Testing of Disinfectants - different methods

Testing of Disinfectants - different methods

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  Testing of Disinfectants Dr.Dhanya KC Associate Professor, Department of Microbiology St. Mary’s College (Autonomous), Thrissur- 680020, Kerala
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  • Disinfectants -must be periodically tested for efficacy • Disinfection Process Validation - establishing documented evidence that a disinfection process will consistently remove or inactivate known or possible pathogens from inanimate objects • Regulated by • U.S. Environmental Protection Agency – disinfectants • Food and Drug Administration - Agents used on humans and animals
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  Several methods oftesting disinfectants • Carrier test • Suspension test • Capacity test • Practical test • In-Use test
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  Carrier tests • Oldesttest - Described by Robert Koch in 1881 • Carrier such as a silk or catgut thread or a penicylinder (a little stick) is used • Carrier is contaminated by submersion into a liquid culture of test organism • The carrier will be dried and brought in contact with disinfectant to be tested • After exposure, carrier is cultured in a nutrient broth; • No growth - disinfectant activity of the • Growth - disinfectant not effective • Use-dilution test is example • Limitation of carrier tests • Number of bacteria dried on a carrier cannot be standardized • Survival of bacteria on carrier during drying is never constant
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  Suspension tests • Asample of bacterial culture is suspended in disinfectant solution • Cultured to identify whether inoculum is killed or not • Preferred - as bacteria are uniformly exposed to disinfectant • Different kinds of suspension tests • Qualitative suspension tests • Quantitative suspension tests • Phenol coefficient test
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  • Qualitative suspensiontest • Loopful of bacterial suspension will be brought into contact with disinfectant • Mixture cultured to analyze bacterial survival - Results - ‘growth’ or ‘no growth’ • Quantitative suspension test • Same above procedure, but number of surviving organisms are counted and compared to original inoculum size • Phenol coefficient test • Phenol coefficient - calculated by dividing dilution of test disinfectant by dilution of phenol that disinfects under predetermined conditions • Rideal Walker method • Chick Martin test • Garrod’s Test
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  Rideal Walker method •Phenol and test disinfectant diluted to different concentrations • Bactericidal activity determined against Salmonella typhi at various time intervals • Dilution of disinfectant which disinfects suspension in a given time is divided by that dilution of phenol which disinfects the suspension in same time - phenol coefficient • If effective dilution was 1:90 for phenol and 1:450 for disinfectant, Phenol coefficient - 5 Disadvantages • No organic matter is included • Microorganism Salmonella typhi may not be appropriate • Time allowed for disinfection is short • It should be used to evaluate phenolic type disinfectants only
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  Chick Martin test •Determines phenol coefficient of test disinfectant • 3% dried human feces - simulate presence of organic matter • Exposure time - 30 minutes • Test organisms – Salmonella typhi and Staphylococcus aureus Garrod’s Test • Modified Chick Martin Test • Organic material added is 5% of yeast suspension
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  Capacity tests • Abilityto retain activity in presence of increasing load of dirt and bacteria • Disinfectant is challenged repeatedly by successive additions of bacterial suspension • Best known capacity test - Kelsey-Sykes test • Triple challenge test - evaluate disinfectant - clean and dirty conditions • In hard water for testing under clean conditions • In yeast suspension for testing under dirty conditions • Test organism - S aureus, P aeruginosa, P vulgaris or E coli • Three successive additions of test bacteria, at 0, 10 and 20 minutes’ interval • Bacterial growth 8 minute after each challenge will be monitored in nutrient media • The disinfectant passes - no bacterial growth after first and second challenge
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  Stability Test • Stabilityupon storage determined- supplementary test • Pseudomonas aeruginosa • Disinfectant solution is prepared in two sets • First Set - inoculated with test organism and incubated for 7 days and tested for bacterial growth after 7 days • Second set - kept aside at room temperature for 7 days and then inoculated with test organism and tested for bacterial growth • If bacteria survive dilution of disinfectant tested, a higher concentration must be tested
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  Practical tests • Underreal-life conditions • Measure time-concentration relationship of disinfectant - quantitative suspension test Surface disinfection tests • Assess effectiveness of agent against surface-adhered microbes • Test surface contaminated with test bacteria and dried • Disinfectant is distributed over carrier • After exposure time number of survivors determined Surface Time kill Test • 24 hour culture is spread on sterile test surfaces • Treated with disinfectant for different time durations • Then test surfaces are placed in a solution that neutralizes disinfectant • Microorganisms surviving are cultured and enumerated • Untreated, inoculated test surfaces kept as control
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  In-use test • Todetect contamination of disinfectants • 1 ml disinfectant added to 9 ml diluent that contains inactivator • 10 drops of this diluted disinfectant placed on two nutrient agar plates • One incubated at 37o C for 3 days • Other incubated at room temperature for seven days • Five or more colonies on either plate indicate contamination of disinfectant
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  Testing scheme • Disinfectantexamined at three stages of testing • I - laboratory tests to verify antimicrobial activity - quantitative suspension tests • II - in laboratory using conditions simulating real-life conditions - practical tests • III - field tests and in-use tests • The antimicrobial activity is assessed in terms of activity towards • Vegetative Bacteria - Bactericidal Tests • Fungi And Yeasts - Fungicidal Tests • Mycobacteria - Tuberculocidal Tests • Viruses - Virucidal Tests • Bacterial Spores - Sporicidal Tests
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  Thank You….