Biology Reference
In-Depth Information
3.4.3. Protozoan detection
The major protozoan pathogens found in water contamination are
Cryptosporidium
,
Giardia
, microsporidia,
Entamoeba histolytica
and
Cyclospora
.
Routine monitoring of water samples for protozoan parasites rely on the
detection of FIB, although direct detection of the protozoan can be moni-
tored using immunofluorescent antibodies and microscopy.
31
Since many outbreaks of disease have been attributed to
Cryptosporidium
and
Giardia
, especially in the UK and the US (most likely not indicative
of worse water treatment regimes here, but perhaps detailed public health
surveillance), there are standard, validated monitoring methods which are
widely adopted, especially at all UK water suppliers.
The standard recovery and detection protocol includes a concentration
step, usually through filtration, followed by a purification step, involving
density gradients, and finally an immunofluorescent staining step which
increase the chances that the (oo)cysts can be detected by microscopy. This
protocol, “information collection rule” (ICR) for protozoa, was developed
as a standardized method for the collection and quantification of
Giardia
cysts and
Cryptosporidium
oocysts.
20
It is still being used by some labs and
has been adopted as a standard method for the detection of other parasites,
as well as from effluents, surface and ground water sources. However, due to
low recovery rates associated with the ICR method, a newer method called
Method 1623 was developed. The main difference is the use of immu-
nomagnetic separation which allowed for more efficient separation of the
organisms from other debris resulting in a cleaner slide for microscopy.
Method USEPA 1623 describes the filtration, concentration and sub-
sequent detection of these pathogens, and illustrated in
Fig. 3.7
. Like with
fecal indicator organisms, filtration is used to capture the organisms. How-
ever, since infectious doses are so low and the organisms will not multiply
outside of a host, large water volumes are sampled to increase detection
likelihood. Attempts to develop culture methods have not yet met with
great success.
33
The first stage thus comprises filtration of 1000L, over
a period of 24 h. This cartridge filter is then eluted to release the (oo)-
cysts and a second filtration step is performed, this time with a mem-
brane filter. Next the (oo)cysts are subjected to centrifugation, followed
by specific separation using immunomagnetic beads. Finally, the (oo)cysts
are removed from the beads, placed on a microscope slide, and fluores-
cently stained before they are examined under the microscope by a highly
trained technician.
Search WWH ::
Custom Search