Shinichiro Ohgaki

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.

Detection of Noroviruses in Tap Water in Japan by Means of a New Method for Concentrating Enteric Viruses in Large Volumes of Freshwater

ABSTRACT A virus concentration method using a cation-coated filter was developed for large-volume freshwater applications. Poliovirus type 1 (LSc 2ab Sabin strain) inoculated into 40 ml of MilliQ (ultrapure) water was adsorbed effectively to a negatively charged filter (Millipore HA, 0.45-μm pore size) coated with aluminum ions, 99% (range, 81 to 114%) of which were recovered by elution with 1.0 mM NaOH (pH 10.8) following an acid rinse with 0.5 mM H2SO4 (pH 3.0). More than 80% poliovirus recovery yields were obtained from 500-ml, 1,000-ml, and 10-liter MilliQ water samples and from tap water samples. This method, followed by TaqMan PCR detection, was applied to determine the presence of noroviruses in tap water in Tokyo, Japan. In a 14-month survey, 4 (4.1%) and 7 (7.1%) of 98 tap water samples (100 to 532 liters) contained a detectable amount of noroviruses of genotype 1 and genotype 2, respectively. This method was proved to be useful for surveying the occurrence of enteric viruses, including noroviruses, in large volumes of freshwater.

Effect of pore structure of membranes and module configuration on virus retention

We measured virus retention by many types of membranes including microfiltration membranes, ultrafiltration membranes and nanofiltration membranes. We succeeded in evaluating quantitatively virus retention in a very high retention range by employing coliphage Qβ and T4 as model viruses. Qβ, which is the smaller virus in this study, penetrated all tested pieces of ultrafiltration membranes and nanofiltration membranes, though the retention was very high such as in the range of 99-99.9999%. The analysis of the polyethylene glycol retention data has shown that the leakage of viruses is caused by abnormally larger pores which are not included in the main pore size distribution. The diameter of the abnormal pores was estimated from the results of retention of different coliphages. The leakage of Qβ was also observed in the case of inorganic ceramic ultrafiltration membranes, but T4, which is larger than Qβ, cannot penetrate them. Some types of microfiltration membrane have shown higher retention than ultrafiltration membranes and nanofiltration membranes. This suggests the possibility that we can develop high virus retention membranes with low filtration resistance.

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.

Mass transfer mechanism in a porous riverbed

Abstract The mass transfer mechanism in a porous riverbed was investigated in two experimental open channels which have porous beds composed of 1.9 and 4.08 cm diameter ceramic balls, respectively. Vertical diffusion coefficients in the porous bed were determined by analyzing the diffusional pattern of tracer (NaCl) from the overlying water into the deeper region of the porous bed. Velocity profiles both over and under the water-bed interface and turbulence at the interface was measured. The diffusion coefficients in the porous bed were expressed by the product of velocity component and mixing length along every depth of the porous bed. Near the interface, the diffusion coefficient was expressed by the product of the turbulent intensity and the void scale of the porous layer. In the deeper region of the porous bed, the diffusion coefficient was expressed by the product of the time-averaged velocity and the void scale.

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).

Biological powdered activated carbon (BPAC) microfiltration for wastewater reclamation and reuse

Abstract The experiment was conducted to evaluate a biological powdered activated carbon (BPAC) microfiltration (MF) system as an alternative for wastewater reclamation and reuse. A synthetic secondary sewage effluent contains refractory organic compounds such as humin, tannin, lignin, protein and high molecular carbohydrates as well as coliphage Qs as a model virus. The system performance was investigated at the activated carbon concentration of 20 g/1, water temperature 25°C and transmembrane pressure of 55 kPa. It was noted that organic removal occurred mainly at the membrane module. This was caused by the accumulation of the powdered activated carbon in the membrane module. The average organic removal efficiency was 83%, resulting in an effluent TOC concentration of 1–2 mg/1. The performance of the process did not deteriorate at water temperature of 15°C, showing an organic removal efficiency of 89.6%. It was estimated that the higher removal efficiency at lower water temperature was mainly due to the less self-degradation of microorganisms because the permeate flux of the membrane was maintained at the same level by increasing the transmembrane pressure up to 80 kPa. The removal of virus by the BPAC-MF system was significant. From the mass balance at steady state, 99.9997% of fed coliphage Qs was removed from the system. Especially coliphage Qs showed a strong adsorbability on powdered activated carbon (PAC). For 1 h contact with PAC, the removal of Qs was 99.999% even at PAC concentrations of 0.55 g/l. It was obvious that the virus removed was inactivated in the system.

Effect of turbulence on nitrifying biofilms at non-limiting substrate conditions

Abstract The effect of turbulence on nitrifying biofilms was studied in five cylindrical PVC (polyvinyl chloride) reactors, each having ten biofilm sampling taps, over a period of 196 days. Bulk water in the reactors was stirred by paddles at 32, 92, 140, 278 and 500 rpm and the turbulent intensities measured at 10 mm from the wall were 0.6, 1.5, 2.6, 4.4 and 8.9 cm/s. Biofilms appeared as isolated colonies and continued to grow as filament-type biofilms. Higher turbulence resulted in higher NH4-N flux and higher areal biomass density. Turbulent diffusion of substrates and by-products in the vicinity of filament-type biofilms must have resulted in the above phenomena. Photographic observation of the biofilm surfaces on sampling taps showed uniform biofilm filaments at higher turbulent intensities and large variation in the height of filaments at low turbulent intensities. Substrate flux and biofilm structure (areal density, filament height and cross-sectional area of filament) are inter-related parameters and are strongly affected by turbulence near the biofilm. Substrate flux is expressed as a power function of turbulent intensity, volumetric density and substrate concentration for filament-type biofilm when substrates are non-limiting.