healthmedicinet

Study design and area

A cross sectional study was conducted in school age children in Tach Armachiho Ethiopia
from May to June 2013. The Tach Armachiho is one of the woredas in the Amhara National
Regional states of Ethiopia. It is found 260 km away from Bahir Dar, the capital city
of the region. The prevalence of intestinal parasites is high in the region especially
in remote area like Tach Armachiho due to low sanitation and hygiene. As a result,
we were interested to conduct this research in Tach Armachiho area.

Sample collection procedure

A total of 422 primary school age children were selected as study participants. Of
these, 135 (31.9 %) were males and 287 (68 %) were females having age ranges from
6–11 years. From registration list, simple random sampling method was employed to
select students from each section using a table of random numbers and when the selected
student was absent, the student before or after the indicated one was sampled for
replacement.

Students who were under taking anti parasitic drugs during the data collection date
or refused in participation were excluded from the study.

Single stool sample was collected from each participant by sterilized stool cup. Three
stool slides smears were processed for Wet mount, FEC and Kato Katz techniques from
each sample and the slides were examined microscopically. The performance of Wet mount,
Kato-Katz and FEC techniques were evaluated in terms of “Gold” standard. The Wet mount
preparation and examination was performed as soon as the stool arrives in the laboratory.
The FEC and Kato Katz techniques were processed depending on the test protocol and
smeared on different slides. Finally, labeling of the slides was done with the students
ID number.

Diagnostic techniques

Wet Mount: In the Wet mount, fresh stool samples (approximately 2 mg of stool) were
put on a slide with wooden applicator, emulsified with a drop of physiological saline
(0.85 %) for diarrheic and semi solid or Iodine for formed stools, covered with cover
slide and examined under microscope using first 10 × objectives and then 40 × objectives.

Formol ether concentration: This test was performed by mixing 0.5 g of faeces in 10 ml
of normal saline in a glass container and mix thoroughly. Two layers of gauze were
placed in a funnel and strain the contents into a 15 ml centrifuge tube. 2.5 ml of
10 % formaldehyde and 1 ml of ether was added. The solution was mixed well and centrifuge
at 1000 revolution for 3 min. The supernatant was removed and the slides were prepared
from the sediment. Two slides were prepared (one for saline and the other for iodine),
covered with cover slide and finally examined with microscope.

Kato Katz technique: The Kato Katz quantitative cellophane thick smear method (Mahidol
University, Thailand following the manufacturer’s instruction) was used. It was performed
by transferring the sieved stool to the templates which delivers 41.7 mg of stool.

The stool was covered with cellophane which was previously immersed with malachite
green. Identification of the ova or cyst of the parasites and quantification of the
ova of S. mansoni and geo-helminths were done. Eggs counted per slide were multiplied by 24 to convert
into number of eggs per gram (epg) of stool. The parasite load or intensity was defined
as light (1–100 epg), moderate (101–400 epg) and heavy based (400 epg) of stool for
S. mansoni and geo-helminthes according to World Health Organization (WHO) guideline 13], 14].

In all cases, the results of the Wet mount slides were determined earlier to FEC and
Kato Katz techniques with strict blinding. The independent readings of the Wet mount,
FEC and Kato Katz slides by the experienced laboratory personnel were checked by the
principal investigator. To avoid observer bias, two experienced laboratory personnel
performed the microscopic examination of Wet mount, FEC and Kato Katz slide smears
blindly and independently. The results of their observation were recorded for later
comparison on separate sheets. A quality control was done by repeating all discordant
results. The discordant smears were rechecked by the principal investigator using
100 × objectives.

Data entry and analysis

Data were entered and analyzed using SPSS version 20. Since, there is no “Gold” standard
method to detect intestinal parasites; the operational characteristics [sensitivity
and negative predictive value (NPV)] of diagnostic tests were estimated using the
combined results from the three methods as diagnostic “Gold” standard 15]. The combination of Wet mount, FEC and Kato-Katz tests was used as a “Gold” standard
diagnostic test. Sensitivity, specificity, positive predictive value, NPVs and Kappa
value of Wet mount, FEC and Kato-Katz techniques were calculated against the Gold
standard.

Ethical considerations

Ethical clearance was obtained from College of Medicine and Health Science ethical
review committee, Bahir Dar University and from Tacharmachiho Woreda Education Office.
Written informed consent was obtained from every study participant including the parents
and the guardians of the children. Intestinal parasite positive cases were treated
with antihelminthics and anti protozoals depending on the type of species identified.