Saliva as a proven, non-invasive sample type formolecular malaria testing and surveillance using
OMNIgene®•DISCOVER at ambient temperature
David Visanuvimol

DNA Genotek, Ottawa, Ontario, Canada
2014-09-19

Background
Saliva collection is a non-invasive alternative from RDT’s for malaria detection and as asurveillance tool. Both P. falciparum and P. vivax DNA have been detected in saliva samples ofmalaria patients.1, 2, 3, 4 It has been demonstrated that 1 mL of whole saliva samples harbours
detectable levels of Plasmodium spp. DNA for downstream sequencing of pfdhfr and 18S rRNA.5, 6
Therefore, saliva can be sampled for high sensitivity and specificity molecular-based malaria
diagnosis. Additionally, participant compliance is increased with pain-free and easy saliva
collection; therefore, providing greater access to Plasmodium DNA for improved monitoringof malaria transmission, identification of sub-patent or mixed Plasmodium species infections,and patient screening in artemisinin-resistance-emerging regions/elimination settings.

OMNIgene®•DISCOVER for sample stability
and testing scalability
OMNIgene•DISCOVER enables non-invasive and pain-free collection of Plasmodium spp.
via saliva. The OMNIgene•DISCOVER chemistry stabilizes Plasmodium DNA in 1 mL of saliva
at ambient temperature for up to 1 year, eliminating the cost and complexity of cold storage
making it ideal for field collection in remote and low-resource settings. The easy-to-use andreliable nature of the OMNIgene•DISCOVER kits improves patient compliance in both adultsand children.
The following data is from ongoing pilot studies that show promising indications forthe use of OMNIgene•DISCOVER as a non-invasive alternative for malaria diagnostics.
P. falciparum DNA from 1 mL of saliva is a reliable sample typefor malaria detection (n=100)(Courtesy of Dr. Collins Ouma, Maseno University, Kenya, as presented
at the 6th MIM Pan-African Malaria Conference in Durban, South African).

ID

Temperature (°C)

Microscopy

mRDT

PCR

21179

37.3

0/100

Negative

Negative

21166

38.3

Pf.313/200

Positive

Negative

21048

37.8

0/100

Negative

Negative

21355

39.0

Pf.1744/200

Positive

Positive

21165

37.6

Pf.158/200

Positive

Positive

21356

37.9

Pf.1064/200

Positive

Positive

Table 1: Patient information, microscopy, mRDT and PCR results.

“The OMNIgene•DISCOVER kit proved successful in the majority of the tested samples
with comparable results as detected by mRDT and microscopy. Therefore, thenon-invasive OMNIgene•DISCOVER kit offers potential for use in malaria diagnosis.”- Dr. Deus Ishengoma
P. falciparum DNA collected from febrile participants shows high specificity
and sensitivity compared to blood (n=11)(Courtesy of Dr. Kenji Obadiah Mfuh, John A. Burns School of Medicine,University of Hawaii at Manoa, USA)120.00%100.00%80.00%
60.00%
40.00%

Saliva PCR

20.00%RDT results

0.00%
Saliva PCR

Specificity
87.50%

Sensitivity
100%

PPV
75%

NPV
100%

RDT results

25%

100%

33.30%

100%“Malaria parasite DNA is present in the saliva of infected individuals and that saliva
can be used as an alternative for non-invasive sample for the diagnosis of malaria
in a PCR-based reaction. This pilot study also proves that the OMNIgene•DISCOVER
kit is very effective in preserving malaria DNA at room temperature.”- Dr. Kenji Obadiah Mfuh
S= Pf PCR from OMNIgene/saliva kit
O= Pf PCR using DNA obtained from blood
M= Molecular weight marker

Additional testimony“We have successfully extracted P. falciparum from the OM-501 kits. Parasite genetic
diversity using MSP1 and MSP2 allelic families was also determined. I’m happy to saythat we have sufficient amount of DNA for genetic analysis from the kits (n=69).”

Figure 1: PCR results demonstrate DNA from the OMNIgene/saliva sample kits performs
as well as DNA from blood in detection of circulating Plasmodium falciparum parasites.

“I recommend that future approaches should utilize OMNIgene•DISCOVER self-collection
kits to avoid invasive sample collections, improve patient recruitment, improve thepatient experience, and enhance malaria diagnostics and research.”- Dr. Collins Ouma
1 mL of saliva stabilized in OMNIgene•DISCOVER can detect P. falciparumin symptomatic patients ranging from 6,000-70,000 parasites/µL (n=6)(Courtesy of Dr. Deus Ishengoma, National Malaria Research Council, Tanzania).

13 12 11 1098765432- Dr. Magatte Ndiaye, Université Cheikh Anta Diop (University of Dakar), Senegalon the use of OM-501 for temporal dynamics of molecular markers of anti-malarial
drug resistance in P. falciparum parasite populations in Senegal.

Conclusions
Given that only 14% of malaria cases are detected globally and several limitations exist tocurrent diagnostic testing methods, there is a need to explore other approaches that enhance
malaria diagnostics and national surveillance programs. OMNIgene•DISCOVER is a saliva-based
collection and stabilization kit that allows for the non-invasive, proactive sampling and storageof Plasmodium DNA at ambient temperature.

1

References

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21179 (saliva sample)
21166 (saliva sample)
21048 (saliva sample)
21355 (saliva sample)
21165 (saliva sample)
21356 (saliva sample)
Negative control

Interpretations key
8. Negative control
9. Positive P. falciparum control Pf (3D7)
10. Positive P. falciparum control (FCR3)
11. P. falciparum known sample
12. Positive P. malariae control
13. Negative control (ddH20)
14. 50 bp ladder marker

Figure 2: Agarose gel electrophoresis results of samples and control from PCR products.

Superior samples
Proven performance

1 Putaporntip et al. Improved performance with saliva and urine as alternative DNA
sources for malaria diagnosis by mitochondrial DNA-based PCR assays. Clin Microbiol
Infect 17: 1484–1491 (2011)
2 Nwakanma et al. Quantitative Detection of Plasmodium falciparum DNA in Saliva, Blood,and Urine. J Infect Dis 199:1567–74 (2009)
3 Gbotosho et al. Rapid Detection of Lactate Dehydrogenase and Genotyping of Plasmodium
falciparum in Saliva of Children with Acute Uncomplicated Malaria. Am J Trop Med Hyg 83:
496–501 (2010)
4 Singh et al. Rapid detection of Plasmodium vivax in saliva and blood using loop mediatedisothermal amplification (LAMP) assay. J Infect 67:245-247 (2013)
5 Ongagna-Yhombi et al. Improved assay to detect Plasmodium falciparum using anuninterrupted, semi-nested PCR and quantitative lateral flow analysis. Malaria Journal
12:74 (2013)
6 Singh et al. Comparison of three PCR-based assays for the non-invasive diagnosisof malaria: detection of Plasmodium parasites in blood and saliva. Eur J Clin Microbiol
Infect Dis 33:1631–1639 (2014)© 2014 DNA Genotek Inc., a subsidiary of OraSure Technologies, Inc., all rights reserved.
OMNIgene®•DISCOVER is for research use only, not for use in diagnostic procedures.
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®OMNIgene is a registered trademark of DNA Genotek Inc. All brands and names contained herein are the property of their respective owners.
Patent (www.dnagenotek.com/legalnotices) MK-00430 Issue 2/2014-10

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