Dennis J. Lye

Rooftop runoff as a source of contamination: A review☆

Scientific reports concerning chemical and microbiological contaminant levels of rainwater runoff from rooftop collection in both urban and rural areas are reviewed. This alternative source of water has been documented to often contain substantial amounts of contaminants. Studies describing levels of heavy metal contamination specific to runoff from rooftop catchment areas containing exposed metal surfaces are discussed. Depending upon the intended use, scientific evidence is also accumulating that various treatments and disinfections will be required prior to release of roof-runoff water either into surface waters or for more direct consumer usage. For microbial contamination, current proposed standards and guidelines regarding this type of water source are shown to vary widely worldwide. Scientific literature reveals a lack of clarity regarding water quality guidelines and health related standards for certain types of rooftop runoff. Studies suggests that rainwater collection systems which are properly designed, maintained, and treated may provide a valuable supplement to existing water supplies by reducing demand on community water supplies/infrastructure costs, enhancing effective management of storm water runoff, and increasing restoration of underground reservoirs through controlled infiltration.

Survey of ground, surface, and potable waters for the presence of Legionella species by EnviroAmpR PCR Legionella Kit, culture, and immunofluorescent staining

Abstract A total of 116 samples from numerous aquatic sources including water from faucets, showerheads, dental units, fire sprinklers, and surface waters were examined for the presence of Legionella by the EnviroAmpR Legionella PCR Kit, culture on BCYEα, or direct fluorescent antibody (DFA) staining. In addition, 58 ground water samples collected across the U.S. were also examined for the presence of Legionella species by PCR and by culture on BCYEα and BCYEα-GVPC. Of the 58 ground water samples tested by polymerase chain reaction (PCR) assay, 93% were positive by PCR, while only 7% were culture positive. Analysis of the remaining samples indicated that 52.7% of the samples were positive by PCR, 50.0% were positive by DFA, while 8.3% were culture positive. Although Legionella species were frequently detected by PCR in all water types examined, concentrations were typically less than 103 cells per ml. Legionalla species were detected much more frequently than was Legionella pneumophila in all aquatic samples. DFA results correlated well to PCR results but culture results were often negative for Legionella due to overgrowth, competition, or inhibition by background organisms. Results of this type of survey confirm the ubiquity of Legionella in aquatic environments, even ground waters. This study represents the first in depth survey of ground water across the United States.

Microbiology of rainwater cistern systems : a review

Abstract Many parts of the world do not have community water systems or have systems so badly in need of maintenance and repair that individually maintained water systems are still considered to be an appropriate and important source of potable water. The collection of rainwater by individuals using a cistern storage system is currently practiced in almost every nation in the world. A survey of American State Health Departments revealed that there are an estimated 253,815 cistern systems still in use in the United States. The scientific community is often reluctant to promote individually maintained water systems (such as rainwater catchment ‐ cistern storage systems) as being equal in quality to centrally maintained community water systems because domestic cistern systems are typically under maintained, not disinfected, and fall short of current microbiological guidelines for potable water. This review article provides information about the current use of rainwater catchment systems in the United States.

Development of an internal control for evaluation and standardization of a quantitative PCR assay for detection of Helicobacter pylori in drinking water.

ABSTRACT Due to metabolic and morphological changes that can prevent Helicobacter pylori cells in water from growing on conventional media, an H. pylori-specific TaqMan quantitative PCR (qPCR) assay was developed that uses a 6-carboxyfluorescein-labeled probe (A. E. McDaniels, L. Wymer, C. Rankin, and R. Haugland, Water Res. 39:4808-4816, 2005). However, proper internal controls are needed to provide an accurate estimate of low numbers of H. pylori in drinking water. In this study, the 135-bp amplicon described by McDaniels et al. was modified at the probe binding region, using PCR mutagenesis. The fragment was incorporated into a single-copy plasmid to serve as a PCR-positive control and cloned into Escherichia coli to serve as a matrix spike. It was shown to have a detection limit of five copies, using a VIC dye-labeled probe. A DNA extraction kit was optimized that allowed sampling of an entire liter of water. Water samples spiked with the recombinant E. coli cells were shown to behave like H. pylori cells in the qPCR assay. The recombinant E. coli cells were optimized to be used at 10 cells/liter of water, where they were shown not to compete with 5 to 3,000 cells of H. pylori in a duplex qPCR assay. Four treated drinking water samples spiked with H. pylori (100 cells) demonstrated similar cycle threshold values if the chlorine disinfectant was first neutralized by sodium thiosulfate.

Characterization of Aeromonas Virulence Using an Immunocompromised Mouse Model

An immunocompromised mouse model was used to characterize Aeromonas strains for their ability to cause opportunistic, extraintestinal infections. A total of 34 isolates of Aeromonas (A. hydrophila [n = 12]), A. veronii biotype sobria [n = 7], A. caviae [n = 4], A. enchelia [n = 4], A. allosaccharophila [n = 2], A. salmonicida (n = 4), and A. bestiarum [n = 1]) were introduced by intraperitoneal injection into immunocompetent or chemically compromised (using cyclophosphamide) mice. The ability of each isolate to persist in the liver and spleen tissue was monitored at 24 hours after exposure. A majority of A. hydrophila and A veronii v. sobria strains, but none of the isolates of other Aeromonas species, were capable of persistent colonization (<300 cells/mg spleen and liver tissue at 24 hours). The presence or absence of several putative virulence factors (cytotoxicity to HEp-2, lipase activity, elastase activity, and hemolysis) were determined for each isolate using in vitro tests. There were no correlations between the presence or absence of biochemical test results for putative virulence factors and persistence of the isolate in spleen and liver tissue at 24 hours.

Effects of Prolonged Chlorine Exposures upon PCR Detection of Helicobacter pylori DNA

The effect of low doses of free chlorine on the detection of Helicobacter pylori (H. pylori) cells by qPCR in tap water was monitored. Detection of sequences targeted to the ureA gene from preparations containing 107 cells/ml decreased about 2–4 logs by days 9 and 14, respectively. When duplicate suspensions of the 107 cells/ml were exposed to higher levels of chlorine, 0.2–2.2 mg/l, by day 9 and 14 there were 5 and 6 log decreases, respectively, in the detection of ureA gene. H. pylori target sequences (within suspended, intact cells at densities of 102–103 cells /ml) were rendered undetectable by qPCR analysis after 17 h of continuous exposure to low chlorine levels common to treated drinking water distribution systems. The persistence of DNA sequences within treated distribution systems detectable by qPCR may be as brief as 17 h especially for bacteria such as H. pylori which are known to occur in very low numbers within treated distribution systems. This study suggests that degradation of H. pylori DNA target sequences by chlorine levels commonly found within treated water distribution systems occurs within the average water retention times (2–3 days) commonly found in these systems.

A mouse model for characterization of gastrointestinal colonization rates among environmental Aeromonas isolates.

The colonization rates of 10 different environmental Aeromonas isolates were determined using a novel mouse–streptomycin pretreatment method. As demonstrated, alterations to the colon flora of mice pretreated with streptomycin allowed transient colonization by bacterial species normally excluded by host competition. A novel procedure is described for determining the colonization abilities of Aeromonas isolates under these conditions. The colonization rates of A.salmonicida, A. encheleia, and A. allosaccharophila were either negative or occurred randomly at low levels with respect to concentrations of the dosage consumed by the animals. In contrast, A. hydrophila, A. veronii biovar sobria, and A. caviae exhibited relatively high rates of mouse colon tissue colonization.

Evaluating virulence of waterborne and clinical Aeromonas isolates using gene expression and mortality in neonatal mice followed by assessing cell culture’s ability to predict virulence based on transcriptional response

Aims:  To assess the virulence of Aeromonas spp. using two models, a neonatal mouse assay and a mouse intestinal cell culture.

Gastrointestinal Colonization Rates for Human Clinical Isolates of Aeromonas Veronii Using a Mouse Model

A variety of environment-associated gastrointestinal infections have been associated with the Aeromonas group of bacteria which contain both non-virulent strains as well as virulent strains within a particular species. This study monitors the colonization rates of colon tissue in a mouse-streptomycin dose/response model involving isolates of Aeromonas veronii biovar sobria obtained from human clinical specimens. The ability to successfully colonize mouse colon tissues by the human clinical isolates was then compared with the rates achieved in a previous study of Aeromonas isolates obtained from environmental drinking water samples. Results suggest that strains of Aeromonas isolated from drinking water environmental samples contain pathogenic and virulence capabilities similar to those seen in Aeromonas veronii clinical isolates from human infections.

Predicting virulence of Aeromonas isolates based on changes in transcription of c-jun and c-fos in human tissue culture cells.

Aims:  To screen for the virulence potential of Aeromonas isolates based on the change in regulation of c‐jun and c‐fos in the human intestinal tissue culture cell line Caco‐2.