Research Journal of Recent Sciences _________________________________________________ ISSN 2277-2502
Vol. 1 (ISC-2011), 361-364 (2012)
Res.J.Recent Sci.

Short communication

Evaluation of one Rapid Method for Diagnosis of Malaria
The Optional and Better Replacement of Microscopy
Panchal Hetal K.1 and Desai Pratibha B.2
1

Dolat Usha Institute of Applied Sciences, Valsad; Veer Narmad South Gujarat University, Surat, Gujarat, INDIA
2
Shree Ram Krishna Institute of Comp. Edu. and Applied Sciences; Veer Narmad South Gujarat University, Surat, Gujarat, INDIA

Available online at, www.isca.in
(Received 27th September 2011, revised 16th January 2012, accepted 28th January 2012)

Abstract
Microscopy has been the most trustable technique for the diagnosis of malaria in India. Reduction of morbidity and mortality rate
of malaria highly influenced by earlier and proper diagnosis. This study was carried out at Valsad, Gujarat. It involved use of
microscopy i.e. field’s stain and detection of Plasmodium falciparum - HRP II antigen, Plasmodium vivax - pLDH antigen detection
by one rapid diagnostic Test (RDT) SD Bioline. Present study was carried out from 966 EDTA anticoagulated samples collected
from clinical laboratories and hospitals of Valsad. Microscopic examinations of stained thick and thin films, shows 8.39%, 13.97%,
0.21% were detected as Plasmodium falciparum, P. vivax and mix respectively. Whereas with Rapid Diagnostic test using HRP II,
p-LDH antigens 9.05% and 13.87% were detected as P. falciparum, P. vivax respectively. The study shows reasonable harmony
between microscopy and RDT. Among two methods RDT was found to have high sensitivity (97.70%) and specificity (98.93%)
compared to microscopy. Though the microscopic method is cost effective but laborious and needs an expertise. The RDT results
were highly accurate and can be used where microscopy is inaccurate or in case of unavailability of expert.
Keywords: Malaria, malaria diagnosis, rdt, microscopy.

Introduction
Malaria is most important parasitic disease in tropical areas.
Around 300 million malaria cases reported each year in the
world, causes 1 to 3 million deaths. Nearly 3 billion people
lives with the risk of malaria. In India during running year
2011 total malaria cases reported were 336,545. Among them
53.75% were due to Plasmodium falciparum and rest of the
cases were due to P. vivax1. The disease is caused by
Plasmodium species namely P. falciparum, P. vivax, P. ovale
and P. malariae transmitted through biting of female
Anopheles mosquito2,3. Valsad district of Gujarat state, India
is considered as one of the malaria endemic area4.
Microscopic examination of thick and thin blood smears
stained with Romanosky’s stain is the most common
technique to diagnose malaria since last hundred years5,6.
Microscopy continues to be the gold standard for
identification of Plasmodium species in the laboratories7. The
method is easy to apply and cost effective in the laboratories
where skilled professionals are available who can even detect
very low level of parasite like 10 to 50 parasites/µl. So the
sensitivity of microscopy may fluctuate depending upon the
skill of technician. In these consequences WHO has
recognized the need for simple and cost effective diagnostic
test for malaria to overcome the deficiencies of microscopy
and clinical diagnosis3. To overcome this problem one most
easy, cheaper, faster and reliable method available is Rapid

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Diagnostic Test (RDT). RDT detects P. falciparum HistidineRich-Protein II (HRP II) antigen and parasite Lactate
Dehydrogenase (pLDH) antigen present in all four
Plasmodium species8,9. It does not require any special
equipment and give results within 15 to 30 minutes10,11. To
estimate the impact of RDT SD Bioline on malaria diagnosis,
we analyzed all samples to see the difference between
conventional diagnostic microscopy and RDT SD Bioline.

Material and Methods
Total 966 samples were collected from various clinical
laboratories of Valsad, Gujarat during December 2010 and
July to September 2011. Approximately 1 ml blood sample
was collected from each patient in a vaccutainer containing
an anticoagulant EDTA. All samples were tested by both
diagnostic methods microscopy and RDT. Thick and thin
smears were prepared on slide, stained with Field’s stain B
and A for 5 and 12 seconds respectively12. Thick smears
were used to confirm malaria and to count parasites/µl.
Smears were considered negative if no parasite was observed
in 200 consecutive fields of thick smear in oil immersion
objective. Parasites were counted against 200 to 500
leucocytes. For the parasite estimation it was assumed that
8000 leucocytes present in 1 µl of blood 13,14. Thin smears
were used to identify and differentiate parasites. RDT SD
Bioline malaria antigen detection test was purchased from
SD Bio Standard Diagnostic Pvt. Ltd. The test cassette

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Research Journal of Recent Sciences ____________________________________________________________ ISSN 2277-2502
Vol. 1 (ISC-2011), 361-364 (2012)
Res.J.Recent Sci.
contains a membrane strip, precoated with one monoclonal
antibody against Plasmodium falciparum HRP II antigen and
other polyclonal antibodies specific to pLDH of all 4 human
malaria Plasmodium species as two separate lines. The test is
one step, rapid, qualitative and differential for Plasmodium
falciparum and Plasmodium vivax, the two prominent
parasites found in India. For all samples, the cassettes were
removed from pouch, approximately 5µl blood samples were
placed in small, circular wells with loop, and 4 drops of
assay diluent were placed in square assay diluent wells.
Results were recorded at the end of 15 minutes and
maximum within 30 minutes.

Figure- 1
Microscopic Observation of Malarial Parasites

Results and Discussions
Out of 966 blood samples 213 were recognized positive by
both tests. Total 218 and 221 cases were found positive by
Microscopy and RDT respectively. Detail results are shown
in Table 1 and 2. Figures showing observation of ICT and
microscopy are shown as figure 1 and 2 respectively. The
sensitivity and specificity of the test found was also high
about 97.70% and 98.93% respectively. 08 samples were
detected as false positive and 05 samples were detected as
false negative. Comparison of microscopy and RDT results
are also shown in graph 1.
Table - 1
Results of Microscopy and RDT analysis
Technique

Positive for
P. f

Microscopy

81 (8.39%)

RDT

87 (9.05%)

Positive for
P. v
135
(13.97%)
134
(13.87%)

Positive for
mix
02 (0.21%)
00 (0.00%)

Negative
748
(77.43%)
745
(77.12%)

Table -2
Results of Microscopy and RDT analysis
Microscopy/RDT Positive Negative Total
213
05
218
Positive
08
740
748
Negative
221
745
966
Total

150

Total
966
966

Figure-2
Rapid Diagnostic Test Results
In this study RDT has shown high level of agreement with
microscopy. The sensitivity and specificity of the test was
also very high. Out of 08 false negative patients 05 were
previously treated with chemoprophylaxis against malaria.
Even in microscopic analysis of these samples the
parasites/µl counts were as low as like 60, 311, 170, 296, 318
etc. Compared with microscopic diagnosis, the SD Bioline
RDT was found false positive in 05 patients. This may be
due to persistent antigenemia following treatment of malaria
in 03 patients with recent history of malaria. Because in
some cases antigenemia may remain positive 3-28 days after
disappearance of circulating parasites15. Other 02 false
positive results may be due to rheumatoid factor, hepatitis etc
16,17,18,19
. This may be due to a non specific reaction of
rheumatoid factor, hepatitis antigens with coated antibodies.

100

Microscopy

50

RDT

0

P. f

P. v

Mix

Graph-1
Comparative Results of Microscopy and RDT

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Our study was valuable because the sample size was quite
large about to access the acceptability of RDT. Microscopy
involves good time and tough microscopic observation.
Skilled professional is required to observe the same.
Sometime when parasitemia is very low even a keen
observation may lead to false negative diagnosis. It is also
not easy to differentiate different Plasmodium species
without ample experience. In rural areas where skilled
malaria detecting experts are not available diagnosis may be
delayed and lead to improper diagnosis of malaria and there

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Research Journal of Recent Sciences ____________________________________________________________ ISSN 2277-2502
Vol. 1 (ISC-2011), 361-364 (2012)
Res.J.Recent Sci.
by treatment which some time even lead to death. In this
situation alternatively we suggest RDT as optional method to
diagnose malaria. RDT can be performed within couple of
minutes. It is easy to perform that even a new lab technician
or a layman can also perform it and interpret the results.
Even the RDT was able to differentiate between P.
falciparum and P. vivax. It is unable to differentiate P. vivax,
P. ovale and P. malariae but in India malaria is mainly
caused due to P. vivax and P. falciparum. The sensitivity and
specificity of the test found was also high. It suggests RDT
as a better option of microscopy in diagnosis of malaria.

Conclusion
In India malaria is mainly found due to P. falciparum and p.
vivax. Both can be differentiated well by RDT. Results
obtained by RDT are suggesting that it can be used for
malaria diagnosis. It makes diagnosis faster, better and
reliable. Even can be used at areas where experts are not
available or results are needed in emergency.

Acknowledgment
The authors wish to thank to staff of all clinical laboratories
of Valsad district, who gave their valuable support. The
authors also wish to thank management and staff of Shree
RamKrishna Institute of Computer Education and Applied
Sciences, Surat; Dolat Usha Institute of Applied Sciences,
Valsad.

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Res.J.Recent Sci.
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