A system for growing Geobacter sulfurreducens under anaerobic conditions in chemostats was developed in order to study the physiology of this organism under conditions that might more closely approximate those found in the subsurface than batch cultures. Geobacter sulfurreducens could be cultured under acetate-limiting conditions with fumarate or Fe(III)-citrate as the electron acceptor at growth rates between 0.04 and 0.09 h(-1). The molar growth yield was threefold higher with fumarate as the electron acceptor than with Fe(III), despite the lower mid-point potential of the fumarate/succinate redox couple. When growth was limited by availability of fumarate, high steady-state concentrations were detected, suggesting that fumarate is unlikely to be an important electron acceptor in sedimentary environments. The half-saturation constant, Ks, for acetate in Fe(III)-grown cultures (10 microM) suggested that the growth of Geobacter species is likely to be acetate limited in most subsurface sediments, but that when millimolar quantities of acetate are added to the subsurface in order to promote the growth of Geobacter for bioremediation applications, this should be enough to overcome any acetate limitations. When the availability of electron acceptors, rather than acetate, limited growth, G. sulfurreducens was less efficient in incorporating acetate into biomass but had higher respiration rates, a desirable physiological characteristic when adding acetate to stimulate the activity of Geobacter species during in situ uranium bioremediation. These results demonstrate that the ability to study the growth of G. sulfurreducens under steady-state conditions can provide insights into its physiological characteristics that have relevance for its activity in a diversity of sedimentary environments.

It has previously been demonstrated that [14C]-labeled polycyclic aromatic hydrocarbons (PAHs) can be oxidized to 14CO2 in anoxic, PAH-contaminated, marine harbor sediments in which sulfate reduction is the terminal electron-accepting process. However, it has not previously been determined whether this degradation of [14C]-PAHs accurately reflects the degradation of the in situ pools of contaminant PAHs. In coal tar-contaminated sediments from Boston Harbor, [14C]-naphthalene was readily oxidized to 14CO2, but, after 95 d of incubation under anaerobic conditions, there was no significant decrease in the detectable pool of in situ naphthalene in these sediments. Therefore, to better evaluate the anaerobic biodegradation of the in situ PAH pools, the concentrations of these contaminants were monitored for ca. 1 year during which the sediments were incubated under conditions that mimicked those found in situ. There was loss of all of the PAHs that were monitored (2-5 ring congeners), including high molecular weight PAHs, such as benzo[a]pyrene, that have not previously been shown to be degraded under anaerobic conditions. There was no significant change in the PAH levels in the sediments amended with molybdate to inhibit sulfate-reducing bacteria or in sediments in which all microorganisms had been killed with glutaraldehyde. In some instances, over half of the detectable pools of in situ 2-3 ring PAHs were degraded. In general, the smaller PAHs were degraded more rapidly than the larger PAHs. A distinct exception in the Boston Harbor sediment was naphthalene which was degraded very slowly at a rate comparable to the larger PAHs. In a similar in situ-like study of fuel-contaminated sediments from Liepaja Harbor, Latvia, there was no decline in PAH levels in samples that were sulfate-depleted. However, when the Latvia sediments were supplemented with sufficient sodium sulfate or gypsum to elevate pore water levels of sulfate to approximately 14-25 mM there was a 90% decline in the naphthalene and a 60% decline in the 2-methylnaphthalene pool within 90 days. These studies demonstrate for the first time that degradation by anaerobic microorganisms can significantly impact the in situ pools of PAHs in petroleum-contaminated, anoxic, sulfate-reducing harbor sediments and suggest that the self-purification capacity of contaminated harbor sediments is greater than previously considered.

In situ poly(hydroxyalkanoate) (PHA) levels and repeating-unit compositions were examined in stratified photosynthetic microbial mats from Great Sippewissett Salt Marsh, Mass., and Ebro Delta, Spain. Unlike what has been observed in pure cultures of phototrophic bacteria, the prevalence of hydroxyvalerate (HV) repeating units relative to hydroxybutyrate (HB) repeating units was striking. In the cyanobacteria-dominated green material of Sippewissett mats, the mole percent ratio of repeating units was generally 1HB:1HV. In the purple sulfur bacteria-dominated pink material the relationship was typically 1HB:2HV. In Sippewissett mats, PHA contributed about 0.5 to 1% of the organic carbon in the green layer and up to 6% in the pink layer. In Ebro Delta mats, PHA of approximately 1HB:2HV-repeating-unit distribution contributed about 2% of the organic carbon of the composite photosynthetic layers (the green and pink layers were not separated). Great Sippewissett Salt Marsh mats were utilized for more extensive investigation of seasonal, diel, and exogenous carbon effects. When the total PHA content was normalized to organic carbon, there was little seasonal variation in PHA levels. However, routine daily variation was evident at all sites and seasons. In every case, PHA levels increased during the night and decreased during the day. This phenomenon was conspicuous in the pink layer, where PHA levels doubled overnight. The daytime declines could be inhibited by artificial shading. Addition of exogenous acetate, lactate, and propionate induced two-to fivefold increases in the total PHA levels when applied in the daylight but had no effect when applied at night. The distinct diel pattern of in situ PHA accumulation at night appears to be related, in some phototrophs, to routine dark energy metabolism and is not influenced by the availability of organic nutrients.

The 1996 Safe Drinking Water Act amendments require the US Environmental Protection Agency and the Centers for Disease Control and Prevention to develop a national estimate of the occurrence of waterborne infectious disease that is attributable to public drinking water systems in the United States. Much of the information for developing the national estimate will be derived from epidemiologic data, and the primary outcome of this effort will be an estimate of the number of cases of gastrointestinal illness. While quantifying the number of these cases provides some measure of waterborne disease impact, the usefulness of this measure may be limited because the full spectrum of societal impact also involves consideration of the additional effects of these diseases such as hospitalization costs and lost productivity. If decision-makers wish to compare the impact of waterborne infectious diseases to the impact of some other public health concern (e.g. to aid in resource allocation decisions), then a comparison of case numbers may prove inadequate. Case numbers alone do not provide sufficient information about the severity of different illnesses. Society may value the avoidance of a few cases of severely debilitating illness more than it values the avoidance of many cases of mild illness. In order to compare disparate public health concerns, “burden of disease” measures that incorporate indicators of disease severity, costs, or societal values may prove essential for some types of decisions. We describe epidemiologic measures of severity, quality adjusted life years (QALYs), disability adjusted life years (DALYs), willingness-to-pay, and cost-of-illness methods commonly used for burden of disease estimates, and discuss how some of these summary measures of burden might be used for waterborne disease estimates.

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