Chlorine disinfection of blended municipal wastewater effluents.

Blending is used in the wastewater industry to manage wet-weather events. Wastewater is treated through primary clarification, with flows in excess of the hydraulic capacity of the secondary system being directed to effluent disinfection. Before disinfection, the primary clarified effluent is "blended" with effluents that have been treated through the secondary system. The combined or "blended" effluents are then disinfected before being discharged to receiving waters. This study evaluated the effectiveness of chlorine to disinfect blended effluents. Experiments were conducted at bench-scale on primary and secondary effluents and three ratios of primary to secondary effluent (1:9, 3:7, and 5:5) from three publicly owned treatment works. Results from this study found that blending 10% or more primary effluent with secondary reduces the efficacy of chlorine disinfection, and coliphage survived chlorine disinfection better than bacterial indicator organisms. A simple empirical model for predicting indicator organism densities following chlorine disinfection was developed using data from this research.

The widespread occurrence of the enterohemolysin gene ehlyA among environmental strains of Escherichia coli.

The putative virulence factor enterohemolysin, encoded by the ehlyA gene, has been closely associated with the pathogenic enterohemorrhagic Escherichia coli (EHEC) group. Escherichia coli isolates from effluents from seven geographically dispersed municipal wastewater treatment plants were screened for the presence of enterohemolysin. A total of 338 E. coli isolates were found to express the ehlyA gene. However, none of the isolates contained the toxin-encoding genes (stxA or stxB) associated with EHEC. Two of the 338 isolates possessed the virulence factor intimin, encoded by the eae gene. These findings suggest that the ehlyA gene may be widely distributed among non-EHEC isolates in the environment.

The role of flushing dental water lines for the removal of microbial contaminants.

OBJECTIVES: This study was designed to determine the role of flushing dental water lines for the removal of heterotrophic plate count bacteria, Legionella spp., and free-living protozoa. METHODS: Forty dental offices were surveyed in the study. An initial sample and a sample taken after three minutes of flushing were obtained from the air/water syringe at each location. All samples were quantitatively analyzed for heterotrophic bacteria using three bacteriological procedures. The samples were analyzed for the presence of Legionella spp. using cultural, immunological, and molecular procedures and for the occurrence of free-living protozoa using a killed bacteria plate procedure. RESULTS: The flushing process reduced the level of heterotrophic plate count bacteria by 1.1 to 1.5 log10 CFU/ml. Compliance with recommendations for bacterial levels varied depending on the methodology employed in the analysis. The flushing process did not reduce the occurrence of Legionella spp. or free-living protozoa. CONCLUSION: The results support recent U.S. Centers for Disease Control and Prevention recommendations that the process of flushing dental water lines cannot be relied upon as a sole means of reliably improving the quality of water used in dental treatment.

Propidium monoazide reverse transcriptase PCR and RT-qPCR for detecting infectious enterovirus and norovirus.

Presently there is no established cell line or small animal model that allows for the detection of infectious human norovirus. Current methods based on RT-PCR and RT-qPCR detect both infectious and non-infectious virus and thus the conclusions that may be drawn regarding the public health significance of positive findings are limited. In this study, PMA RT-PCR and RT-qPCR assays were evaluated for selective detection of infectious poliovirus, murine norovirus (MNV-1), and Norwalk virus. Viruses were inactivated using heat, chlorine, and ultraviolet light (UV). Infectious and non-infectious viruses were treated with PMA before RT-PCR and RT-qPCR. PMA RT-PCR was able to differentiate selectively between infectious and heat and chlorine inactivated poliovirus. PMA RT-PCR was able to differentiate selectively between infectious and noninfectious murine norovirus only when inactivated by chlorine. However, PMA RT-PCR could not differentiate infectious Norwalk virus from virus particles rendered non-infectious by any treatment. PMA RT-PCR assay was not able to differentiate between infectious and UV inactivated viruses suggesting that viral capsid damage may be necessary for PMA to enter and bind to the viral genome. PMA RT-PCR on naked MNV-1 and Norwalk virus RNA suggest that PMA RT-PCR can be used to detect intact, potentially infectious MNV-1 and Norwalk viruses and can be used to exclude the detection of free viral RNA by PCR assay.

Detection of intrinsic vancomycin resistant enterococci in animal and human feces.

Fecal samples from animal species and humans were analyzed by quantitative culture for enterococci and vancomycin resistant enterococci (VRE). Each host species carried enterococci which exhibited intrinsic intermediate resistance to vancomycin and sensitivity to teicoplanin (Van C phenotype). The carriage rate in humans was 9%. Carriage rates varied among animal species with the highest percentages being found in deer, duck, goose, horse and turkey.

Survival of Salmonella in waste egg wash water.

Waste wash waters from chicken egg-processing facilities can harbor high densities of bacteria, including salmonellae. For this study, we enumerated total coliforms, Escherichia coli, and Salmonella spp. in the egg wash waters of a large egg producer. We then determined how long these organisms would survive at temperatures of 5, 15, and 25 degrees C. We found that the fraction of salmonellae surviving over time at a given temperature was comparable to the fraction of indicator organisms that survived. We also found that the survival of these organisms varied with temperature, with 16, 8, and < 2 days being required for a 90% reduction of Salmonella in waste wash water held at 5, 15, and 25 degrees C, respectively. Finally, we noted that the response of laboratory-derived cultures to environmental stresses mimics the response of the indigenous microbial population, but individual cells within that population may survive for longer periods than laboratory-cultured strains.

Production of Enterotoxin by Vibrio vulnificus Isolates.

Weakly virulent isolates of Vibrio vulnificus that were lethal only to simultaneously iron-overloaded and immunosuppressed mice were tested for ability to cause fluid accumulation in the permanently ligated rabbit ileal loop. Unlike the highly virulent isolates, which caused septicemia and death in rabbits, these isolates caused significant fluid accumulation in the rabbit loops. Fluid accumulation was also observed when culture filtrates were tested, indicating the existence of an enterotoxin. Enterotoxin activity did not correlate with the hemolysin or protease activities. Only one of three enterotoxigenic isolates caused diarrhea when administered to temporarily ligated rabbit ileal loops, suggesting involvement of some other pathogenic determinant(s) such as colonization.

Oral Infectivity of Vibrio vulnificus in Suckling Mice.

Lethal doses of 11 clinical and environmental isolates of Vibrio vulnificus were determined in suckling mice after oral challenge. With one exception, isolates that were virulent to iron-overloaded adult mice after intraperitoneal inoculation were highly lethal to the infant mice (>50% lethality at 105 CFU/mouse). The virulent isolate that failed to kill infant mice at 105 CFU had lost its invasiveness. Conditionally virulent isolates that were virulent only to simultaneously iron-overloaded and immunosuppressed adult mice required > 109 CFU to kill the infant mice. Avirulent isolates failed to kill at >109 CFU/mouse. There were no significant differences in the lethalities of clinical and environmental isolates. These findings demonstrated a close correlation between virulence in the iron-overloaded adult mouse and infectivity by the oral route.