Effect of Sediment Particle Size and Temperature on Fecal Bacteria Mortality Rates and the Fecal Coliform/Fecal Streptococci Ratio

Extended survival of fecal bacteria in sediment can obscure the source and extent of fecal contamination in agricultural settings. The variability in fecal coliform/fecal streptococci (FC/FS) ratios with time and discrepancies between observable fecal sources and measured FC/ FS ratios in shallow surface water from agricultural watersheds may be explained by examining FC and FS mortality rates in response to ambient temperature and sediment particle size. We measured FC and FS mortality rates at three different temperatures and in three feces-amended sediments with different particle size in a laboratory study. In controlled conditions, using physiological saline to reduce cell death by osmotic shock, FC mortality rates exceeded FS mortality rates. These rates declined as sediment particle-size shrank and as temperature decreased. There was no interaction between these two factors in determining fecal bacteria persistence. The apparent halflives of FCs exceeded those of FS, even though mortality rates were higher, because of FC regrowth shortly after deposition. The FC/FS ratio is influenced by temperature, the presence of sediment, and sediment particle size. In warm conditions, FC regrowth increases FC/FS ratios to levels indicative of human contamination even where none clearly exists. These factors interfere with the interpretation of the FC/FS ratio and contribute to the difficulty of its use in agricultural settings. F BACTERIA concentrations in underlying sediment increase when cattle (Bos taurus) have direct access to streams. These bacteria survive for prolonged periods in stream sediment. Monitoring programs in agricultural watersheds that use indicator bacteria, such as FCs and FS, to assess water quality, may be biased as a result. Since bottom disturbance resuspends fecal bacteria in sediment (Gary and Adams, 1985; Sherer et al., 1988), they may also appear in overlying water for extended periods (Biskie et al., 1988; Jawson et al., 1982; Stephenson and Rychert, 1982). Consequently, Department of Agronomy, Univ. of Kentucky, Lexington, KY 405460091. Contribution of the Kentucky Agric. Exp. Stn., Lexington, KY. Received 12 June 1995. Corresponding author ([email protected]. edu). Published in J. Environ. Qual. 25:1216-1220 (1996). streams flowing through pastures typically exceed EPA limits for primary contact water (200 FCs/100 mL) long after cattle are removed (Howell et al., 1995; Jawson et al., 1982; Stephenson and Street, 1978). Small sediment particle size increases Escherichia coli (FC) survival (Tate, 1978). Extended survival of E. coli and Salmonella in sediments with high clay contents has been attributed to organic matter and nutrients held by the clay (Burton et al., 1987) and to physical protection in macropores. Moisture content, pH, sunlight, competition from indigenous soil microflora, and ambient temperature are other environmental factors that affect fecal bacteria survival in soil (Gerba et al., 1975). Higher temperature usually increases bacterial mortality but it can also promote FC regrowth in aquatic environments (Doran and Linn, 1979; Stephenson and Street, 1978). Howell et al. (1995) noted that the FC/FS ratio increased in streams of central Kentucky as temperature increased during spring. Ideally, the FC/FS ratio could be used to assess whether nonpoint source pollution comes from fecal contamination by humans (FC/FS > 4), domestic animals (FC/FS between 0.1 and 0.6), or wild animals (FC/FS<0.1) (Geldreich, 1976). However, the ratio has had questionable applicability in agricultural settings (Doran and Linn, 1979; Howell et al., 1995; Jawson et al., 1982) and is no longer recommended to distinguish between human and animal sources of contamination (APHA, 1992). This is due, in part, to the variability of FC/FS ratios. We were interested in accounting for the variability in FC/FS ratios that we have observed on a seasonal basis in agricultural watersheds (Howell et al., 1995). While the individual effect of ambient temperature and particle size on fecal bacteria survival have been studied, the interactive effect of these parameters has not. Furthermore, FC growth shortly after manure deposition may explain some of the variability we have observed in Abbreviations: EPA, Environmental Protection Agency; FC, fecal coliform; FS, fecal streptococci. HOWELL ET AL. : EFFECTS OF SEDIMENT PARTICLE water-monitoring studies (Howell et al., 1995). Our objectives were to observe whether interactive effects between sediment particle size and temperature influenced fecal bacteria growth and mortality in sediment. We also wanted to demonstrate that these effects, individual and interactive, confound use of the FC/FS ratio as an indicator of fecal contamination sources in agricultural settings. MATERIALS AND METHODS Treatment design for the experiment was a factorial arrangement of four substrates (physiological saline, sand, loam, and clay) and three incubation temperatures (4, 25, and 35°C). Particle-size distribution in each sediment ype was determined by sieving and the hydrometer method (Table 1). Two hundred and fifty grams of substrate was placed in 500-mL Nalgene bottles and a slurry containing 10 + 0.05 g of fresh cattle manure and 200 mL of sterile physiological saline (8.5 g NaCI/L water) was added to each bottle. Sediment were nonsterile but contained no fecal bacteria prior to adding the manure slurry. The mixtures were shaken on a mechanical flatbed shaker for 30 min to ensure thorough mixing and allowed to settle for 2 h. Fecal coliforms and FS were subsequently enumerated to determine initial concentrations. Each particle size and water treatment was incubated at 4, 25, and 35°C for 40 d. All sediment treatments were saturated and maintained at a constant volume by adding sterile physiological saline. The bottle caps were loosened during the incubations to permit air exchange while minimizing evaporation. The experiment was replicated three times with each replicate done in a different 40-d period.