Gary K. Bissonnette

TREATMENT OF DOMESTIC WASTEWATER BY THREE PLANT SPECIES IN CONSTRUCTED WETLANDS

Three common Appalachian plant species (Juncus effusus L., Scirpus validus L., and Typha latifolia L.) were planted into small-scale constructed wetlands receivingprimary treated wastewater. The experimental design includedtwo wetland gravel depths (45 and 60 cm) and five plantingtreatments (each species in monoculture, an equal mixture of the three species, and controls without vegetation), with two replicates per depth × planting combination. Inflow rates (19 L day-1) and frequency (3 times day-1) were designed to simulate full-scale constructed wetlands as currently used for domestic wastewater treatmentin West Virginia. Influent wastewater and the effluent from each wetland were sampled monthly for ten physical, chemical and biological parameters, and plant demographic measurements were made. After passing through these trough wetlands, the average of all treatments showed a 70% reduction in total suspended solids (TSS) and biochemical oxygen demand (BOD), 50 to 60% reduction in nitrogen (TKN), ammonia and phosphate, anda reduction of fecal coliforms by three orders of magnitude. Depth of gravel (45 or 60 cm) had little effect on wetland treatment ability, but did influence Typha and Scirpus growth patterns. Gravel alone provided significant wastewater treatment, but vegetation further improved many treatment efficiencies. Typha significantly out-performedJuncus and Scirpus both in growth and in effluent quality improvement. There was also some evidence that the species mixture out-performed species monocultures.Typhawas the superior competitor in mixtures, but a decline in Typha growth with distance from the influent pipe suggested that nutrients became limiting or toxicities may have developed.

Fate of physical, chemical, and microbial contaminants in domestic wastewater following treatment by small constructed wetlands.

In order to evaluate the efficacy of constructed wetlands for treatment of domestic wastewater for small communities located in rural areas, small-scale wetland mesocosms (400 L each) containing two treatment designs (a mixture of Typha, Scirpus, and Juncus species; control without vegetation) were planted into two depths (45 or 60 cm) with pea gravel. Each mesocosm received 19 L/day of primary-treated domestic sewage. Mesocosms were monitored (inflow and outflow samples) on a monthly basis over a 2-year period for pH, total suspended solids (TSS), 5-day biochemical oxygen demand (BOD(5)), total Kjeldahl nitrogen (TKN), dissolved oxygen (DO), and conductivity. Microbiological analyses included enumeration of fecal coliforms, enterococci, Salmonella, Shigella, Yersinia, and coliphage. Significant differences between influent and effluent water quality for the vegetated wetlands (p<0.05) were observed in TSS, BOD(5), and TKN. Increased DO and reduction in fecal coliform, enterococcus, Salmonella, Shigella, Yersinia, and coliphage populations also were observed in vegetated wetlands. Greatest microbial reductions were observed in the planted mesocosms compared to those lacking vegetation. Despite marked reduction of several contaminants, wetland-treated effluents did not consistently meet final discharge limits for receiving bodies of water. Removal efficiencies for bacteria and several chemical parameters were more apparent during the initial year compared to the second year of operation, suggesting concern for long-term efficiency and stability of such wetlands.

Antibiotic resistant gram-negative bacteria in rural groundwater supplies☆

Abstract More than 250 coliform and noncoliform bacteria, isolated by standard membrane filtration methods from rural, untreated groundwater supplies, were examined for resistance to 16 antibiotics. All of the noncoliforms and 87% of the coliforms were resistant to at least one antibiotic, with resistance most commonly directed toward novobiocin, cephalothin, and ampicillin. The frequency of multiple antibiotic resistance (MAR) within each species was also determined. Approximately 60% of the coliforms were MAR, including 14, 64, and 94% of Escherichia coli, Citrobacter freundii, and Enterobacter cloacae isolates, respectively. In comparison, MAR was demonstrated by more than 95% of the noncoliforms and included isolates of Acinetobacter calcoaceticus, Aeromonas hydrophila, and Serratia marcescens. Ampicillin-resistant environmental isolates were assayed for ability to transfer resistance to ampicillin-sensitive strains of E. coli and Salmonella typhimurium. Environmental E. coli isolates were capable of in vitro transfer to both recipients in a nutrient-rich environment (trypticase soy broth) at frequencies ranging from 1.1 × 10−5 to 1.0 × 10−4. Isolates of C. freundii and Klebsiella pneumoniae exhibited resistance transfer to the E. coli recipient under similar conditions at reduced rates. None of the environmental isolates demonstrated transfer of ampicillin resistance when placed in an environment containing filter-sterilized well water. The presence of antibiotic resistant bacteria, and particularly MAR bacteria, in rural groundwater supplies used as drinking water sources may have important public health implications.

Assessment of the bacteriological quality of rural groundwater supplies in Northern West Virginia

A bacteriological survey was performed on 155 untreated, individual, rural groundwater supplies which included drilled wells, dug wells, and springs. Of these, 105 exceeded the Environmental Protection Agency (EPA) maximum contaminant level of one total coliform per 100 mL. Repeat sampling for 87 of the unacceptable 105 supplies indicated that 76 again exceeded the EPA standard limit. Masked coliforms were detected through confirmation tests in 11 water supplies that exhibited excessive noncoliform colonies (> 50 CFUs/membrane filter). Also, 48% of the supplies contained fecal coliforms and 62% contained fecal streptococci. Bacterial densities were related to the type of water supply with drilled wells containing fewer total coliforms, fecal coliforms, fecal streptococci, and heterotrophic plate count bacteria than dug wells or springs. Water supplies that were shallower, older, and lacking adequate casing characteristically were more heavily contaminated with sanitary indicator bacteria than supplies that were deeper, of more recent construction, and with sufficient casing.

Freshwater mussels as monitors of bacteriological water quality

A species of commercially obtained freshwater mussel (Elliptio complanata) was shown to actively filter, concentrate, and retain fecal coliform (FC) bacteria from a variety of freshwater stream environments. Fecal coliform densities in mussel viscera were maximum within 48 hr of in situ exposure. Significant FC reproduction did not occur in mussel viscera at temperatures below 30 °C. Variation in replicate FC determinations from mussels exposed in situ for 48 hr was less than that obtained directly from water samples. Numbers of FC concentrated in mussel viscera were retained for several hours after FC densities declined in the surrounding water, suggesting the utility of mussels to record recent episodes of fecal pollution.

Effect of acid mine water on Escherichia coli: structural damage

Escherichia coli B/5 12-h cultures were exposed to filter-sterilized acid mine water (AMW), fixed in situ, and examined for morphological changes by transmission electron microscopy, scanning electron microscopy, and x-ray spectrometry. Thin sections showed that layers of the Gram-negative envelope were altered and often lacking. Additionally, polar regions of the cell were frequently devoid of cytoplasm. AMW-exposed cells were distorted and had an amorphous substance associated with them. Spectra obtained by x-ray spectrometry suggested that this amorphous substance was cytoplasm rather than a mineral precipitate from AMW. Morphometric analyses of control and AMW-exposed populations showed significant differences in mean volume, length, and width of cells stressed in AMW; this indicates that smaller cells were selectively destroyed by the action of AMW. We concluded that loss of cytoplasm and cell lysis were the consequence of AMW damage to the bacterial envelope.

In situ persistence of indicator bacteria in streams polluted with acid mine drainage

Abstract Persistence of indicator organisms (total coliforms, fecal coliforms, and fecal streptococci) associated with natural samples of raw sewage was studied following in situ exposure to five aquatic environments. Three of these streams contained significant amounts of acid mine water (AMW) while the other two were relatively uncontaminated. Indicator organisms were rapidly killed upon exposure to the acid mine systems, whereas little reduction in numbers was observed in the uncontaminated streams. Seasonal changes affected survival of indicators as reflected by prolonged persistence at colder in situ water temperatures. The fecal coliform group was most susceptible to the AMW stress, while the fecal streptococci were most persistent. An enrichment technique resulted in substantially enhanced recovery of certain species of sublethally injured survivors of acid stress. Enrichment was particularly beneficial for recovery of AMW-injured fecal coliforms. Relatively little improvement in recovery of fecal streptococci was afforded by the enrichment technique.

Mobility and enhanced biodegradation of a dilute waste oil emulsion during land treatment

SummaryThe mobility and biodegradability in soil of a dilute waste oil emulsion generated by an aluminium rolling industry was investigated. Laboratory simulations and field evaluation of waste disposal suggested that the majority of the oil emulsion was retained in surface soil following application. However, potential leaching of waste to the subsurface was demonstrated, particularly at higher loading rates in soils of sandy texture. Strategies to enhance rates of biodegradation in surface soils were investigated, including fertilization and microbial inoculation. A single strain inoculum was obtained from a group of 81 isolates selected for their ability to partially mineralize the waste oil emulsion, and was tentatively characterized as a hydrocarbonoclasticCorynebacterium sp. Inoculation did not effectively stimulate waste removal in soil compared with fertilization, which significantly increased respiration and biodegradation. The maximum loss of the applied oil emulsion from soil was 30% during a 56-day in vitro incubation. Fertilized, aerated liquid waste emulsion was more rapidly degraded, resulting in loss of 65% of the waste emulsion within 18 days.

Comparative survival and injury of Candida albicans and bacterial indicator organisms in streams receiving acid mine drainage

Abstract Microbiological studies were conducted to assess the survival and injury characteristics of Candida albicans and indicator bacteria in streams impacted by acid mine water (AMW) and organic wastes. Persistence of pure cultures of C. albicans in three AMW-polluted streams was studied in situ using environmental membrane diffusion chambers. Survival of the fungus (at least 3 days in AMW) indicated prolonged tolerance to acid conditions. In comparison, Escherichia coli was killed within several hours of acid stress. Persistence studies also demonstrated that C. albicans was less sensitive to seasonal water temperature fluctuations than E. coli or Streptococcus faecium . In addition to its prolonged survival, C. albicans incurred minimal sublethal injury whereas identical conditions of exposure resulted in significant injury to traditional indicator bacteria. The failure of standard microbiological procedures to detect AMW-damaged bacteria compromises the accuracy of public health safety determinations in these waters. On the other hand, the ability of C. albicans to survive in AMW streams with marked resistance to injury suggests its potential as a favorable alternative sanitary indicator organism for such environments.

Injury and repair of Escherichia coli damaged by acid mine water

Abstract Pure culture suspensions of Escherichia coli B/5 were stressed by exposure to filter-sterilized acid mine water (AMW). Sublethally injured survivors were examined for their ability to repair in several resuscitation media under different conditions of pH, temperature and oxygen availability. The repair process was monitored as a function of time by periodically removing samples from the repair media and simultaneously plating on nonselective and selective media. E. coli was severely damaged by AMW; however, sublethally injured survivors repaired when placed under favorable conditions. Optimal repair occurred in trypticase soy broth supplemented with 0.3% yeast extract (TSYB) at pH 7.0 and 35°C. Resuscitation did not occur in TSYB at pH 9.0, at an incubation temperature of 20°C, or in the absence of oxygen. Lauryl tryptose broth (LTB), which is recommended for the presumptive isolation of fecal coliforms, was unable to facilitate repair of injury. The presence of the surfactant, sodium dodecyl sulfate, as well as the nutrient composition of LTB, appeared to be responsible for the inability of this medium to permit recovery of AMW-stressed E. coli.