Movement of traditional fecal indicator bacteria and source-tracking targets through septic drainfields.

The past three decades' data on outbreaks in the United States indicate that homes dependent on untreated groundwater (e.g. wells) for household drinking water that are also reliant on onsite treatment of household wastewater (e.g. septic systems) may be at greater risk for waterborne disease. While groundwater quality monitoring to protect public health has traditionally focused on the detection of fecal indicator bacteria, the application of emerging source tracking strategies may offer a more efficient means to identify pollution sources and effective means of remediation. This study compares the movement of common fecal indicator bacteria (E. coli and enterococci) with a chemical (optical brighteners, OB) and a molecular (Bacteroides HF183) source tracking (ST) target in small scale septic drainfield models in order to evaluate their potential utility in groundwater monitoring. Nine PVC column drainfield models received synchronized doses of primary-treated wastewater twice daily, with influent and effluent monitored bi-weekly over a 7-month period for all targets. Results indicate that E. coli and enterococci concentrations were strongly associated (Spearman's rank, p<0.05), and correlations between enterococci and optical brighteners were moderately strong. Bacteroides HF183 was significantly, but not strongly, associated with optical brighteners and both indicator bacteria (Point-biserial correlation, p<0.05), most likely due to its sporadic detection. Application of human ST marker monitoring in groundwaters at risk of contamination by human sewage is recommended, although consistent interpretation of results will rely on more detailed evaluation of HF183 incidence in source contamination waters.

Enterococcus growth on eelgrass (Zostera marina); implications for water quality.

Enterococci are fecal indicator bacteria used to monitor fecal pollution of recreational waters. When enterococci levels exceed health standards, fecal pollution is assumed as the cause. Enterococci growing on plants limit their usefulness as fecal indicator bacteria. Here we examined enterococcal growth on eelgrass in Mission Bay, CA where enterococci levels have exceeded water quality thresholds. A total of 69 eelgrass samples were collected from six sites, shaken to remove enterococci attached to plant surfaces and the eluant filtered onto culture media. Isolates were then identified to species using biochemical methods, and DNA typing by pulsed-field gel electrophoresis was done to assess clonality of strains. Enterococci concentrations among eelgrass ranged from 8 to 14 000 CFU g(-1) dry weight. The most predominant enterococcal species found were Enterococcus casseliflavus and E. hirae followed by E. faecalis. Cluster analysis indicated a high level of clonality among isolates across all species, with clonal isolates consistently associated with individual eelgrass samples. Finding high densities of E. casseliflavus, E. hirae and E. faecalis on eelgrass that included clonal strains indicates the capability of enterococcal growth on eelgrass. Amplification of enterococci on eelgrass presents challenges for regulatory agencies that interpret elevated levels of these bacteria as an indication of fecal pollution.

Associations between fecal indicator bacteria prevalence and demographic data in private water supplies in Virginia.

Over 1.7 million Virginians rely on private water sources to provide household water. The heaviest reliance on these systems occurs in rural areas, which are often underserved with respect to available financial resources and access to environmental health education. This study aimed to identify potential associations between concentrations of fecal indicator bacteria (FIB) (coliforms, Escherichia coli) in over 800 samples collected at the point-of-use from homes with private water supply systems and homeowner-provided demographic data (household income and education). Of the 828 samples tested, 349 (42%) of samples tested positive for total coliform and 55 (6.6%) tested positive for E. coli. Source tracking efforts targeting optical brightener concentrations via fluorometry and the presence of a human-specific Bacteroides marker via quantitative real-time polymerase chain reaction (qPCR) suggest possible contamination from human septage in over 20 samples. Statistical methods implied that household income has an association with the proportion of samples positive for total coliform, though the relationship between education level and FIB is less clear. Further exploration of links between demographic data and private water quality will be helpful in building effective strategies to improve rural drinking water quality.

A high-precision mechanical absolute-rotation sensor.

We have developed a mechanical absolute-rotation sensor capable of resolving ground rotation angle of less than 1 nrad/√Hz above 30 mHz and 0.2 nrad/√Hz above 100 mHz about a single horizontal axis. The device consists of a meter-scale beam balance, suspended by a pair of flexures, with a resonance frequency of 10.8 mHz. The center of mass is located 3 µm above the pivot, giving an excellent horizontal displacement rejection of better than 3 × 10(-5) rad/m. The angle of the beam is read out optically using a high-sensitivity autocollimator. We have also built a tiltmeter with better than 1 nrad/√Hz sensitivity above 30 mHz. Co-located measurements using the two instruments allowed us to distinguish between background rotation signal at low frequencies and intrinsic instrument noise. The rotation sensor is useful for rotational seismology and for rejecting background rotation signal from seismometers in experiments demanding high levels of seismic isolation, such as Advanced Laser Interferometer Gravitational-wave Observatory.

Comparison of Enterococcus species diversity in marine water and wastewater using Enterolert and EPA Method 1600.

EPA Method 1600 and Enterolert are used interchangeably to measure Enterococcus for fecal contamination of public beaches, but the methods occasionally produce different results. Here we assess whether these differences are attributable to the selectivity for certain species within the Enterococcus group. Both methods were used to obtain 1279 isolates from 17 environmental samples, including influent and effluent of four wastewater treatment plants, ambient marine water from seven different beaches, and freshwater urban runoff from two stream systems. The isolates were identified to species level. Detection of non-Enterococcus species was slightly higher using Enterolert (8.4%) than for EPA Method 1600 (5.1%). E. faecalis and E. faecium, commonly associated with human fecal waste, were predominant in wastewater; however, Enterolert had greater selectivity for E. faecalis, which was also shown using a laboratory-created sample. The same species selectivity was not observed for most beach water and urban runoff samples. These samples had relatively higher proportions of plant associated species, E. casseliflavus (18.5%) and E. mundtii (5.7%), compared to wastewater, suggesting environmental inputs to beaches and runoff. The potential for species selectivity among water testing methods should be considered when assessing the sanitary quality of beaches so that public health warnings are based on indicators representative of fecal sources.

Quantitative analysis of microbial contamination in private drinking water supply systems.

Over one million households rely on private water supplies (e.g. well, spring, cistern) in the Commonwealth of Virginia, USA. The present study tested 538 private wells and springs in 20 Virginia counties for total coliforms (TCs) and Escherichia coli along with a suite of chemical contaminants. A logistic regression analysis was used to investigate potential correlations between TC contamination and chemical parameters (e.g. NO3(-), turbidity), as well as homeowner-provided survey data describing system characteristics and perceived water quality. Of the 538 samples collected, 41% (n = 221) were positive for TCs and 10% (n = 53) for E. coli. Chemical parameters were not statistically predictive of microbial contamination. Well depth, water treatment, and farm location proximate to the water supply were factors in a regression model that predicted presence/absence of TCs with 74% accuracy. Microbial and chemical source tracking techniques (Bacteroides gene Bac32F and HF183 detection via polymerase chain reaction and optical brightener detection via fluorometry) identified four samples as likely contaminated with human wastewater.

Picoradian deflection measurement with an interferometric quasi-autocollimator using weak value amplification.

We present an "interferometric quasi-autocollimator" that employs weak value amplification to measure angular deflections of a target mirror. The device has been designed to be insensitive to all translations of the target. We present a conceptual explanation of the amplification effect used by the device. An implementation of the device demonstrates sensitivities better than 10 picoradians per root hertz between 10 and 200 Hz.

Fecal bacterial diversity of human-habituated wild chimpanzees (Pan troglodytes schweinfurthii) at Mahale Mountains National Park, Western Tanzania.

Although the intestinal flora of chimpanzees has not been studied, insight into this dynamic environment can be obtained through studies on their feces. We analyzed fecal samples from human-habituated, wild chimpanzees at Mahale Mountains National Park, Tanzania, and compared microbial community profiles to determine if members of the same social group were similar. Between July and December 2007, we collected fresh fecal samples from 12 individuals: four juveniles, four adolescents, and four adults, including three parent-offspring pairs. Each sample was analyzed using Terminal-Restriction Fragment Length Polymorphism of amplified 16S rRNA genes. Twelve different profiles were generated, having between 1 and 15 Terminal-Restriction Fragments (T-RFs). Overall, a total of 23 different T-RFs were produced. Putative assignments of T-RFs corresponded to the phyla Firmicutes (Clostridia, Bacilli, and Lactobacilli), Bacteroidetes, Tenericutes (Mollicutes Class), Actinobacteria, and Proteobacteria, as well as to uncultured or unidentified organisms. Firmicutes and Bacteroidetes phyla and Mollicutes Class were the most commonly assigned in 11, 8, and 8 of the samples, respectively, with this being the first report of Mollicutes in wild chimpanzees. Principal Components Analysis (PCA) revealed clustering of nine samples, and 80.5% of the diversity was accounted for by three samples. Morisita indices of community similarity ranged between 0.00 and 0.89, with dissimiliarity (<0.5) between most samples when compared two at a time. Our findings suggest that, although phylotypes are common among individuals, profiles among members of the same social group are host-specific. We conclude that factors other than social group, such as kinship and age, may influence fecal bacterial profiles of wild chimpanzees, and recommend that additional studies be conducted.

Exposing water samples to ultraviolet light improves fluorometry for detecting human fecal contamination.

Fluorometry identifies human fecal contamination by detecting optical brighteners in environmental waters. Because optical brighteners are sensitive to sunlight, we determined if we could improve fluorometry by exposing water samples to ultraviolet (UV) light to differentiate between optical brighteners and other fluorescing organic compounds. Optical brighteners were likely present when the relative percentage difference in fluorometric value of the water before and after UV light exposure was >30% (glass cuvettes, 30 min exposure) or >15% (polymethacrylate cuvettes, 5 min exposure). In a blind study, we correctly identified the presence or absence of optical brighteners in 178 of 180 (99%) of the samples tested with a more expensive field fluorometer and in 175 of 180 (97%) of the samples tested with a less expensive handheld fluorometer. In the field, the method correctly identified two negative and three positive locations for human fecal contamination. When combined with counts of fecal bacteria, the new fluorometric method may be a simple, quick, and easy way to identify human fecal contamination in environmental waters.

Comparison of seven protocols to identify fecal contamination sources using Escherichia coli.

Microbial source tracking (MST) uses various approaches to classify fecal-indicator microorganisms to source hosts. Reproducibility, accuracy, and robustness of seven phenotypic and genotypic MST protocols were evaluated by use of Escherichia coli from an eight-host library of known-source isolates and a separate, blinded challenge library. In reproducibility tests, measuring each protocol's ability to reclassify blinded replicates, only one (pulsed-field gel electrophoresis; PFGE) correctly classified all test replicates to host species; three protocols classified 48-62% correctly, and the remaining three classified fewer than 25% correctly. In accuracy tests, measuring each protocol's ability to correctly classify new isolates, ribotyping with EcoRI and PvuII approached 100% correctclassification but only 6% of isolates were classified; four of the other six protocols (antibiotic resistance analysis, PFGE, and two repetitive-element PCR protocols) achieved better than random accuracy rates when 30-100% of challenge isolates were classified. In robustness tests, measuring each protocol's ability to recognize isolates from nonlibrary

Phenotypic library-based microbial source tracking methods: efficacy in the California collaborative study.

As part of a larger microbial source tracking (MST) study, several laboratories used library-based, phenotypic subtyping techniques to analyse fecal samples from known sources (human, sewage, cattle, dogs and gulls) and blinded water samples that were contaminated with the fecal sources. The methods used included antibiotic resistance analysis (ARA) of fecal streptococci, enterococci, fecal coliforms and E. coli; multiple antibiotic resistance (MAR) and Kirby-Bauer antibiotic susceptibility testing of E. coli; and carbon source utilization for fecal streptococci and E. coli. Libraries comprising phenotypic patterns of indicator bacteria isolated from known fecal sources were used to predict the sources of isolates from water samples that had been seeded with fecal material from the same sources as those used to create the libraries. The accuracy of fecal source identification in the water samples was assessed both with and without a cut-off termed the minimum detectable percentage (MDP). The libraries (approximately 300 isolates) were not large enough to avoid the artefact of source-independent grouping, but some important conclusions could still be drawn. Use of a MDP decreased the percentage of false-positive source identifications, and had little effect on the high percentage of true-positives in the most accurate libraries. In general, the methods were more prone to false-positive than to false-negative errors. The most accurate method, with a true-positive rate of 100% and a false-positive rate of 39% when analysed with a MDP, was ARA of fecal streptococci. The internal accuracy of the libraries did not correlate with the accuracy of source prediction in water samples, showing that one should not rely solely on parameters such as the average rate of correct classification of a library to indicate its predictive capabilities.

Antibiotic resistance profiles to determine sources of fecal contamination in a rural Virginia watershed.

Antibiotic resistance analysis (ARA) was used to determine if enterococci of human origin were present in a stream (Spout Run) that passes through a rural nonsewered community (Millwood, VA). Millwood consists of 82 homes, all served by individual septic systems, and Spout Run drains a 5,800-ha karst topography watershed that contains large populations of livestock and wildlife. Periodic monitoring by state regulatory officials had resulted in Spout Run being placed on the Virginia impaired stream list and Millwood being categorized as an at-risk community. Stream samples were collected monthly and analyzed for fecal coliforms and enterococci (May 1999-May 2000); ARA was performed on enterococci stream isolates on a quarterly basis. All 117 stream samples were positive for fecal coliforms, and 32% exceeded the Virginia recreational water standard (1,000 fecal coliforms/100 mL). A library of 1,174 known source Enterococcus isolate antibiotic resistance profiles was constructed, and yielded correct classification rates of 94.6% for 203 human isolates, 93.7% for 734 livestock isolates, and 87.8% for 237 wildlife isolates. Antibiotic resistance analysis of 2,012 enterococcal isolates recovered from stream samples indicated isolates of human origin appeared throughout the stream as it passed through Millwood, with a yearly average of approximately 10% human, 40% wildlife, and 50% livestock. There were no human origin isolates in samples collected upstream from Millwood, and the percent human origin isolates declined downstream from Millwood. While a human signature was found in Spout Run, it was small compared with the proportion of isolates from livestock and wildlife.