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European Journal of Applied Sciences – Vol. 11, No. 5
Publication Date: October 25, 2023
DOI:10.14738/aivp.115.15370
Mwongula, A. W., Siamba, D. N., Mwamburi, L. A., & Mailu, M. (2023). Concurrent Infections Among Pyretic Children Seeking
Treatment at Alupe Sub County Hospital, Busia County, Kenya. European Journal of Applied Sciences, Vol - 11(5). 38-45.
Services for Science and Education – United Kingdom
Concurrent Infections Among Pyretic Children Seeking
Treatment at Alupe Sub County Hospital, Busia County, Kenya
Albert W. Mwongula
Department of Biological and Agricultural Sciences, School of Science, Technology
and Engineering (SSTE), Alupe University P.O. Box 845, 50400, Busia, Kenya
Donald N. Siamba
Department of Biological and Environmental Sciences, Faculty of
Science, Kibabii University, P.O. Box 1699 – 50200, Bungoma, Kenya
Lizzy A. Mwamburi
Department of Biological Sciences, School of Science,
University of Eldoret, P.O. Box 1125 – 30100, Eldoret, Kenya
Matilu Mailu
Centre for Infectious Parasitic Diseases Control Research,
Kenya Medical Research Institute, P.O Box 3 – 50400, Busia, Kenya
ABSTRACT
Fever is a frequently occurring medical symptom and may result from many
divergent conditions ranging from mild to potentially serious. Children typically get
high and fast – occurring fevers, reflecting the effects of the pyrogens upon an
inexperienced immune system. Symptoms and signs of Chikungunya virus
infections are quite similar to those of malaria and typhoid fever. Malaria and
typhoid investigations are routinely carried out to establish the cause of pyrexia of
unknown origin and treatment follows with complete neglect of Chikungunya virus
infections. Thus, Chikungunya virus fever cases can sometimes be misdiagnosed or
occur simultaneously with malaria, typhoid fever or both. This study was
conducted to determine the concurrent infections of malaria and/or typhoid fever
with Chikungunya virus, among febrile children aged 1 – 12 years seeking
treatment in Alupe Sub County Hospital, Busia,Kenya. Blood smears were prepared
for detection of malarial parasites and serum sample for widal testto detect typhoid
fever. Enzyme-linked Immunosorbent Assay and Plaque Reduction Neutralisation
Test were performed to detect the Chikungunya virus antibodies. The median
age/interquartile range age for the febrile children was 4.5 years and 55.5% were
female. Concurrent infections of Chikungunya virus with malaria or typhoid were
9.6% and 7%, respectively, using the Enzyme-linked Immunosorbent Assay
technique and 10.5% and 9.9%, respectively, using Plaque Reduction
Neutralisation Test. This supports the recommendation that Chikungunya virus
should be tested for using both serological and molecular diagnostics in cases of
patients presenting with fever.
Keywords: Malaria, Chikungunya virus and Typhoid fever
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Mwongula, A. W., Siamba, D. N., Mwamburi, L. A., & Mailu, M. (2023). Concurrent Infections Among Pyretic Children Seeking Treatment at Alupe
Sub County Hospital, Busia County, Kenya. European Journal of Applied Sciences, Vol - 11(5). 38-45.
URL: http://dx.doi.org/10.14738/aivp.115.15370
INTRODUCTION
Fever is a non – specific physiological mechanism of the immune system’s attempt to gain
advantage over infectious agents, such as viruses and bacteria. A fever makes the body less
favorable as a host for replicating viruses and bacteria, which are temperature sensitive.
Children typically get higher and fast – occurring fevers, reflecting the effects of the pyrogens
upon an inexperienced immune system (1). Besides malaria and typhoid fever as the diseases
which are associated with fever, Chikungunya virus according to sero-survey studies is an
endemic disease in Africa that cause fever.
Previous studies reveal that in malaria holoendemic regions; such as Western Region in Kenya
where Alupe is located, mosquitoes of Aedes species are found in large populations (2) The
species of mosquitoes that transmit malaria in the Western province are competent vectors for
Chikungunya virus transmission indicating a possibility of the virus circulating in Western
Province.
Chikungunya infections go undiagnosed or delayed diagnosis at the health centre level due to
lack of diagnostic tests and the children are usually treated with antimalarial drugs or
antibiotics empirically. Clinical and epidemiological similarities with dengue fever, malaria and
typhoid fever make Chikungunya diagnosis challenging (3), which may lead physicians to
misdiagnose Chikungunya as dengue fever, malaria or typhoid (3); therefore, the incidence of
Chikungunya occurrence may actually be higher than currently reported (3-5).
In addition, local transmission of Chikungunya into previously nonendemic areas by travelers
with viremia has been reported in tropical or subtropical areas of Africa (6). Chikungunya has
been responsible for significant human morbidity for several hundred years; yet in spite of its
prevalence; there are few studies which have looked into the epidemiology, mechanisms of
virulence and pathogenesis of Chikungunya especially in regions endemic for the potential
vectors (7).
Early recognition of local transmission followed by prompt, aggressive vector control and other
public health measures might prevent long-term establishment of the virus in new areas (5). So
far, no studies have been conducted to establish the prevalence and incidences of Chikungunya
in high-risk areas of Western Kenya which borders Uganda and served by Busia and Malaba
border crossing points.
METHODS
Study Site
The study was conducted in Alupe, Busia County, Kenya. This research was conducted in two
health facilities; Alupe Sub County Hospital and the Centre for Infectious and Parasitic Disease
Control Research (CIPDCR), Kenya Medical Research Institute (KEMRI).
Busia County has two border crossing points into Uganda – Busia and Malaba towns. The main
economic activity is trade with neighbouring Uganda which is extended to Rwanda and
Democratic Republic of Congo, with Busia and Malaba towns being the cross-border centers.
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European Journal of Applied Sciences (EJAS) Vol. 11, Issue 5, October-2023
According to the National census conducted in Kenya in 2009, Busia County had a total
population of 488,075 out of which 232, 075 were male and 256,000 female (m:f=1:1), (8).
Ethics Statement
Approval was sought from Ethical Review Committee at KEMRI (SSC PROTOCOL No. 2109-3RD
REVISION) through the Centre of Infectious and Parasitic Control Centre Research, Alupe Busia,
Kenya.
Study Design
This study utilized a hospital-based cross-sectional design. Serum samples were collected from
febrile children aged 1 to 12 years with symptoms or clinical features suggestive of
Chikungunya infection; these symptoms include fever, headache, myalgia, joint pain with or
without swelling, and the presence or absence of a rash on the body.
Study Population
The study comprised of both rural and urban populations of febrile children who sought
treatment at Alupe Sub County Hospital from January to December 2010. Criteria for inclusion
in the study were: those Children whose parents or guardians gave consent to be included in
the study, patients aged one year and above (to avoid maternal antibodies), all children who
had clinical presentations suggestive of Chikungunya infection and both sexes of patients.
Criteria for exclusion were: children who had no fever, children whose parent or guardians
declined to consent or patients aged 13 years and above.
Sample Collection, Storage, Transportation and Processing
Upon signing of the consent forms by the client’s, detailed history was taken to obtain
information on socio-demographic and clinical manifestations by the study clinicians. The
clients underwent phlebotomy using standard precautions in the laboratory where 5 ml of
whole blood was obtained in yellow capped vacutainer tubes. The blood sample was
centrifuged for 5 minutes at 5000 r.p.m and stored kept in liquid nitrogen before being
transported to KEMRI, CIPDCR where it was stored at -80oC until used.
Laboratory Sample Analysis of Chikungunya Virus
The study adopted an Indirect Enzyme Linked Immunosorbent Assay protocol as described by
Mwongula. et al., (9). Presence of specific anti-Chikungunya antibodies was confirmed by PRNT
done at Centre for Microbiology Research, KEMRI in Nairobi. The IgG, IgM, and IgA were
measured for the determination of the sero-prevalence of Chikungunya in febrile children
visiting the Alupe Sub County Hospital.
The plates were then read on an ELISA plate reader (Thermoscientific Multiskan ex. Version,
Tokyo, Japan) at a wavelength of A492nm (Ascent software version 2.6-Deafulte.See, Shangai,
China). The Optical Densities (OD) of positive-to-negative (P/N) ratios of >1.0 was considered
positive, <0.5 was considered negative and ≥0.5 was considered borderlines. All sera that were
positive and borderlines for Chikungunya indirect ELISA were further confirmed using the
Plaque Reduction Neutralization Test (PRNT).
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Mwongula, A. W., Siamba, D. N., Mwamburi, L. A., & Mailu, M. (2023). Concurrent Infections Among Pyretic Children Seeking Treatment at Alupe
Sub County Hospital, Busia County, Kenya. European Journal of Applied Sciences, Vol - 11(5). 38-45.
URL: http://dx.doi.org/10.14738/aivp.115.15370
Plaque Reduction Virus Neutralization Tests (PRNT)
A plaque reduction neutralization test was used to confirm the presence of specific neutralizing
antibodies to the Chikungunya in patient sera already regarded as positive or border by ELISA.
The study adopted a PRNT protocol as described by Mwongula. et al., (9).
The PRNT was determined by linear regression (probit) analysis using either probit paper or a
computer software program, or determine highest dilution that results in <50% of input plaque
count.
Laboratory Sample Analysis of Endemic Diseases That Cause Fever
Giemsa Staining of Thick Blood Smear for Malaria Microscopy:
Thick film was dried completely and 10% Giemsa stain gently poured on the slide using a
pipette on the staining rack for 15 minutes. The stain was gently flushed off the slide with a
clean tap water and the slide placed on the drying rack. The slide was observed under oil
immersion for malarial trophozoite parasites.
Widal Test:
A clean glass marked with circles was labeled “H” and "O”. One drop of undiluted serum was
placed in the two circles with the help of a sterile pasture pipette.
One drop of H antigen was added to the first circle and one drop of O antigen was added in the
second circle. With separate applicator sticks, serum and the antigen were mixed together and
spread well to fill the whole of the individual circle. The slides were observed for agglutination.
Data Presentation and Analysis
All data was recorded in purpose designed questionnaires. Clinical and laboratory data was
maintained as excel databases. Data was processed by a microcomputer using Genstat 4th
edition. All qualitative data was summarized using frequency tables and charts. Difference in
the prevalence of Chikungunya, malaria and typhoid were analyzed using Pearson Chi-square
test. The results were assumed to be significant when P≤0.05.
RESULTS
Demographic Characteristics
Three hundred and eighty-four patients aged between 1 and 12 years were recruited for the
study. The characteristics of the study population were as described by Mwongula. et al., 2013.
(9)
Misdiagnosis and Concurrent Infections
Symptoms and signs of Chikungunya infections are quite similar to those of other febrile
illnesses such as malaria and typhoid fever. Malaria and typhoid investigations are routinely
carried out to establish the cause of pyrexia of unknown origin (P.U.O) and treatment follows
with complete neglect of for Chikungunya infection. Chikungunya fever cases can sometimes be
misdiagnosed or occur simultaneously with malaria or typhoid fever.
Out of the 270 children that tested negative for malaria, 9.3% (25) tested positive for
Chikungunya. Among 114 children that tested positive for malaria, 9.6% (11) were positive for
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European Journal of Applied Sciences (EJAS) Vol. 11, Issue 5, October-2023
Chikungunya as shown in Table 1. The P value is greater than 0.05 and therefore there is no
significant relationship between Chikungunya and malaria.
Table 1: Proportion of Children with Chikungunya using ELISA and Malaria amongst the
febrile children seeking treatment at Alupe Sub County Hospital from January to
December 2010
Malaria test results Chikungunya
Border Negative Positive
Negative 20 (7.4%) 225 (83.3%) 25 (9.3%)
Positive 8 (7%) 95 (83.3%) 11 (9.6%)
NNT: No Neutralisation and NT: Neutralisation.
χ
2 = 0.00 DF=1 P=1.000
Among the 70 children that tested negative for typhoid, 20% (14) tested positive for
Chikungunya (Table 2). Out of 314 children that tested positive for typhoid, 7% (22) tested
positive for Chikungunya virus. The P value is less than 0.05 and therefore there is a significant
relationship between typhoid and Chikungunya virus.
Table 2: Proportion of Children with Chikungunya using ELISA and Typhoid amongst
the febrile children seeking treatment at Alupe Sub County Hospital from January to
December 2010
Typhoid test results Chikungunya
Border Negative Positive
Negative 8 (11.4%) 48 (68.6%) 14 (20%)
Positive 20 (6.4%) 272 (86.6%) 22 (7%)
NNT: No Neutralisation and NT: Neutralisation.
χ
2 = 13.43 DF=1 P˂0.001
Out of 270 children who tested negative for malaria, 11.9% (32) tested negative for
Chikungunya. Among 114 children who tested positive for malaria, 10.5% (12) tested positive
for Chikungunya as shown in Table 3. The P value is greater than 0.05 and therefore there is no
significant relationship between Chikungunya virus and malaria.
Table 3: Proportion of Children with Chikungunya using PRNT and Malaria amongst the
febrile children seeking treatment at Alupe Sub County Hospital from January to
December 2010
Malaria test results Chikungunya
NNT NT
Negative 238 (88.1%) 32 (11.9%)
Positive 102 (89.5%) 12 (10.5%)
NNT: No Neutralisation and NT: Neutralisation.
χ
2 = 0.14 DF=1 P=0.709
Out of 70 children who tested negative for typhoid, 18.6% (13) tested negative for
Chikungunya. Among 314 children who tested positive for typhoid, 9.9% (31) tested positive
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Mwongula, A. W., Siamba, D. N., Mwamburi, L. A., & Mailu, M. (2023). Concurrent Infections Among Pyretic Children Seeking Treatment at Alupe
Sub County Hospital, Busia County, Kenya. European Journal of Applied Sciences, Vol - 11(5). 38-45.
URL: http://dx.doi.org/10.14738/aivp.115.15370
for Chikungunya as shown in Table 4. The P value is less than 0.05 and therefore there is a
significant relationship between typhoid and Chikungunya virus.
Table 4: Proportion of Children with Chikungunya using PRNT and Typhoid amongst
the febrile children seeking treatment at Alupe Sub County Hospital from January to
December 2010
Typhoid test results Chikungunya
NNT NT
Negative 57 (81.4%) 13 (18.6%)
Positive 283 (90.1%) 31 (9.9%)
NNT: No Neutralisation and NT: Neutralisation.
χ
2 = 4.27 DF=1 P=0.039
DISCUSSION
Misdiagnosis of Chikungunya Virus infection as Malaria and Typhoid or Both
In the present study, 9.3% (25 out of 270) of children who tested negative for malaria were
reported to have Chikungunya and 20% (14 out of 70) of children who tested negative for
typhoid fever had Chikungunya using the ELISA technique. In addition, 11.9% (32 out of 270)
of children who tested negative for malaria had Chikungunya and 18.6% (13 out of 70) of
children who tested negative for typhoid fever were Chikungunya positive using PRNT
technique.
In 2004, Kenya experienced two outbreaks of Chikungunya fever. The first outbreak in Lamu
resulted in an estimated 13,500 cases, which represents 75% of the population of the island
(10). The outbreak was initially thought to be due to malaria, but eventually, laboratory
screening determined that Chikungunya was the cause.
Symptoms and signs associated with Chikungunya infection such as fever, joint pains, and
myalgias are non-specific that makes clinical diagnosis difficult because they are
indistinguishable from other febrile diseases such as dengue, Rift Valley fever, malaria and
typhoid. However, pronounced persistent severe joint pains that affect wrists, elbows, fingers,
and knees in some patients should raise the suspicion of alphavirus infection, especially
Chikungunya disease or O’nyong nyong fever, which also occurred in epidemic form in East
Africa in the late 1990s (11-13).
Concurrent infection of Chikungunya Virus infection with Malaria or Typhoid or Both
In the current study, 9.6% (11 out of 114) of children who were positive for malaria had
Chikungunya and 7% (22 of 314) children who tested positive for typhoid fever had
Chikungunya using the ELISA technique. In addition, 10.5% (12 out of 114) of children that
tested positive for malaria had Chikungunya and 9.9% (31 out of 314) of children who tested
positive for typhoid fever had Chikungunya using PRNT technique.
There was no significant difference between Chikungunya infection and malaria. Chikungunya
infection was positively correlated to typhoid. These may be explained by the fact that a disease
could exist independently while in other conditions a disease may exist in association with
other diseases. Death caused by Chikungunya infections appears to be rare. However, increases
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European Journal of Applied Sciences (EJAS) Vol. 11, Issue 5, October-2023
in crude death rates have been reported during the 2004–2008 epidemics (14 - 16).
Chikungunya infections, individuals with underlying medical conditions and individuals with
co-infections appear to be more likely to suffer complications and to have a higher risk of death
(17 -19).
In Kenya, febrile illnesses have been restricted to malaria and typhoid investigations and
treatment normally done with complete neglect of Chikungunya due to lack of cost-effective
diagnostics at the point of care.
Chikungunya infection was positively correlated with typhoid fever (P<0.05) and but
negatively correlated with malaria (P>0.05).
Chikungunya infections were misdiagnosed with the common febrile illnesses; 25/36 cases as
malaria and 14/36 cases as typhoid and 32/44 cases as malaria and 13/44 cases as typhoid
using PRNT and ELISA, respectively. Concurrent infection of Chikungunya with malaria or
typhoid was 9.6% and 7% using the ELISA technique and 10.5% and 9.9% using PRNT
technique, respectively. A significant number of patients had co-infection with typhoid (ELISA;
P=0.001 and PRNT; P=0.039).
Considerable number of patients had co-infection with malaria and typhoid hence even if
patient is diagnosed with other more prevalent infections, Chikungunya should be tested for.
The differential diagnosis of Chikungunya includes infections of other alphaviruses that cause
the fever such as Sindibis virus, O’nyong nyong virus, Ross River. In addition, Chikungunya
should be differentiated from dengue, malaria, typhoid fever or other more prevalent infectious
diseases for proper treatment and management. Serology and molecular diagnosis should be
used simultaneously for better case detection.
ACKNOWLEDGEMENT
The authors wish to express their gratitude to Kenya Medical Research Institute (KEMRI) –
Busia, Kenya in collaboration with Nagasaki University, Japan for sponsoring this project. Also,
technical assistance of Mr. T. Mokaya, Mrs. O. Makwaga, Carolyne Kirwaye, Najma Salim, Janet
Awando, Lucy Okubi, Angella Mutuku and F. Adung’o is highly appreciated.
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