Detection of Malaria Parasites: Optimization and Application to a Cohort of Ugandan Children

Hsiang¹,

Lin²,

Dokomajilar

et al. 2010

J Clin Microbiol

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“…The DNA extraction was performed at the UCSD-PRISMA Laboratory, Iquitos, from the head+thorax portion of each specimen following the manufacturer's instructions (Qiagen DNA blood tissue extraction kit, Valencia, CA). Aliquots of the head+thorax DNA were used for the Plasmodium Cyt-B-PCR protocol 18 for detection of parasites in Iquitos, and for the molecular identification of the anopheline species at the Wadsworth Center in Albany, New York. The latter procedure was undertaken with the PCR-fragment length polymorphism-internal transcribed spacer 2 (RFLP-ITS2) using the restriction endonucleases Alu I and BsrB I as previously described.…”

Section: Introductionmentioning

confidence: 99%

Molecular Taxonomy of Anopheles (Nyssorhynchus) benarrochi (Diptera: Culicidae) and Malaria Epidemiology in Southern Amazonian Peru

Conn

Moreno²,

Saavedra³

et al. 2013

The American Society of Tropical Medicine and Hygiene

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Abstract. Anopheline specimens were collected in 2011 by human landing catch, Shannon and CDC traps from the malaria endemic localities of Santa Rosa and San Pedro in Madre de Dios Department, Peru. Most specimens were either Anopheles (Nyssorhynchus) benarrochi B or An. (Nys.) rangeli, confirmed by polymerase chain reaction-restriction fragment length polymorphism-internal transcribed spacer 2 (PCR-RFLP-ITS2) and, for selected individuals, ITS2 sequences. A few specimens from Lupuna, Loreto Department, northern Amazonian Peru, were also identified as An. benarrochi B. A statistical parsimony network using ITS2 sequences confirmed that all Peruvian An. benarrochi B analyzed were identical to those in GenBank from Putumayo, southern Colombia. Sequences of the mtDNA COI BOLD region of specimens from all three Peruvian localities were connected using a statistical parsimony network, although there were multiple mutation steps between northern and southern Peruvian sequences. A Bayesian inference of concatenated Peruvian sequences of ITS2+COI detected a single clade with very high support for all An. benarrochi B except one individual from Lupuna that was excluded. No samples were positive for Plasmodium by CytB-PCR.

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“…New Jersey light traps and Mosquito Magnets were set each afternoon between 1700 and 1730 h, and the collections were picked up the following morning between 0800 and 0830 h. Resting adults were caught using aspirators inside the cattle shed, particularly from the walls, roof, and other parts of the building, between 2000 and 2200 h. Collected specimens were removed, taken to the laboratory, sorted, initially identified morphologically (Tanaka et al 1979, Rueda 2005, and stored in the freezer prior to polymerase chain reaction (PCR) assay. Only the head and thorax of mosquitoes were assayed by PCR for species identification and confirmation (Wilkerson et al 2003, Li et al 2005 and by single step and seminested multiplex-PCR to identify P. vivax sporozoite infections (Rubio et al 1999, Hasan et al 2009). The abdomen was separated from the mosquito body prior to assay to reduce the probability of detecting DNA from oocysts in the hind gut.…”

mentioning

confidence: 99%

Anopheles belenrae, a Potential Vector of Plasmodium vivax in the Republic of Korea

Rueda

Li²,

Kim³

et al. 2010

Journal of the American Mosquito Control Association

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Implementation of a novel PCR based method for detecting malaria parasites from naturally infected mosquitoes in Papua New Guinea

Cited by 23 publications

References 46 publications

PCR-Based Pooling of Dried Blood Spots for Detection of Malaria Parasites: Optimization and Application to a Cohort of Ugandan Children

PCR-Based Pooling of Dried Blood Spots for Detection of Malaria Parasites: Optimization and Application to a Cohort of Ugandan Children

Molecular Taxonomy of Anopheles (Nyssorhynchus) benarrochi (Diptera: Culicidae) and Malaria Epidemiology in Southern Amazonian Peru

Anopheles belenrae, a Potential Vector of Plasmodium vivax in the Republic of Korea

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