**6. Concluding remarks**

354 Environmental Monitoring

the parasites. However, the methods used in this study are in accordance with those recommended for concentration and detection of microorganisms by the *Standard Methods for the Examination of Water and Wastewater* (Clesceri, 1998). They are easily applied, do not pose a great risk to the technician, and are low cost techniques, which can be employed by

The performance of some rapid gravity filters was evaluated in England, using turbidity measurement and particle counts in filtered water as parameters for monitoring and controlling *Cryptosporidium* sp. oocysts as an indicator microorganism (Geldreich, 1996), a

The *in vitro* amplification of DNA fragments of *Cryptosporidium* sp. obtained sensibility and specificity. Nevertheless, the amplification was only possible using Nested-PCR primers (AWA995f/AWA1206R and LAX469F/LAX869R). The primer LAX469F/LAX869R amplifies the regions of *C. parvum*/*C. hominis*, but *C. parvum* diagnosis was confirmed by the difference in diameter, since its oocyst is approximately 5 µm in diameter, while *C. hominis*

Nested-PCR presents the advantage of concentrating a smaller quantity of PCR inhibitors (Kirkpatrick & Green, 1985). In environmental samples, there are several Taq DNA polymerase inhibitors, such as fecal hemoglobin and phenolic compounds, and it might

It was possible to obtain satisfactory amplification with the two methods of DNA extraction applied. Furthermore, they are quick and low-cost, although close attention should be paid

As adverse environmental factors have been proven to alter the morphology of cysts and oocysts (Hsu BM, 2001), making their detection more difficult, this may justify the low positivity found in the present study using parasitological methods. Other factors might have had influence as well, such as the concentration of *Cryptosporidium* sp. oocysts, based almost exclusively on particle size (Hsu, 2001). Also, the level of protozoa may vary according to the season, and an increase in their resistant forms in rainy periods, winter and

Temperature has also been considered a factor that influences protozoa and autochthonous microorganism survival in rivers (Howe, 2002). In this study, we observed just small variations of water temperature in the rivers and lakes sampled during the period of study, although within the limits that allow the survival and viability of protozoa. Using univariate logistic regression (p = 0.066), we demonstrated that temperature was not a statistically significant variable, whereas humidity (p = 0.958) was. In the region of sample collection there are two well-defined seasons, the dry (from April to September) and the rainy (from October to March) seasons, the latter characterized by torrential rain and runoff, which

Due to the low number of protozoa found in this work, i.e. two *Cryptosporidium* sp. and four *Giardia* sp., we could not infer if the protozoan levels vary by season, but only observe the

 Standardization and application of parasitological and molecular techniques in the analysis and seasonal monitoring of opportunistic protozoa were successfully carried

beginning of spring has already been reported (Atherholt 1998, Ong et al. 2002)

technicians trained to monitor water for human consumption.

have been the case of the samples processed in the present research.

to the phenol/chloroform method since it is toxic and corrosive.

certainly makes the detection of parasites more difficult.

out for environmental samples;

**5. Conclusion** 

qualitative inference of their presence in the bodies of water monitored.

The rivers and lakes of Goiânia are contaminated with opportunistic protozoa;

method similar to the one used in our study.

oocyst is approximately 4 µm in diameter.

*Cryptosporidium* is considered a coccidia resistant (Carey et al. 2004), because oocysts have characteristics that favor its rapid spread in the environment, such as the ability to withstand the action of commonly used disinfectants (formaldehyde, phenol, ethanol, lysol), able to cross some water filtration systems due to its small size, the ability to float, remain in the environment by a few weeks or months and tolerance in certain temperatures and salinity (Fayer et al. 2004). Given the scope of the aquatic environment coupled with the wide distribution of different species in Brazilian waters, make the control measures of Cryptosporidium limited.

Therefore, to minimize the risks inherent in the spread of cryptosporidiosis in the populations of free-living mammals, it is of fundamental importance to environmental control, through the adoption of agricultural practices to prevent pollution of rivers by the faeces of animals (Graczyk et al. 2000), as well as encouraging the adequacy of sanitation facilities, protection of water sources, education and guidance on waste discharges from vessels during nautical activities. Regarding the control measures of captive aquatic mammals, so as to minimize or eliminate the risks inherent in the spread of coccidian, several studies should be adopted.

Finally, it must be remembered that currently monitoring systems treated water are based on the frequency of fecal coliforms and Escherichia coli as indicators of pollution, and that this methodology is insufficient to predict the presence of other pathogens such as parasites. Thus, it is imperative the use of alternative methods for the diagnosis, investigation and monitoring of large amounts of water of these pathogens. For in this way can be proposed reorganization measures that contribute to reducing the incidence of opportunistic diseases emerging in water of human use, especially for children, elderly, immunocompromised and immunosuppressed patients.
