Polytechnic University of the Philippines

College of Science, Department of Biology

Sta. Mesa, Manila

Mary Grace Abracia

BS Biology 4-2

Detection of Cryptosporidium sp. and Giardia sp. from

Commercial Bivalve Shellfish (Perna viridis)

I. Introduction

Cryptosporidium spp. are contamination by cryptosporidial oocysts is

monoxenous protozoan parasites that
cause gastrointestinal diseases in humans
and animals. Shellfish harvesting areas can
become contaminated by the infectious
stage of the parasite and humans are
therefore at risk of infection either by
consumption of shellfish, or by taking part
in recreational activities in these areas.
Transmission of Cryptosporidium
spp. occurs mainly by ingestion of oocysts
either by fecal-oral contact or through
contaminated food or drinking water.
Localized epidemics of food-borne
cryptosporidiosis have been associated with
uncooked sausage, offal, raw milk, apple
cider, or foodstuffs, but waterborne
transmission seems to play a more
prominent role and is implicated in most
outbreaks of human cryptosporidiosis. The
presence of Cryptosporidium oocysts in
drinking water supplies has been well
documented since 1984, and waterborne
epidemics of cryptosporidiosis have been
reported frequently in the United States,
United Kingdom, and Japan, among other
countries. The potential for water
high in areas where dumping of raw sewage
is a common practice . In addition, the
presence of waterborne Cryptosporidium
spp. oocysts of animal origin needs to be
considered, since a single neonatal
ruminant can shed up to 1010 oocysts
during the course of infection . The
presence of oocysts in river waters may also
be a source of contamination of the marine
environment. Rivers polluted by
anthropogenic and livestock fecal
discharges could play a major role in
contamination by oocysts of shellfish in
estuaries and coastal environments.
The study was conducted to
determine the potential role of mussels
(Perna viridis) as reservoirs of
Cryptosporidium spp. oocysts and their
value as biological indicators of the
presence of the parasite in water.
II. Materials and Methods III. Results and Discussions

MATERIALS The results were negative due to the

• Mortar and pestle process of homogenizing the meat of the
• 10 test tubes mussle. It was really hard for the mussle to
• Test tube rock
be infected because of its morphological
• Stirring rod
characteristics, unless the water of the area
• Rubber stopper
of the mussle is contaminated with
REAGENTS parasite.
• Sucrose solution
• Methanol Table 1 illustrates the dimension of
• Basic fucshin the mussle L-W- H- Wt. The experiment
• 50% Alcohol show a negative result that gives the
• Decolorizer researcher to explain that the mussle in the
• Methylene blue area of the market of Sta. mesa is negative
to have a parasite like Cryptosporidium spp.
Green mussels (Perna viridis)
Specimen of Green mussels (Perna Table 1. Shows the Dimension of the mussle
viridis) was purchased from Altura Wet (Mytilus sp.) and their average
Market. The dimension of bivalves shellfish measurement.
was get in terms of their length, height,
width and weight. The shell was disrupted
and meat was extracted and homogenized. Species L W H Wt
Afterwards in 1 ml of meat, 10 ml of sucrose 1 4.96 2.38 1.31 6.69
solution was added and centrifuged it at 2 4.9 2.2 1.68 5.26
2500 rpm within 5 mins. Or let it stand for 3 5.9 2.62 1.9 10.15
20-30 mins. Dropped the small amount of 4 5.06 2.4 1.7 9.04
sample at center of the slides and it was 5 4.96 2.3 1.6 8.04
ready for Kinyoun Acid Fast staining. 6 5.1 2.38 1.7 7.91
7 5 2.2 1.7 4.7
Kinyoun Acid Fast Staining 8 5 2.4 1.69 5.75
The slides with samples were dried, 9 5.1 2.3 1.8 6.15
and then submerged it with Methanol 10 5.36 2.38 1.7 6.69
within 1 min. and rinsed with top water. Average 5.134 2.356 1.678 7.038
Afterwards the slides were immersed with *L-length: W- width, H-height; WT- Weight
Basic Fucshin for 5 mins. 50% alcohol was
used to wash it and rinsed with top water.
Decolorizer was added for 2 mins until the IV. Conclusion
color runs and rinsed with top water. The The mussle in the area of the market of
slides were submerged with Methylene Sta.Mesa was not infected of the parasite
Blue for 1 min and lastly washed it by top Cryptosporidium spp. and Giardia sp.
water and ready for microscopic
examination.
References

Gómez-Couso, H., F. Freire-Santos, C. F. L.

Amar, K. A. Grant, K. Williamson, M. E.
Ares-Mazás and J. McLauchlin 2003.
Detection of Cryptosporidium and
Giardia in molluscan shellfish by
multiplexed nested-PCR

Norling, P., and N. Kautsky 2007. Patches of

the mussel Mytilus sp. are islands of
high biodiversity in subtidal sediment
habitats in the Baltic Sea

Shields, J.M, E.R. Gleim, and M.J. Beach

2005. Prevalence of Cryptosporidium
spp. and Giardia intestinalis in
Swimming Pools, Atlanta, Georgia