Use of Tetrahymena thermophila to study the role of protozoa in inactivation of viruses in water.

The ability of a ciliate to inactivate bacteriophage was studied because these viruses are known to influence the size and diversity of bacterial populations, which affect nutrient cycling in natural waters and effluent quality in sewage treatment, and because ciliates are ubiquitous in aquatic environments, including sewage treatment plants. Tetrahymena thermophila was used as a representative ciliate; T4 was used as a model bacteriophage. The T4 titer was monitored on Escherichia coli B in a double-agar overlay assay. T4 and the ciliate were incubated together under different conditions and for various times, after which the mixture was centrifuged through a step gradient, producing a top layer free of ciliates. The T4 titer in this layer decreased as coincubation time increased, but no decrease was seen if phage were incubated with formalin-fixed Tetrahymena. The T4 titer associated with the pellet of living ciliates was very low, suggesting that removal of the phage by Tetrahymena inactivated T4. When Tetrahymena cells were incubated with SYBR gold-labeled phage, fluorescence was localized in structures that had the shape and position of food vacuoles. Incubation of the phage and ciliate with cytochalasin B or at 4 degrees C impaired T4 inactivation. These results suggest the active removal of T4 bacteriophage from fluid by macropinocytosis, followed by digestion in food vacuoles. Such ciliate virophagy may be a mechanism occurring in natural waters and sewage treatment, and the methods described here could be used to study the factors influencing inactivation and possibly water quality.

Transport behaviour and natural attenuation of organic contaminants at spill sites.

Organic contaminants pose a significant threat to groundwater resources. These contaminants are often released as non-aqueous phase liquids (NAPLs) during spills of, for example, gasoline, crude oil, creosote, coal tar or chlorinated solvents. Once released, the liquids seep downward and dissolve into the groundwater. In many cases, the impacted groundwater contains a mixture of contaminants, either due to the complexity of the NAPL (e.g., gasoline) or due to co-disposal/co-spillage (e.g., landfill leachates). Many organic contaminants are hazardous to human health and the environment and therefore threaten our potable water resources and natural ecosystems. Active remediation of contaminated groundwater is often very expensive so that cost-effective alternatives have to be found. If natural attenuation is intended to be used as a means of achieving specific remedial objectives at a contaminated site, it will require a sound understanding of the ongoing processes as well as careful control and monitoring ("monitored natural attenuation" (MNA)). Therefore, a major goal of remediation research today is to develop methods to predict the mass fate of multiple organic compounds in heterogeneous aquifers under natural conditions.

Laboratory evidence of MTBE biodegradation in Borden aquifer material.

Mainly due to intrinsic biodegradation, monitored natural attenuation can be an effective and inexpensive remediation strategy at petroleum release sites. However, gasoline additives such as methyl tert-butyl ether (MTBE) can jeopardize this strategy because these compounds often degrade, if at all, at a slower rate than the collectively benzene, toluene, ethylbenzene and the xylene (BTEX) compounds. Investigation of whether a compound degrades under certain conditions, and at what rate, is therefore important to the assessment of the intrinsic remediation potential of aquifers. A natural gradient experiment with dissolved MTBE-containing gasoline in the shallow, aerobic sand aquifer at Canadian Forces Base (CFB) Borden (Ontario, Canada) from 1988 to 1996 suggested that biodegradation was the main cause of attenuation for MTBE within the aquifer. This laboratory study demonstrates biologically catalyzed MTBE degradation in Borden aquifer-like environments, and so supports the idea that attenuation due to biodegradation may have occurred in the natural gradient experiment. In an experiment with batch microcosms of aquifer material, three of the microcosms ultimately degraded MTBE to below detection, although this required more than 189 days (or >300 days in one case). Failure to detect the daughter product tert-butyl alcohol (TBA) in the field and the batch experiments could be because TBA was more readily degradable than MTBE under Borden conditions.