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British Journal of Healthcare and Medical Research - Vol. 9, No. 2
Publication Date: April, 25, 2022
DOI:10.14738/jbemi.92.12151. Oyewole, I. O., Mustapha, A. K., Adeogun, A., & Awolola, T. S. (2022). Malaria infection and Entomological Survey of Anopheline
Species in Some Communities in Ila Local Government Area of the State of Osun, Nigeria. British Journal of Healthcare and Medical
Research, 9(2). 117-128.
Services for Science and Education – United Kingdom
Malaria infection and Entomological Survey of Anopheline
Species in Some Communities in Ila Local Government Area of the
State of Osun, Nigeria
Isaac Olayinka Oyewole
Biology Department, Osun State College of Education
Ila Orangun, Osun State, Nigeria
Basic Sciences Department,Babcock University
Ilisan Remo, Ogun State, Nigeria
Abdur-Rahman Kolawole Mustapha
Biology Department, Osun State College of Education,
Ila Orangun, Osun State, Nigeria
Department of Zoology, Osun State University
Osogbo, Osun State, Nigeria
Adedapo Adeogun
Department of Biology, Lead City University
Ibadan, Oyo State, Nigeria
Nigerian Institute of Medical ResearchYaba, Lagos State, Nigeria
Taiwo Samson Awolola
Nigerian Institute of Medical Research, Yaba, Lagos State, Nigeria
ABSTRACT
A cross-sectional survey of the population of mosquitoes and incidence of malaria
was carried out in Iyana-Ofa Community and some selected villages around the
College campus (College Community) in Ila Local Government Area, State of Osun,
Nigeria for a period of one year (dry and rainy seasons). Mosquitoes were collected
using the aspirator tube and pyrethrum spray catch (PSC). Morphological and PCR
methods were used for identification. ELISA was employed to determine circum- sporozoite antigens of Plasmodium falciparum. Human biting rate (HBR) was
determined to calculate the Entomological inoculation rate (EIR). Blood film
examination using Giemsa staining techniques was used to detect malaria parasites
in the blood of 427 participants. A Thick film was used to determine the density of
the malaria parasite. Out of 946 mosquitoes collected, 605 (64%) were Anopheles
gambiae. No significant difference in mosquito population density (p>0.05) from
the study communities. EIR from the study areas was moderately high and no
significant difference was observed. P. falciparum predominates among
participants positive for malaria parasites. Parasite density ranged from 100 to
4,000 parasite/μl blood, the mean parasite density generally declined with age
while that 51-60 yr recorded the least. There was no significant difference in the
malaria infection rates in the two communities (P>0.05), but there was a
correlation between the mosquito population density and the prevalence of
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British Journal of Healthcare and Medical Research (BJHMR) Vol 9, Issue 2, April - 2022
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malaria. This study provides baseline information necessary to formulate effective
control interventions in the area.
Keywords: Malaria transmission, Parasites, Anopheles mosquito, Rural community
INTRODUCTION
Malaria is one of the insect-transmitted diseases and remains a major source of illness and
death, particularly in the tropics due to the high frequency of transmission of Plasmodium
species by a large number of Anopheles mosquitoes. The vector transmits malaria to more than
700 million people annually in the affected regions; here 1.2 billion are reported to be at risk of
the infection [1,2] . Sub-Saharan Africa alone accounts for 92% of the global malaria burden
where an estimated 200 million cases of malaria was reported in 2017 [3].
Plasmodium species implicated for human malaria infection in Nigeria include P. falciparum, P.
ovale, and P. malariae. However, P. falciparum had been identified as the parasite responsible
for most malaria-related deaths worldwide and is the predominant Plasmodium species in Sub
Saharan Africa. Findings showed that P. falciparum is responsible for more than 90% of all
malaria cases transmitted through the bites of competent female mosquitoes of the genus
Anopheles [3].
Anopheles species are widely distributed across the different ecological zones in Nigeria where
suitable sub-Saharan climatic conditions exist [4]. The implicated species are found to reflect
varying mosquito–human interactions which have impacted their capacity to transmit the
parasite [5]. Despite the major attempts over the past century to control malaria, vector
resistance to insecticides and malaria parasite resistance to multiple drugs have stood in the
way of malaria control [6,7]. According to the recommendation of the World Health
Organization, vector control is one of the important components of the global strategy for the
prevention of malaria [8]. It has also been documented that vector identification forms an
essential component of the strategy [9].
Previous reports had shown that prevalence in malaria varies spatially and temporally with
topography [10], climatological factors [11], altitude [12], availability of breeding sites [10, 13]
and level of urbanization [14,15]. Findings also indicate that variation in micro geographical
factors with disease prevalence could be an important determinant. For instance, in localities
constituting malaria-endemic regions, some localities have shown a high malaria burden while
others were more or less disease-free [16, 17]. In recent times most of the efforts directed
towards the control of malaria include the interruption in the disease transmission by reducing
man-mosquito contact [18, 19]. To achieve reasonable success through these efforts, there is
the need to adequately identify the implicated species and their parasite transmission dynamics
in each locality.
Due to a paucity of information on the malaria transmission dynamics of the identified
mosquito species in these locations, a study was therefore carried out to investigate its
population dynamics as related to malaria incidence to form baseline information for a vector
control strategy in these localities.
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Oyewole, I. O., Mustapha, A. K., Adeogun, A., & Awolola, T. S. (2022). Malaria infection and Entomological Survey of Anopheline Species in Some
Communities in Ila Local Government Area of the State of Osun, Nigeria. British Journal of Healthcare and Medical Research, 9(2). 117-128.
URL: http://dx.doi.org/10.14738/jbemi.92.12151
MATERIALS AND METHODS
Study Area
The study was carried out in Ila Orangun; a cosmopolitan town (8° 01' 1.70" N 4° 54' 15.16" E)
located 350 km from Osogbo, capital of the State of Osun, south-western Nigeria. The climate of
the area is characteristic of the forest zone with two distinct seasons. The rainy season is from
April to October while the dry season is from November to March. The mean annual rainfall is
2,500 and 3,000 mm while mean relative humidity ranges from 60-90%. The highest and lowest
monthly mean relative humidity was observed during rainy and dry seasons, respectively. The
mean temperature is 24°C during the wet and 30°C during the dry season. Adjoining villages
along Iyana-Ofa (Iyana-Ofa community) and the college (College community) constitute the
study area. Iyana-Ofa community is located along the express road leading to Ofa, a metropolis
in Kwara State, Nigeria. The residents are a mixture of Yorubas and nomadic Fulanis. The
housing structures consist mainly of modern homes with brick walls and corrugated iron roofs
while few are traditional houses built with mud walls and either thatched or corrugated iron
roofs. The inhabitants are mainly artisans and traders while few are cattle rearers and farmers.
The community consists of a modern structured hospital and a community health centre. The
adjoining villages constituting the College community include Ajebandele, Baba Eko, and
Jagilegbo which is about 3km from the main town. The community is surrounded by a tropical
deciduous forest with a nearby stream or its tributary in the forest. The houses in the
surrounding villages are mainly of mud and thatched or corrugated iron roofs. Houses are often
dark and damp with or without windows. People in this area often stay outdoors to relax or
take fresh air in the evening after the day's job, most often without protective measures while
children are found playing around naked at the mercy of mosquito bites.
STUDY DESIGN AND METHODS
Sample collection
A cross-sectional survey was carried out from randomly selected 65 houses to sample for
Anopheles mosquito during the dry and raining season for a period of one year while 427
volunteers (age range between 0 and 60 yrs) were sampled for malaria parasites.
Subject selection
Inclusion criteria
Only participants who were permanent residents (resident in the area >6 months) and
currently having symptoms of malaria were included in the study.
Exclusion criteria
Subjects who were on transitory assignment in the area were not included in the study.
Sampling technique
A multistage random sampling technique was used in choosing the required samples from
subjects (427) whose ages ranged from 0 to 60 yrs and were subdivided into 7 age groups of
ten years interval each. House enumeration was carried out while a systematic random
sampling technique using a sample interval of two and three was used to choose 65 houses in
the selected communities.
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Entomological survey
Mosquito collection was carried out in Iyana-Ofa community and College community
comprising of Ajebandele, Baba Eko, and Jagilegbo around the Osun State College of Education,
Ila- Orangun.
Mosquito collections
Mosquitoes resting indoors were collected in the study communities twice a month using hand
collection and indoor residual spray (IRS) methods. Collections cover the early part of the
morning between 0600 and 0900 h in selected houses, that is, for 15 minutes in each of the
study communities as per standard techniques. The number of mosquitoes caught per hour
and the hour biting rate (HBR) were recorded. Samples were stored individually over
desiccated silica in Eppendorf tubes before identification and ELISA test.
Species identification
Morphological identification
Identification of mosquitoes was done using morphological keys of Gillies and De Meillon [20]
and Gillies and Coetzee [21].
Molecular identification
A molecular assay was carried out using the species-specific PCR [22] with minor modifications
as detailed in Van Rensburg et al. [23] for the confirmatory identification of the members within
An. gambiae complex.
ELISA tests
The circum-sporozoite proteins of Plasmodium species present on the head and thorax of the
Anopheles mosquitoes were tested following the method of Wirtz et al. [24]. Sporozoite rates
were determined photometrically as described by Beier et al. [25].
Entomology inoculation rate (EIR)
The entomological inoculation rate (EIR) was calculated for each species as the product of the
sporozoite and human biting rates.
Parasitological survey
A Parasitological survey was conducted in the Iyana-Ofa community and College community
comprising of Ajebandele, Baba Eko, and Jagilegbo around the Osun State College of Education,
Ila- Orangun. During the monthly visit, blood samples were collected from volunteers using a
venepuncture technique (Onyido et al., 2011). Two millilitres of venous blood collected from
each of the 427 volunteers were stored in an ethylene-diamine-tetra-acetic acid (EDTA)
(anticoagulant) specimen bottle for further analysis.
Laboratory Analysis
The blood sample was analyzed within 1 to 2 h of collection. Thick and thin films were prepared,
stained, and examined for malaria parasite under the microscope using the oil immersion
objective followed WHO (2000) protocol. A finger prick blood sample was also used to assess
Plasmodium spp. infection in the peripheral blood using Rapid Diagnostic Test (RDT). Parasite
density per micro litre of blood was estimated from the thick film. Parasites were counted
against 200 white blood cells and converted into counts per micro litre assuming an average
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Oyewole, I. O., Mustapha, A. K., Adeogun, A., & Awolola, T. S. (2022). Malaria infection and Entomological Survey of Anopheline Species in Some
Communities in Ila Local Government Area of the State of Osun, Nigeria. British Journal of Healthcare and Medical Research, 9(2). 117-128.
URL: http://dx.doi.org/10.14738/jbemi.92.12151
count of 8000/μl blood. Blood films were examined microscopically using 100X (Oil
immersion) objectives [26].
Data analysis
Parasite density per micro litre of blood was estimated from the thick film, taking the number
of leucocytes per micro litre of blood as 8,000 (8000/μl blood) and was expressed as:
Parasite density = Parasite count x 8,000
No of WBC counted
Percentage parasitemia was expressed as:
% parasitemia = No of infected red blood cells x 100
Total no of red blood cells
Malaria parasite density was defined as log10 of the number of asexual malaria parasites/μl
blood in the thick blood films of malaria-positive volunteers to approximate normality.
Statistical analysis
Data collected were analyzed using STATA statistical package (STATA Corp LP, USA, Version
9), while ANOVA was used as test statistics (P-value 0.05).
ETHICAL CONSIDERATIONS
All aspects of the study were approved by the Osun State Ministry of Health Ethical Committee
and written Informed consent was obtained from the respondents. They were assured of
voluntary participation, the confidentiality of their responses, and the opportunity to withdraw
at any time without prejudice in line with the Helsinki Declaration [27].
RESULTS
A cross-sectional survey of the mosquito population in the study area produced 946 catches
from both communities and these comprised of 605 anophelines and 341 culicines. The highest
collection was recorded in September which coincided with the peak of rain in the area. An.
gambiae predominate the anopheline species in the study area comprising 64% of the total
catch. However, there was a significant difference (p>0.05) in the abundance of mosquitoes in
the sampled communities. The relative abundance of the mosquito species in each of the
locations is shown in Table 1. The collections showed seasonal significant difference between
the two communities (p>0.05). Molecular identification of An. gambiae (Fig. 1)
Entomological profiles of malaria transmission as presented in Table 2 indicate no significant
difference in the Human biting rate, Sporozoite, and Entomological inoculation rates in the
study areas (P=1.004, P>0.05). However, the peak of these rates was also recorded in
September corresponding with the period of an abundance of Anopheles mosquito.
Parasitological profiles of malaria transmission as analyzed in this study showed the trend of
the prevalence of malaria parasite among volunteers whose ages ranged between 0 and 60 yrs.
Participants within the age bracket of 21-30 years had the highest prevalence of malaria
85(90.4%) while the age group 51-60 years has the least 19(50%) and the results were
significant (P=0.016, P<0.05) (Table 3). Of the 427 participants examined for malaria parasites
in the study area, 319 were infected with a prevalence rate of 74.7%. A total of 197 participants
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were examined from the College community only 112(56.8%) carried infected parasites. At the
Iyana-Ofa community, 207(90%) were infected out of 230 examined for malaria parasite.
Generally, the participants at Iyana-Ofa possessed higher parasite density than those from the
College community (Table 4). There was no statistical difference in the malaria infection rates
between the two communities (p≥0.05), although more participants were available for
examination at Iyana-Ofa.
Detection of the malaria parasites in the blood film showed that P. falciparum was predominant
in the study area.
Meteorological data shows that January recorded the lowest rainfall (0.0mm) and relative
humidity (48%). The highest range of rainfall, temperature, and relative humidity was recorded
in September (288.0mm), February (36.4oC), and September (91.0%) respectively. The results
showed that the trend of mosquito abundance and infection rate corresponds with the increase
in rainfall (Fig. 2 and 3).
Table 2. Entomological Profiles of Malaria Transmission in the Study areas
Entomological Profiles College community Iyana-Ofa Community
Human biting rate 24.57± 5.60 a 26.33±4.23a
(mosquito/man/night)
Sporozoite rate (%) 2.05±1.92b 2.46±2.41b
Entomological inoculation rate 0.81±0.21c 1.07±0.44c
(infective bites/man/night)
Value followed by same superscript alphabet in a row are not significantly different at p=0.05
level of significant
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Fig.1: PCR product showing the amplified fragment using the species –specific assay for the
identification of member of the An. gambiae s.s.
Figure 2. Relationship between rainfall and number of persons infected with malaria
Fig.1:
Monthl
y mean
for Pa
Figure 3. Relationship between rainfall and mosquito abundance
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RAINFALL (MM)
NO OF INFECTED PERSON
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Monthly Mean rainfall (mm) Monthly Mean Parasitemia
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NO OF MOSQUITO COLLECTED
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Monthly Mean rainfall (mm) Monthly Collection of Anopheles gambiae