19

THE APPLICATION OF DISINFECTION AND STERILIZATION TO INFECTIOUS

WASTE MANAGEMENT

Eugene C. Cole, Dr.P.H. School of Medicine

University of North Carolina, Chapel Hill, NC

The application of the principles of disinfection and sterilization

to effective infectious waste (IW) management must be viewed carefully.

While in general, both processes involve the inactivation of microbial

forms, the methods for achieving suitable disinfection and sterilization

for the on-site treatment of infectious wastes are very limited. I

might mention that on-site treatment has 3 potential advantages:

.assurance that wastes are properly treated, (2) minimization of

(1)

potential risk to personnel as material moves through the waste stream,

and (3) cost-effectiveness.

Disinfection

A disinfectant can be described as an agent, usually chemical, which

destroys disease or other harmful microorganisms except, ordinarily,

bacterial spores.

Disinfectants may inactivate cells in a variety of ways including cell

It refers to substances applied to inanimate objects.

wall and cytoplasmic membrane damage, electron transport interference,

and the coagulation of proteins and nucleic acids.

disinfection is normally a chemical process, it is not the only one.

While indeed,

Ultraviolet (W) radiation has long been popular for the inactivation of

airborne and surface microbes within the close vicinity of the

-. generating lamp. UV radiation however, provides poor penetrability and

is therefore not effective as a means of IW treatment.

20 I

Chemical disinfection is appropriate for the inactivation of liquid

wastes, such as cultures of etiologic agents, associated biologicals,

and human blood and blood products. It can also serve to decontaminate

some solid infectious wastes in a small clinic or office laboratory

where contaminated swabs, disposable culture loops, etc., are placed in

jars of disinfectant when steam sterilization or incineration are

unavailable.

The ideal disinfectant, in addition to being microbicidal, should

possess the characteristics listed in Table 1. Obviously, there is no

ideal disinfectant, so decisions must be made as to which factors are

most important in regard to the environment in question. Additionally,

in assessing the efficacy of a chemical disinfectant, one must consider

the important factors listed in Table 2.

When selecting a suitable disinfectant, consider first the type or

types or infectious agents that are of concern. Next, consider those

products with demonstrated efficacy against those agents.

becoming knowledgeable by reading and understanding product labels and

literature and consulting other appropriate references.

This warrants

Normally it is inadequate to pour liquid waste (other than very

small amounts) into a disinfectant solution. Preferably, an amount of

concentrated disinfectant is placed into an appropriate container so

that when the liquid waste is added, the final use-dilution will be that

which is recommended. Mixing may be required. Following approximate

“inactivation, or at the end of the day, the container is emptied into

2 1

Table 1. Characteristics of an Ideal Disinfectant

~~

Microbicidal

Easy to use

Detergent activity

Non - toxic

Non-irritating

Harmless to surfaces

Rapid action

Activity in presence of organic matter

Activity in presence of hard water

Stability

Residual activity

Inexpensive

22

Table 2. Factors Affecting Disinfectant Efficacy

~ ~~~~

Hydrogen in concentration

Concentration

Exposure time

Presence of interfering substances

Temperature

Numbers of microorganisms

Types of microorganisms

23

the sanitary sewer system (check local codes), and the system is flushed

with tap water to dilute the disinfectant and avoid damage to plumbing.

Solid waste items that have been decontaminated may then be regarded as

non-infectious trash and disposed of accordingly. One should always

remember that chemical disinfectants are toxic, and the use of proper

personal protective equipment is recommended.

Classes of Disinfectants

The following are the most commonly used classes of chemical

disinfectants :

A. Alcohols. (60-90%)

Advantages - bactericidal, tuberculocidal, virucidal (except isopropanol and against hydrophilic viruses), non-staining,

non-irritating, rapid action.

Disadvantanes - non-sporicidal, organic matter interference, incompatible with rubber and some plastics, highly flammable,

relatively expensive.

B. Quaternary Ammonium Compounds.

Advantanea - bactericidal (especially against gram-positive organisms), virucidal (against lipophilic viruses), fungicidal,

pleasant odor, inexpensive.

pi sadvantu - non-tuberculocidal, non-sporicidal, organic matter interference, non-virucidal (against hydrophilic

viruses).

24

C. Phenolics.

Advantapes - bactericidal, fungicidal, tuberculocidal, inexpensive.

Disadvantapes - questionable virucidal activity, non- sporicidal, toxic, skin irritant, unpleasant odor,

corrosiveness.

D . Iodophors.

Advantapes - bactericidal, virucidal, fungicidal, detergent action, storage stability.

Disadvantanes - prolonged exposure for tuberculocidal and sporicidal activity, corrosiveness, inactivation by organic

matter, relatively expensive.

E. Gluteraldehydes.

Advantaees - bactericidal, virucidal, fungicidal, tuberculocidal, sporicidal, lack or organic matter

interference, generally non-corrosive.

Disadvantaeeg - irritant, limited shelf life, expensive. F. Hypochlorites. (2 500 ppm free available chlorine)

Advantaees - bactericidal, virucidal, tuberculocidal, fungicidal, inexpensive.

D i s advantane s - non-sporicidal, toxic, corrosive, bleaching agents.

G . Hydrogen Peroxide. (2 3%)

Advantages - bactericidal, virucidal, tuberculocidal, fungicidal, sporicidal.

Disadvantaees - corrosive, expensive.

25

Chemical Inactivation of HIV (AIDS virus)

The AIDS virus has been found to be extremely susceptible to

chemical disinfection. Disinfectants used in lower than normal

concentrations and yet able to inactivate lo5 HIV during a 10 min

exposure at room temperature include: ethyl alcohol, isopropyl alcohol,

sodium hypochlorite ( 5 0 ppm), phenolics, and hydrogen peroxide.

Chemical Inactivation of HeDatitis B virus 6 High concentrations (10 ) of hepatitis B virus were found to be

inactivated within 10 min at 20C by sodium hypochlorite ( 5 5 0 ppm),

alkaline glutaralhdehyde (2%) , glutaraldehyde-phenate (0.13%/0.44%),

isopropyl alcohol (70%), and iodophor (80 ppm).

I

26

Sterilization

Sterilization is the act or process, physical or chemical, which

destroys all forms of life, especially microorganisms. Common

sterilization techniques include steam heat, dry heat, ethylene oxide,

and ionizing radiation. Of all the methods, heat, and particularly

moist heat, is the most reliable and widely used.

Ethylene oxide may present a carcinogenic, mutagenic, genotoxic,

reproductive, neurologic, and sensitization hazard to personnel and is

not recommended for IW treatment.

to solid infectious waste.

however, and energy requirements are extensive, dry heat treatment of IW

is not preferred. Ionizing radiation is an effective, low temperature

sterilization method that is used extensively for a wide range of

medical products.

large scale sterilization.

Drv heat inactivation may be applied

As sterilization times are prolonged,

Because of its high cost it is only suitable for

In considering all of the aforementioned sterilization methods,

steam sterilization is preferred.

the irreversible coagulation and denaturation of enzymes and structural

proteins. The basic principle of steam sterilization, as accomplished

in an autoclave, is to expose each item to direct steam contact at the

required temperature and pressure for the specified time. Thus, there

are four parameters of steam sterilization: pressure, temperature,

time, and steam. Recognized exposure periods for sterilization of clean

wrapped supplies (not infectious waste) are 30 min @ 121C in a gravity

displacement sterilizer, and 4 min @ 132C in a prevacuum unit.

Moist heat destroys microorganisms by

At

I

27

constant temperatures, sterilization times vary depending on the size

and type of load as well as the sterilizer type.

In the gravity displacement unit, steam is admitted to the top of

the chamber and because steam is lighter than air it forces air out the

bottom of the chamber through the drain vent. Such autoclaves are

primarily used to process culture media, water, pharmaceutical products,

infectious waste, and non-porous articles whose surfaces have direct

steam contact. For gravity displacement units, the penetration time is

prolonged because of incomplete air elimination. High speed prevacuum

sterilizers are similar to the gravity displacement type, except they

are fitted with a vacuum pump to insure air removal from the sterilizing

chamber and load before the steam is admitted. The advantage is nearly

instantaneous steam penetration, even into porous loads.

Autoclave monitoring is an essential part of the steam sterilization

process.

Periodic preventive maintenance should include calibration of gauges and

indicators. Biological indicators (using spores of Bacillus

stearothermoDhilus) should be run with actual loads on a daily or weekly

basis depending on frequency of use.

This includes in-use monitoring of temperature and pressure.

In 1982, Rutala et al. published data from a study of a gravity

displacement steam autoclave that was tested to determine the operating

parameters that affected sterilization of microbiological waste.

Commercially available 1.5 mil polyethylene biohazard bags were used.

They were tested in two modes:

of the bag folded down to expose the top layer of petri plates, and (2)

(1) in the open position, with the sides

28

with the opening in the bag loosely constricted with a twist tie. Four

holes were punched in the tips of all twist tied bags. Loads were

tested both with and without 500 ml of water added to the bags. 5, 10,

and 15 lb. loads of contaminated petri dishes were tested. They

contained 67, 136, and 205 plates, respectively. An average of 8 5 % of

the plates were contaminated with viable bacteria. The waste bags were

placed into shallow stainless steel (ss) or polypropylene (pp)

containers. The bags were monitored for time-temperature profiles by a

digital potentiometer, and for sterilization efficacy by a biological

indicator (spores of E. stearothermoDhilus) within the load. At the end

of the cycle, contents were sampled and cultured for viable microbes,

both aerobically and anaerobically.

StaDhvlococcus auxeus, StaDhvlococcus eDidermidis, Klebsiella

Bacteria included Escherichia coli,

pneumoniae, and species of Acinetobacter, Enterobacter, Pseudomonas,

Proteus, Streptococcus, and Bacillus.

When 5 lbs of microbiological waste in ss containers with or without

water, or pp containers with water was exposed to a steam sterilizing

cycle of 30 min, no growth of vegetative or sporeforming bacteria

occurred. In a pp container without water, all organisms were killed

after a 45 min cycle. When 10 lbs of microbiological waste was tested

in ss containers with water, 121C was reached in 45 min, and all

organisms except the indicator spores were killed. Without water at 45

min, all organisms with the exception of the indicator spores were

killed, but the temperature within the load did not reach 121C.

Utilizing the ss containers, either with or without water, the indicator

spores were not killed until a 90 min cycle was used. When the pp

containers were used, either with or without water, 121C could not be

29

reached and indicator spores survived even when a 90 min cycle was used.

All other organisms were killed after 45 min in the presence of water,

and after 60 min without water. The 15 lb load data were essentially

the same as for the 10 lb loads.

The investigators thus concluded that factors that facilitated heat

transfer and the sterilization of microbiological waste included the

type of container in which the waste was placed, the physical

characteristics of the load, and the autoclave bag. They also noted

that the bag closest to the door heated more slowly than the middle and

Pack bags and that the tops of bags must be adjusted to allow for the

free passage of air and steam.

necessity of using a cycle (90 min) that will kill all the spores of the

indicator, &. stearothermoDhilus. Since those spores are much more heat

resistant than the average organism it is unrealistic to require the

elimination of all spores in order to render waste "non-infectious".

Depending on the characteristics of the load, as already stated, spore

forming bacteria other then 8 . stearothermoDhiluS will be killed after

45 or 60 min.

The question was also raised as to the

The use of microwave oven ir radiation as a method for sterilizing

They undertook a bacterial waste was reported by Latimer and Matsen.

quantitative study to determine the effect of timed microwave

irradiation on commonly encountered laboratory bacteria, using an oven

operating at 2 , 4 5 0 MHz.

microwaves for 5 min included $. coli, E. plirabilis , E. aerueinosa, S .

marcescenq, S. Bureus , S. eDidermidig, and enterococcus. All organisms

Organisms grown in broth culture and exposed to

30

were killed within the 5 min period.

stearothermoDhilus spores were likewise exposed, with none surviving

after a 5 min exposure.

(about 100/load) exposed to the microwaves were rendered sterile within

5 min. The authors conclude that the utilization of microwave ovens for

bacterial decontamination in laboratories is entirely feasible. It

appears to be a practical time and energy saving method for the

treatment of bacterial waste. The treatment of fungal, viral, and

mycobacterial waste however, warrants additional investigation.

Spore strips containing viable E.

Loads of contaminated plastic petri dishes

Lastly, concern exists over the proper treatment of combined

'infectious/radioactive waste, Normally, the component representing the

greatest hazard is addressed first, with the final disposal of the

material subject to the regulations of the Nuclear Regulatory Commission

( N R C ) . If the waste is considered "highly infectious" and is

contaminated with low level radioisotopes, then extended autoclaving

followed by storage for decay, or approved incineration (for solids), or

autoclaving with release to the sanitary sewer (for liquids) may be

utilized.

result in the death of the infectious agent.

Control (CDC) recommends treating radioactive blood and urine by

chemical disinfection using sodium hypochlorite or hydrogen peroxide to

inactivate the biological component prior to approved disposal.

However, if chemical inactivation is not feasible, the waste should be

steam-sterilized, tagged non-infectious, and disposed of according to

the NRC.

In general, however, the time of storage for decay will

The Centers for Disease

3 1

References

1.

2.

3.

4.

5.

6.

7.

8.

9.

10.

11.

I' 12.

Rutala, W.A., M.M. Stiegel, and F.A. Sarubbi, Jr. 1982. Decontamination of laboratory microbiological waste by steam sterilization. Appl Envir Microbiol 43:1311-1316.

Latimer, J.M., and J.M. Matsen. 1977. Microwave oven irradiation as a method for bacterial decontamination in a clinical microbiology laboratory. J Clin Microbiol 6:340-342.

Gardner, J.F., and M.M. Peel. 1985. Introduction to sterilization and disinfection. Churchill Livingstone Inc., New York.

National Committee for Clinical Laboratory Standards. Clinical laboratory hazardous waste; proposed guideline. NCCLS document GP5-P. Villanova, Pennsylvania.

Martin, L.S., J . S . McDougal, and S.L. Loskoski. 1985. Disinfection and inactivation of the human T lymphotropic virus type III/lymphadenopathy-associated virus. J Infect Dis 152:400- 403.

Kobayashi, H., M. Tsuzuki, K. Koshimizu, H. Toyarma, N. Yoshihara, T. ShiKata, K. Abe, K. Mizuno, N. Otomo, and T. Oda. 1984. Susceptibility of hepatitis B virus to disinfectants or heat. J Clin Microbiol 20:214-216.

Bond, W.W., M.S. Favero, N.J. Petersen, and J.W. Ebert. 1983. Inactivation of hepatitis B virus by intermediate-to-high level disinfectant chemicals. J Clin Microbiol 18:535-538.

Songer, J.R. 1986. Decontamination--A probabilistic pursuit, p. 71-88. In Richardson, J.H., E. Schoenfeld, J.J. Tulis, and W.M. Wagner (eds.), Proceedings of the 1985 Institute on critical issues in health laboratory practice: health laboratory. E . I . duPont de Nemours & Co. , Wilmington, Delaware.

Safety management in the public

Wenzel, R.P., and D.H.M. Groschel. 1984. Sterilization, disinfection and disposal of hospital waste. In Mandell, G.L., R.G. Douglas Jr., and J .E . Bennett (eds.), Principles and practices of infectious disease, 2nd ed. John Wiley & Sons, New York.

Klein, M., and A. DeForest. 1963. The inactivation of viruses by germicides.

Collins, C.H., M.C. Allwood, S . F . Bloomfield, and A. Fox (eds.). 1981. Disinfectants: Their use and evaluation of effectiveness. Academic Press, London.

Chem Specialists Manuf Assoc Proc 49:116-118.

Block, S.S. (ed.). 1983. Disinfection, sterilization, and preservation, 3rd. ed. Lea & Febiger, Philadelphia.

32

13. Rutala, W.A., 1987. Disinfection, sterilization, and waste disposal. In Wenzel, R.P., Prevention and control of nosocomial infections. Williams & Wilkins, Baltimore.

14. Russell, A . D . , W.B. Hugo, and G.A.J. Ayliffe (eds.). 1982. Principles and practice of disinfection, preservation, and sterilization. Blackwell Scientific Publications, Boston.

15. The Environmental Protection Agency. 1986. EPA guide for infectious waste management. Service, Springfield, Virginia.

The National Technical Information