| 1. ObjectUse both the Adenosine triphosphate (ATP) bioluminescence assay and the colony counting method to detected the different concentrations of 6 bacteria, and to evaluated the relationship between the two methods; Investigated the stability, repeatability, and the disinfectant infect one the ATP bioluminescence assay; Use ATP bioluminescence assay evaluated the effect of cleaning and disinfection, the results were used to support that ATP bioluminescence assay would be a rapid method to evaluate the effect of cleaning and disinfection.2. Method2.1 Use ATP test instrument and ATP kit to ration detected of 6 representative bacteria (Escherichia coli for intestinal infection, Staphylococcus aureus for pyogenic infection; Pseudomonas aeruginosa for nosocomial infection; Candida albicans for fungal infection; Mycobacterium chelonae subsp. Abscessus for M.tuberculosis and Bacillus Subtilis Var.niger Spore), use the colony counting method as parallel comparison, and described the linear correlation of the two methods in statistical chart.2.2 Use ATP bioluminescence assay to continuous tested one Escherichia coli sample for 3 days, the concentration of the sample was 1×101 cfu/ml-1×106 fu/ml detected by colony counting method. Use ATP bioluminescence assay to continuous tested one Escherichia coli sample for 3 days, the concentration of the sample was 1×105cfu/ml detected by colony counting method.Use one-way ANOVA to tested the relationship between ATP assay and colony counting method, Geehouse-Geisser and Huynh-Feldt used for adjusted, to detected the stability of ATP assay.2.3 According to neutralizer selecting test and suspension quantitative germicidal test to chose neutralizer for the 3 tested disinfectants (0.05% sodium hypochlorite,0.55% OPA, and 70% alcohol) and to evaluated the disinfection effect.2.4 Use ATP assay tested the Escherichia coli (ATCC8099) and ATP standard after their infected by 3 disinfectants for different times, to evaluated the disinfectants impact on the ATP assay.2.5 Use the ATP test instrument with sensitivity was 1×10-15mole and special sampling swab to evaluated the cleanness of environmental surfaces of the common hospital ward (III category of hospital environment) after varieties ways of cleaning/disinfection. Use the ATP test instrument with sensitivity was 1×10-18 mole to evaluated the disinfection effect of environmental surfaces of the common hospital ward (Ⅲ category of hospital environment).ATP results were compared with colony forming unit value (cfu, colony counting method) by Mann-Whitney test.2.6 Use ATP test instrument with the sensitivity was 1×10-18 mole to evaluated the cleanness of environmental surfaces after high-level disinfection, ATP results were compared with cfu value by chi-square test. The eligibility criteria was the results decreased 90% after disinfection.3. Result3.1 Compared ATP assay and colony counting method on tested of six bacteria suspensionFor Pseudomonas aeruginosa, Escherichia coli, and Staphylococcus aureus with the concentration of 1.0×101-1.0×106 cfu/ml, the log cfu and the log ATP had a linear correlation. The regression equation of Escherichia coli was Y=0.831X+0.3239 (R2=0.9261),of Staphylococcus aureus was Y=0.9983 X-0.0633 (R2=0.9913),and of Pseudomonas aeruginosa was Y=0.8736X-0.0741 (R2=0.9778).For Candida albicans with the concentration of 1.0×101-1.0×105 fu/ml, the log cfu and the log ATP had a linear correlation (Y=1.4867X-2.707, R2=0.926).For mycobacterium chelonae with the concentration of 1.0×102-1.0×107 cfu/ml, the log cfu and the log ATP had a linear correlation (Y=0.9639X+2.0683,R2=0.979). For Bacillus Subtilis Var.niger Spore with the concentration of 1.0×102-1.0×105 cfu/ml, the log cfu and the log ATP had a linear correlation(Y=1.9897X-0.9677, R2=0.9211).3.2 The stability of the ATP assay in tested of Escherichia coliUse ATP assay continuous tested different concentration of Escherichia coli samples for 3 days. Linear chart showed this method had a high stability. Continuous tested one Escherichia coli sample (the concentration was 1×105cfu/m) for 10 times by ATP assay, sphericity test P=0.896; after adjusted by Geehouse-Geisser, F=0.111, P=0.895; after adjusted by Huynh-Feldt, F=0.111, P=0.896.3.3 The results of neutralizer selecting testThe tested samples were infected by 0.05% sodium hypochlorite,0.55% OPA, and 70% alcohol for 1 minute,5 minutes, and 10 minutes. Suspension quantitative germicidal test showed that with the infected time increased, the log cfu and the long ATP had a linear correlation. The neutralizer can effective stop the disinfect, and had no influence on ATP method.3.4 The results of disinfectants effected on ATP standardThe log ATP value of the ATP standard was 5.95; Use 0.05% sodium hypochlorite infected the ATP standard for 1 minute,5 minutes,10 minutes and 30 minutes, the log ATP value were 5.72,5.32,5.26,5.24, and 5.18. Use 0.55% OPA infected the ATP standard for 1 minute,5 minutes,10 minutes and 30 minutes, log ATP value were 5.65, 5.61,5.53,5.57, and 5.48. Use 70% alcohol infected the ATP standard for 1 minute,5 minutes,10 minutes and 30 minutes, log ATP value were 5.71,5.77,5.60,5.64, and 5.61. With the infected time increased, the log ATP value of the ATP standard had some decreased.3.5 Compared the ATP assay of different sensitivity with colony counting method on evaluated the cleanness of hospital experimental surfaces after cleaning and/or disinfectionThe tested surfaces were cleaned by cotton cloth with tap water or disposable disinfection wet wipes with 0.3% compound quaternary ammonium salt, both the ATP assay and colony counting method showed that the results were higher in cotton cloth with tap water group, the two methods had a higher consistency. Forty samples were tested by ATP handset after cleaning by cotton cloth with tap water, the Md and IQR were 215RLU/100cm2 and 372RLU/100cm2, respectively. Forty samples were tested by colony counting method after cleaning by cotton cloth with tap water, the Md and IQR were 38cfu/cm2 and 117 cfu/cm2,respectively. Forty samples were tested by colony counting method after cleaning by disposable disinfection wet wipes with 0.3% compound quaternary ammonium salt, the Md and IQR were 14 cfu/cm2 and 71 cfu/ cm2, respectively. Twenty samples were compared the ATP and cfu value before and after cleaning by chlorinated disinfectant, the decreased Md and IQR of ATP were 170.5 (2amole/cm2) and 233.75 (2amole/cm2), of cfu were 140cfu/cm2 and 102 cfu/cm2, P=0.137.3.6 ATP instrument with high sensitivity used in external environment to evaluated the disinfection effectThe tests had been done in 5 different sites, and 15 samples were sampling before and after disinfection. Results of ATP assay and colony counting method were tested by chi-square test,P>0.05 (P=0.070).4. ConclusionColony counting method and ATP method had a higher correlation within bacterial concentration in certain limits; ATP method had a higher stability in tested the content of bacteria; The disinfectant had no influence on ATP standard; The ATP method would be a rapid tested method, to achieve the desired effect, it should be chose different sensitivity of ATP instrument according to purposes(for evaluate the cleaning effect or disinfection effect)... |
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