Seasonal retention and release of Cryptosporidium parvum oocysts by environmental biofilms in the laboratory

Abstract

Cryptosporidium is a genus of waterborne protozoan parasites that causes significant gastrointestinal disease in humans. These parasites can accumulate in environmental biofilms and be subsequently released to contaminate water supplies. Natural microbial assemblages were collected each season from an eastern Pennsylvania stream and used to grow biofilms in laboratory microcosms in which influx, efflux, and biofilm retention were determined from daily oocyst counts. For each seasonal biofilm, oocysts attached to the biofilm quickly during oocyst dosing. Upon termination of oocyst dosing, the percentage of oocysts retained within the biofilm decreased to a new steady state within 5 days. Seasonal differences in biofilm retention of oocysts were observed. The spring biofilm retained the greatest percentage of oocysts, followed (in decreasing order) by the winter, summer, and fall biofilms. There was no statistically significant correlation between the percentage of oocysts attached to the biofilm and (i) any measured stream water quality parameter (including temperature, pH, conductivity, and dissolved organic carbon concentration) or (ii) experimental temperature. Seasonal differences in oocyst retention persisted when biofilms were tested with stream water from a different season. These data suggest that seasonal variation in the microbial community and resulting biofilm architecture may be more important to oocyst transport in this stream site than water quality. The biofilm attachment and detachment dynamics of C. parvum oocysts have implications for public health, and the drinking water industry should recognize that the potential exists for pathogen-free water to become contaminated during the distribution process as a result of biofilm dynamics.

FIG. 1.

Seasonal changes in oocyst retention by biofilms using a standard laboratory water temperature (20 to 25°C). The cumulative percentage of oocysts associated with the biofilm over time for summer, fall, winter, and spring biofilm cultures was determined. Error bars show the percent standard error (n = 2) and are smaller than the symbols where not visible. The black line on the time axis indicates the period of oocyst dosing (0 to 72 h).

FIG. 2.

Testing the effect of experimental water temperature on oocyst retention by biofilm. The cumulative percentage of oocysts associated with the biofilm over time with winter culture biofilm grown at 5°C (ambient biofilm collection temperature) and at 20 to 25°C (normal laboratory experimental temperature) was determined. Error bars indicate the percent standard error (n = 2) and are smaller than the symbols where not visible. The black line on the time axis indicates the period of oocyst dosing (0 to 72 h).

FIG. 3.

Cumulative percentage of oocysts associated with the biofilm at the end of oocyst dosing (day 3) and the end of the experiment (day 8). The error bars indicate the percent standard error (n = 2).

FIG. 4.

Effect of changing flow media on oocyst retention by biofilm. The cumulative percentage of oocysts associated with the biofilm over time with fall biofilm grown with fall water, fall biofilm grown with spring water, spring biofilm grown with fall water, and spring biofilm grown with spring water was determined. The error bars show the percent standard error (n = 2) and are smaller than the symbols where not visible. The black line on the time axis indicates the period of oocyst dosing (0 to 72 h).

FIG. 5.

Correlation of several water quality measures and oocyst retention by biofilms. The percentages of oocysts attached to biofilms at day 3 and day 8 versus water temperature at the time of biofilm collection (A), pH (B), conductivity (C), and DOC concentration (D) were determined. Spearman correlation coefficients and P values for each parameter are indicated on each panel.