Kumiko Oguma

Application of Cation-Coated Filter Method to Detection of Noroviruses, Enteroviruses, Adenoviruses, and Torque Teno Viruses in the Tamagawa River in Japan

ABSTRACT The occurrence of human enteric viruses in surface water in the Tamagawa River, Japan, was surveyed for 1 year, from April 2003 to March 2004. Sixty-four samples were collected from six sites along the river, and 500 ml of the sample was concentrated using the cation-coated filter method, which was developed in our previous study. This method showed recovery yields of 56% ± 32% (n = 37) for surface water samples inoculated with polioviruses. More than one kind of tested virus was detected in 43 (67%) of 64 samples by TaqMan PCR. Noroviruses and adenoviruses were detected in a high positive ratio; 34 (53%), 28 (44%), and 29 (45%) of 64 samples were positive for norovirus genotype 1 and genotype 2 and adenoviruses, respectively. The mean concentrations of norovirus genotype 1 or genotype 2 determined by real-time PCR were 0.087 and 0.61 genome/ml, respectively, showing much higher values in winter (0.21 genome/ml for genotype 1 and 2.3 genomes/ml for genotype 2). Enteroviruses were detected by both direct PCR (6 of 64 samples; 9%) and cell culture PCR (2 of 64 samples; 3%). Torque teno viruses, emerging hepatitis viruses, were also isolated in three samples (5%). The concentration of total coliforms and the presence of F-specific phages showed a high correlation with the presence of viruses, which suggested that the simultaneous use of total coliforms and F-specific phages as indicators of surface water may work to monitor viral contamination.

Photoreactivation of Escherichia coli after Low- or Medium-Pressure UV Disinfection Determined by an Endonuclease Sensitive Site Assay

ABSTRACT Photoreactivation of Escherichia coli after inactivation by a low-pressure (LP) UV lamp (254 nm), by a medium-pressure (MP) UV lamp (220 to 580 nm), or by a filtered medium-pressure (MPF) UV lamp (300 to 580 nm) was investigated. An endonuclease sensitive site (ESS) assay was used to determine the number of UV-induced pyrimidine dimers in the genomic DNA of E. coli, while a conventional cultivation assay was used to investigate the colony-forming ability (CFA) of E. coli. In photoreactivation experiments, more than 80% of the pyrimidine dimers induced by LP or MPF UV irradiation were repaired, while almost no repair of dimers was observed after MP UV exposure. The CFA ratios of E. coli recovered so that they were equivalent to 0.9-, 2.3-, and 1.7-log inactivation after 3-log inactivation by LP, MP, and MPF UV irradiation, respectively. Photorepair treatment of DNA in vitro suggested that among the MP UV emissions, wavelengths of 220 to 300 nm reduced the subsequent photorepair of ESS, possibly by causing a disorder in endogenous photolyase, an enzyme specific for photoreactivation. On the other hand, the MP UV irradiation at wavelengths between 300 and 580 nm was observed to play an important role in reducing the subsequent recovery of CFA by inducing damage other than damage to pyrimidine dimers. Therefore, it was found that inactivating light at a broad range of wavelengths effectively reduced subsequent photoreactivation, which could be an advantage that MP UV irradiation has over conventional LP UV irradiation.

Determination of Pyrimidine Dimers in Escherichia coli and Cryptosporidium parvum during UV Light Inactivation, Photoreactivation, and Dark Repair

ABSTRACT UV inactivation, photoreactivation, and dark repair ofEscherichia coli and Cryptosporidium parvum were investigated with the endonuclease sensitive site (ESS) assay, which can determine UV-induced pyrimidine dimers in the genomic DNA of microorganisms. In a 99.9% inactivation of E. coli, high correlation was observed between the dose of UV irradiation and the number of pyrimidine dimers induced in the DNA ofE. coli. The colony-forming ability of E. coli also correlated highly with the number of pyrimidine dimers in the DNA, indicating that the ESS assay is comparable to the method conventionally used to measure colony-forming ability. WhenE. coli were exposed to fluorescent light after a 99.9% inactivation by UV irradiation, UV-induced pyrimidine dimers in the DNA were continuously repaired and the colony-forming ability recovered gradually. When kept in darkness after the UV inactivation, however,E. coli showed neither repair of pyrimidine dimers nor recovery of colony-forming ability. When C. parvum were exposed to fluorescent light after UV inactivation, UV-induced pyrimidine dimers in the DNA were continuously repaired, while no recovery of animal infectivity was observed. When kept in darkness after UV inactivation, C. parvum also showed no recovery of infectivity in spite of the repair of pyrimidine dimers. It was suggested, therefore, that the infectivity of C. parvumwould not recover either by photoreactivation or by dark repair even after the repair of pyrimidine dimers in the genomic DNA.

Efficacy of UV irradiation in inactivating Cryptosporidium parvum oocysts.

ABSTRACT To evaluate the effectiveness of UV irradiation in inactivating Cryptosporidium parvum oocysts, the animal infectivities and excystation abilities of oocysts that had been exposed to various UV doses were determined. Infectivity decreased exponentially as the UV dose increased, and the required dose for a 2-log10 reduction in infectivity (99% inactivation) was approximately 1.0 mWs/cm2 at 20°C. However, C. parvum oocysts exhibited high resistance to UV irradiation, requiring an extremely high dose of 230 mWs/cm2 for a 2-log10 reduction in excystation, which was used to assess viability. Moreover, the excystation ability exhibited only slight decreases at UV doses below 100 mWs/cm2. Thus, UV treatment resulted in oocysts that were able to excyst but not infect. The effects of temperature and UV intensity on the UV dose requirement were also studied. The results showed that for every 10°C reduction in water temperature, the increase in the UV irradiation dose required for a 2-log10 reduction in infectivity was only 7%, and for every 10-fold increase in intensity, the dose increase was only 8%. In addition, the potential of oocysts to recover infectivity and to repair UV-induced injury (pyrimidine dimers) in DNA by photoreactivation and dark repair was investigated. There was no recovery in infectivity following treatment by fluorescent-light irradiation or storage in darkness. In contrast, UV-induced pyrimidine dimers in the DNA were apparently repaired by both photoreactivation and dark repair, as determined by endonuclease-sensitive site assay. However, the recovery rate was different in each process. Given these results, the effects of UV irradiation on C. parvum oocysts as determined by animal infectivity can conclusively be considered irreversible.

Assessment of groundwater pollution in Tokyo using PPCPs as sewage markers.

While the occurrence of pharmaceuticals and personal care products (PPCPs) in groundwater has typically been reported in bank filtration sites, irrigated fields, septic tanks, and sewage disposal practices, fewer studies have been conducted in highly urbanized areas, where infiltration of treated or untreated sewage is not supposed to be a source of groundwater recharge. Furthermore, little is known about the occurrence of various kinds of PPCPs in relation to microbial indicators in groundwater from different types of aquifers. Thus, we examined the city-wide occurrence of selected PPCPs (diethyltoluamide, crotamiton, ethenzamide, propyphenazone, carbamazepine, and caffeine) and E. coli in 50 groundwaters from unconfined aquifers (<30 m in depth) and confined aquifers (up to 500 m in depth) in Tokyo, where unintended groundwater contamination could take place due to decrepit sewer networks. PPCPs were detected in unconfined aquifers and springs (23/34 samples, 68%), and in confined aquifers (7/16 samples, 44%). Compared with published results for sewage influents, concentrations of PPCPs, excluding caffeine, were generally 1-2 orders of magnitude lower, while in some samples concentrations were quite comparable. The high occurrence rate of PPCPs, even in confined aquifers, indicated that such aquifers are not always protected from pollution by sewage near the land surface. Among the PPCPs analyzed, carbamazepine and crotamiton were most frequently detected, which would appear to be owing to their high persistence, combined with the high concentration of crotamiton in sewage. Crotamiton was detected in all four E. coli-positive groundwaters, and thus may potentially serve as a precautionary indicator of E. coli contamination. Using carbamazepine as a sewage marker, we estimated that 0.8%-1.7% of the dry-weather flow of sewage was leaking out into the unconfined aquifers.

Quantitative analysis of human enteric adenoviruses in aquatic environments

Aims:  The aim of this study was to determine human adenoviruses (HuAdVs) in aquatic environments by real‐time polymerase chain reaction (PCR).

Pepper mild mottle virus as an indicator and a tracer of fecal pollution in water environments: comparative evaluation with wastewater-tracer pharmaceuticals in Hanoi, Vietnam.

We analyzed pepper mild mottle virus (PMMoV) in 36 samples taken from surface water, wastewater, groundwater, tap water and bottled water in Hanoi, Vietnam. We then compared the occurrence and fates of PMMoV with pharmaceuticals and personal care products (PPCPs), which are known wastewater tracers. PMMoV was detected in 94% of the surface water samples (ponds, water from irrigated farmlands and rivers) and in all the wastewater samples. The PMMoV concentration ranged from 5.5×10(6)-7.2×10(6)copies/L in wastewater treatment plant (WWTP) influents, 6.5×10(5)-8.5×10(5)copies/L in WWTP effluents and 1.0×10(4)-1.8×10(6)copies/L in surface water. Among the sixty PPCPs analyzed, caffeine and carbamazepine had high detection rates in surface water (100% and 88%, respectively). In surface water, the concentration ratio of PMMoV to caffeine remained unchanged than that in WWTP influents, suggesting that the persistence of PMMoV in surface water was comparable to that of caffeine. The persistence and the large concentration ratio of PMMoV in WWTP influents to the method detection limit would account for its ubiquitous detection in surface water. In comparison, human enteric viruses (HEV) were less frequently detected (18-59%) than PMMoV in surface water, probably because of their faster decay. Together with the reported high human feces-specificity, our results suggested that PMMoV is useful as a sensitive fecal indicator for evaluating the potential occurrence of pathogenic viruses in surface water. Moreover, PMMoV can be useful as a moderately conservative fecal tracer for specifically tracking fecal pollution of surface water. PMMoV was detected in 38% of the groundwater samples at low concentrations (up to 19copies/L). PMMoV was not detected in the tap water and bottled water samples. In groundwater, tap water and bottled water samples, the occurrence of PPCPs and HEV disagreed with that of PMMoV, suggesting that PMMoV is not suitable as an indicator or a tracer in those waters.

Occurrence and formation potential of N-nitrosodimethylamine in ground water and river water in Tokyo

N-nitrosodimethylamine (NDMA), a disinfection byproduct of water and wastewater treatment processes, is a potent carcinogen. We investigated its occurrence and the potential for its formation by chlorination (NDMA-FP Cl2) and by chloramination (NDMA-FP NH2Cl) in ground water and river water in Tokyo. To characterize NDMA precursors, we revealed their molecular weight distributions in ground water and river water. We collected 23 ground water and 18 river water samples and analyzed NDMA by liquid chromatography-tandem mass spectrometry. NDMA-FP Cl2 was evaluated by chlorinating water samples with free chlorine for 24 h at pH 7.0 while residual free chlorine was kept at 1.0-2.0 mg Cl(2)/L. NDMA-FP NH2Cl was evaluated by dosing water samples with monochloramine at 140 mg Cl(2)/L for 10 days at pH 6.8. NDMA precursors and dissolved organic carbon (DOC) were fractionated by filtration through 30-, 3-, and 0.5 kDa membranes. NDMA concentrations were <0.5-5.2 ng/L (median: 0.9 ng/L) in ground water and <0.5-3.4 ng/L (2.2 ng/L) in river water. NDMA concentrations in ground water were slightly lower than or comparable to those in river water. Concentrations of NDMA-FP Cl2 were not much higher than concentrations of NDMA except in samples containing high concentrations of NH(3) and NDMA precursors. The increased NDMA was possibly caused by reactions between NDMA precursors and monochloramine unintentionally formed by the reaction between free chlorine and NH(3) in the samples. NDMA precursors ranged from 4 to 84 ng-NDMA eq./L in ground water and from 11 to 185 ng-NDMA eq./L in river water. Those in ground water were significantly lower than those in river water, suggesting that NDMA precursors were biodegraded, adsorbed, or volatilized during infiltration. The molecular weight of NDMA precursors in river water was dominant in the <0.5 kDa fraction, followed by 0.5-3 kDa. However, their distribution was inconsistent in ground water: one was dominant in the <0.5 kDa fraction, and the other in 0.5-3 kDa. Molecular weight distributions of NDMA precursors were very different from those of DOC. This is the first study to reveal the widespread occurrence and characterization of NDMA precursors in ground water.

Investigating sources and pathways of perfluoroalkyl acids (PFAAs) in aquifers in Tokyo using multiple tracers.

We employed a multi-tracer approach to investigate sources and pathways of perfluoroalkyl acids (PFAAs) in urban groundwater, based on 53 groundwater samples taken from confined aquifers and unconfined aquifers in Tokyo. While the median concentrations of groundwater PFAAs were several ng/L, the maximum concentrations of perfluorooctane sulfonate (PFOS, 990 ng/L), perfluorooctanoate (PFOA, 1800 ng/L) and perfluorononanoate (PFNA, 620 ng/L) in groundwater were several times higher than those of wastewater and street runoff reported in the literature. PFAAs were more frequently detected than sewage tracers (carbamazepine and crotamiton), presumably owing to the higher persistence of PFAAs, the multiple sources of PFAAs beyond sewage (e.g., surface runoff, point sources) and the formation of PFAAs from their precursors. Use of multiple methods of source apportionment including principal component analysis-multiple linear regression (PCA-MLR) and perfluoroalkyl carboxylic acid ratio analysis highlighted sewage and point sources as the primary sources of PFAAs in the most severely polluted groundwater samples, with street runoff being a minor source (44.6% sewage, 45.7% point sources and 9.7% street runoff, by PCA-MLR). Tritium analysis indicated that, while young groundwater (recharged during or after the 1970s, when PFAAs were already in commercial use) in shallow aquifers (<50 m depth) was naturally highly vulnerable to PFAA pollution, PFAAs were also found in old groundwater (recharged before the 1950s, when PFAAs were not in use) in deep aquifers (50-500 m depth). This study demonstrated the utility of multiple uses of tracers (pharmaceuticals and personal care products; PPCPs, tritium) and source apportionment methods in investigating sources and pathways of PFAAs in multiple aquifer systems.

Bacterial contamination of raw vegetables, vegetable-related water and river water in Ho Chi Minh City, Vietnam.

The study attempts to identify the potential routes of bacterial infection via consumption of raw vegetables, drinking water and vegetable-related water in Ho Chi Minh City (HCMC). Vegetables in the markets and restaurants had higher total coliforms and E. coli counts than the vegetables at the vegetable cultivation fields. In search of the potential contamination sources, it was found that vegetables are washed in nearby canals after harvesting. Those canals are contaminated with human and animal excreta, which in turn may contaminate the vegetables. At the markets, although the tap water was found to be free of microbes, contaminated and non-contaminated vegetables are mixed and washed in the same bowl, which may bring about further spreading of infectious bacteria. The results of this study suggested that an integrated countermeasure that incorporates reducing microbial contamination of canals, raising the awareness of microbial infection among the local farmers and wholesalers, and providing enough clean water to the food markets should be implemented to reduce the incidence of food-borne illness in HCMC.