ABSTRACT Pulsed lamps which irradiates ultraviolet(UV) in high luminance has been developed. The experiment to inactivate of cryptosporidium parvum was conducted with the pulsed lamp. Also the effect of inactivate to high turbidity solution was investigated. The experiment examined the comparison and the evaluation methods (growth vitality assays and infectivity assays) with other kinds of UV lamps. As a result, higher capability of the pulsed lamp was demonstrated in the high UV dose area, compared to other UV lamps. The difference between inactivation efficiencies by the low pressure mercury lamp, the medium pressure mercury lamp and the pulsed lamp was not significant in the effect of inactivate to high turbidity solution. However, the tailing phenomenon was not observed with the pulse lamp while it was with the low pressure mercury lamp and the medium pressure mercury lamp when evaluating by excystation method. Moreover, sensitivity of cryptosporidium parvum to UV irradiation was in the order of the cell culture method, the mouse infection method, and excystation method in this experiment
Journal of Water and Environment Technology, Vol.3, No.1, 2005 - 29 - The effect of inactivation for C.parvum by Pulse lamp. T. IWASAKI ※ 1 ，S. KINISHITA ※ 1 ，M. OHTAKI ※ 2 ，K. HIKOSAKA ※ 3 ，Y. NAKAI ※ 3 ※1 Applied Optics Research＆Development Department，Applied Optics Division，Iwasaki Electric Co.,Ltd．1-1 Ichiriyama-cho Gyoda-city，Saitama-pref．361-8505，JAPAN (E-mail：iwasaki-tatsuyuki@eye.co.jp ， kinoshita-shinobu@eye.co.jp) ※2 Graduate School of Humanities and Science，Ochanomizu University，2-1-1 Otsuka， Bunkyo-ku Tokyo 112-8610，JAPAN(E-mail：otaki@cc.ocha.ac.jp) ※3 Department of Animal Health and Management，Tohoku University，1-1 Tsutsumidori- Amamiyamachi,，Aoba-ku，Sendai-city，Miyagi-ptef．981-8555，JAPAN (E-mail：hikosaka@naro.affrc.go.jp，nakai@bios.tohoku.ac.jp) ABSTRACT Pulsed lamps which irradiates ultraviolet(UV) in high luminance has been developed. The experiment to inactivate of cryptosporidium parvum was conducted with the pulsed lamp. Also the effect of inactivate to high turbidity solution was investigated. The experiment examined the comparison and the evaluation methods (growth vitality assays and infectivity assays) with other kinds of UV lamps. As a result, higher capability of the pulsed lamp was demonstrated in the high UV dose area, compared to other UV lamps. The difference between inactivation efficiencies by the low pressure mercury lamp, the medium pressure mercury lamp and the pulsed lamp was not significant in the effect of inactivate to high turbidity solution. However, the tailing phenomenon was not observed with the pulse lamp while it was with the low pressure mercury lamp and the medium pressure mercury lamp when evaluating by excystation method. Moreover, sensitivity of cryptosporidium parvum to UV irradiation was in the order of the cell culture method, the mouse infection method, and excystation method in this experiment. KEYWORDS UV-radiation，pulsed lamp，cryptosporidium，inactivation． INTRODUCTIONI UV disinfection and sterilization are used as alternative technologies to the chlorine disinfection of secondarily processed sewage, with some European countries using these methods to disinfect drinking water. Disinfection through UV means is thus common worldwide. It should also be noted that the struggle against pathogenic microbes such as the Cryptosporidium parvum ("C. parvum") protozoan present in drinking water sources now represents a major issue to be addressed by drinking water authorities in their efforts to supply safe water. C. parvum is particularly resistant to chlorine, meaning that it cannot be easily treated using conventional means of chlorine disinfection. With this in mind, the introduction of UV technology is something which the Ministry of Health, Labour and Welfare is considering as a means of disinfecting drinking water in Japan and one which may well have even more widespread applications. This paper reports a recently conducted comparative test on the ability to render C. parvum inactive featuring low-, medium-pressure mercury lamps(330W&1kW) as well as newly introduced pulse lamps, which irradiate short bursts of high-luminance energy. The report also outlines a comparison of the evaluation method using growth vitality assays and infection evaluation. EXPERIMENTAL METHODS 1) Ultraviolet irradiation experiment An ultraviolet irradiation experiment was conducted using Petri dishes. Journal of Water and Environment Technology, Vol.3, No.1, 2005 - 30 - The ultraviolet radiation sources used were a 20W low-pressure mercury lamp, a 330W medium- pressure mercury lamp, a 1kW medium-pressure mercury lamp, and a pulse lamp (at 500J x 2 pulses/second). A schematic diagram of the experimental apparatus is shown in Fig. 1. However, although not shown on Fig. 1, the 1kW medium-pressure mercury lamp alone was handled such that its irradiation unit was traversed using a belt conveyor running at constant speed without using a shutter. The procedure was then repeated. The experiment was conducted as follows: For runs 1 and 2 (Table 2), glass Petri dishes (5.7 mm dia.) were filled with 10 to 20 mL of a solution of C. parvum (10 6 oocysts/mL). For run 3 (Table 3), actual sludge present in city water was used and adjusted to a turbidity of about 250 NTU before being subjected to an irradiation experiment. In run 4 (Table 4), 20 mL of a solution of C. parvum (10 8 oocysts/mL) was applied to city water and then subjected to an experiment. UV Lamp Irradiation distance (L) Shutter Petri dish Magnetic spinbars Magnetic stirrer Fig. 1. Schematic diagram of the experimental apparatus Table 1. Experimental condition 1 (runs 1 and 2) RUN UV Lamp Experimental condition Irradiation distance (L) Low-pressure mercury lamp 20W 52mm １ Pulse lamp 500J×2 pulses/sec. 52mm Medium-pressure mercury lamp1 330W 97mm Medium-pressure mercury lamp2 1kW 17.5mm ２ Pulse lamp 500J×2 pulses/sec. 21.5mm Table 2. Experimental condition 2 (run 3) UV dose measurements RUN UV Lamp Experimental condition Irradiation distance (L) Not turbid Turbid Low-pressure mercury lamp 20W 17.5mm 0.55 mJ/cm 2 /sec. 0.4 mJ/cm 2 /sec. Medium- pressure mercury lamp2 1kW 17.5mm 2.0 mJ /cm 2 /pass 1.2 mJ/cm 2 /pass ３ Pulse lamp 500J×2 pulses/sec. 17.5mm 2.2 mJ/cm 2 /pulse 1.43 mJ/cm 2 /pulse Table 3. Experimental condition 3 (run 4) RUN UV Lamp Experimental condition Irradiation distance (L) ４ Pulse lamp 500J×2 pulses/sec. 21.5mm Journal of Water and Environment Technology, Vol.3, No.1, 2005 - 31 - The UVdoses of the lamps were measured using coliphage Qβ 1) . Tables 2 to 4 indicate the conditions for UV lamp irradiation in each experiment. In run 3, the cell culture method was conducted with turbidity present. *Beams from the 1kW medium -pressure mercury lamp were passed through 9 wire nets and reduced in output without any change in their wavelength spectra. 2)Inactivation evaluation The inactivation evaluation of C. parvum was conducted mainly using excystation (growth vitality assays). It was combined with excystation and the cell culture method in run 3, and the mouse infection and cell culture method (infectivity assays) in run 4. The testing methods used were as follows: - Excystation method: Samples were subjected to excystation culture and then observed with a microscope. - Cell culture method: HCT-8 cells were used and observed with a fluorescent microscope. - Mouse infection method: SCID mice (4 weeks old, female) were used. They were inspected using sucrose centrifugation suspension. EXPERIMENT RESULTS AND OBSERVATION Figs. 2 and 3 show the experiment results for runs 1 and 2, while figs. 4 and 5 show the experiment results on run 3 and fig. 6 shows the experiment results on run 4. 1) Comparison of the lamps using excystation evaluation Fig. 2 shows that the level of inactivity achieved with the low-pressure mercury lamp rose primarily corresponding to the UV dose delivered. However, the gradient of the change in inactivity varied in the case of the pulse lamps with 80 mJ/cm2 as the turning point. Fig. 3 demonstrates that the pulse lamps displayed superior ability in terms of inactivation than the 330W&1kW medium-pressure mercury lamps. The above findings revealed the pulse lamps to be superior in their ability to inactivate C. parvum by growth vitality assays. Fig. 2. Experiment results on run 1 Fig. 3. Experiment results on run 2 2) Comparison of the lamps in turbid water samples The inactivation effects of the lamps on turbid water with the excystation and the cell culture methods were compared. Fig. 4 shows that, concerning the inactivation effect with the excystation method, the low- and 1kW medium-pressure mercury lamps displayed trends that appeared to tail off, which was not the case with the pulse lamps. However, no clear difference was noted in the actual inactivation speed. -3 -2 -1 0 0 50 100 150 200 UV dose[mJ/cm 2 ] log10 (N/No) -2 -1.5 -1 -0.5 0 0 250 500 UV dose [mJ/cｍ 2 ] log10(N/N0) □ LPUV ● Pulse-Lamp ● Pulse-Lamp ■ 1kW-MPUV △ 330W-MPUV Journal of Water and Environment Technology, Vol.3, No.1, 2005 - 32 - Fig. 5 revealed that there was also no clear difference noted in the inactivation speed when the cell culture method was used between the low- and 1kW medium-pressure mercury lamps and pulse lamps. During the experiment evaluation, the cell culture method was highly uncertain (with a maximum accuracy of 95%), and a limit detection sensitivity of about 4 1ogs. UV dose [mJ/cm 2 ] UV dose [mJ/cm 2 ] Fig. 4. Inactivation effect of the Fig. 5. Inactivation effect of the lamps using the excystation method lamps using the cell culture method 3) Comparison of the evaluation methods used The experiment results (Fig. 6) show that the sensitivity to UV was highest when the cell culture method was used, followed by the mouse infection method. The excystation method was found to be extremely lacking in sensitivity to UV. CONCLUTION 1) Comparison of the measured UV doses demonstrated that, in terms of the ability to inactivate C. parvum, the pulse lamps performed to a higher level in the high-luminance region than the other lamps. 2) The sensitivity to ultraviolet rays was found to be in the following order (descending) cell culture＞mouse infection≫excystation. REFERENCES １）Kamiko N. Ohgaki S.,W.S.T.Vol.21,(.3)， Fig. 6. Run 4 (different pp.227-231(1989) evaluation methods used) ２）Clancy J.(2000).UV rises to the Cryptosporidium challenge, Water21,10,14-16 -6 -5 -4 -3 -2 -1 0 0204060 UV dose [mJ/cm 2 ] log10(Nt/N 0 ） ● LPUV □ Pulse-Lamp ▲ 1k W-MPUV ● LPUV □ Pulse-Lamp ▲ 1kW-MPUV ○ cell culture □ excystation △ mouse infection . Environment Technology, Vol .3, No.1, 2005 - 30 - The ultraviolet radiation sources used were a 20W low-pressure mercury lamp, a 33 0W medium- pressure mercury. Pulse-Lamp ● Pulse-Lamp ■ 1kW-MPUV △ 33 0W-MPUV Journal of Water and Environment Technology, Vol .3, No.1, 2005 - 32 - Fig. 5 revealed that there was also