Use (Final Bleach Concentration)	Bleach Amount	Water Amount	Details	Susceptible Organisms (Concentration Needed to Kill)
Drinking water decontamination (50 ppm)^b	teaspoon or 3.75 mL—depending on clarity	1 gallon, or 3.79 liters	Let stand 30 min before use	Mycoplasma (25 ppm) Vegetative bacteria (<5 ppm)
Food handling equipment, bleach left on as residual (200 ppm)^c	1 tablespoon, or oz, or 15 mL	1 gallon, or 3.79 liters	Rinse surfaces after cleaning. Apply bleach and let sit for at least 5 min, then air dry	Bacillus spores (5 min, 100 ppm) Mycotic agents (1 hr, 100 ppm) 25 unenveloped viruses (10 min) Salmonella, Pseudomonas (10 min, 100 ppm)
Nonabsorbable surface disinfection (400 ppm)	2 tablespoons, or 1 oz, or 30 mL	1 gallon, or 3.79 liters	Rinse surfaces after cleaning. Apply bleach and let sit for at least 5 min, then air dry	Dermatophytes
Nonabsorbable surface disinfection (640 ppm)	1 cup or 8 oz or 237 mL	5 gallons, or 18.9 liters	Clean surface soap and water. Disinfect with bleach solution. Air dry.	Disinfect yeasts, (e.g., Candida [30 sec, 500 ppm])
Food or water utensils, bleach rinsed off after disinfection (1000 ppm)	5 tablespoons or 60 mL	1 cup or 237 mL	Wash with soap and water. Rinse, then sanitize with bleach solution. Air dry.	Mycobacterium tuberculosis Calicivirus or Norovirus (1 min)
Nonabsorbable surface disinfection (1750 ppm)	cup or 4 oz or 120 mL	1 gallon, or 3.79 liters	Wash with soap and water. Rinse, then sanitize with bleach solution. Air dry.	Unenveloped viruses, (e.g., parvoviruses, adenoviruses without organic debris [10 min]) Giardia cysts
Fungal growth on impervious surfaces (3200 ppm)	1 cup or 237 mL	1 gallon, or 3.79 liters	Wash with bleach. Scrub with brush. Rinse with water and air dry.	Clostridium difficile spores (10 min) Retard fungal growth
High-level disinfection in presence of organic debris or 10% plasma (5250 ppm, 10% solution)	1.5 cups or 25 tablespoons or 300 mL	1 gallon, or 3.79 liters	Clean surface, if possible, with water and soak organic debris. Apply bleach solution liberally.	Unenveloped viruses in organic debris (10 min)
Highest level disinfection (26,250 ppm)	1 gallon or 3.79 liters	1 gallon, or 3.79 liters	Wash and soak in bleach. Scrub with brush. Rinse with water and air dry.	Prions (1 hr)

1 ppm = 1 mg/L= 1 µg/mL; 1% bleach = ∼10,000 ppm solution; 0.1% bleach = ∼1000 ppm solution; 1 cup = 236.5 mL; 1 gallon = 3785.4 mL; 1 quart = 946.4 mL; 1 ounce = 30 mL; 1 tablespoon = 15 mL; 1 teaspoon = 5 mL.

Household bleach ranges in concentration from 5.25% to 6.15% sodium hypochlorite depending on the manufacturer. 5.25% bleach = 5.25 g/100mL = 52.5 g/L = ∼52,500 ppm solution. This concentration is used in dilutions noted in the table.

Bleach solutions stored at room temperature in opaque containers lose 50% activity over 1 month. Double strength concentrations should be used where diluted bleach solutions are stored before use. Use unscented 5.25% solution (household bleach). Never mix with other cleaners.

Wear protective rubber gloves and eye protection when using undiluted or handling >50 ppm solutions. Respiratory protection may be needed if fumes are detected.

d Can also use concentrated sodium hydroxide solutions.

^a Information adapted from Centers for Disease Control,54,55,55 and McGlynn W. Food Technology Fact Sheet, Oklahoma State University, www.fapc.okstate.edu. Accessed 2/6/11.

^b Minimum concentration of disinfection. For general washing, residual concentrations of 2–7 ppm are used. Municipal water systems have concentrations of 0.25–2 ppm.

^c Maximum concentration for food contact implements. Higher concentrations require rinsing off chlorine solutions before use. This concentration has been used for decontamination of fruits and vegetables for at least 1 minute, but must be washed off with potable water thereafter. Greater levels may cause bad odors or taste. Carcinogenic trihalomethanes form if higher concentrations are used.

Physical Agents

Heat

Use of either moist or dry heat is one of the oldest physical controls of microorganisms. Of the two, moist heat, especially under pressure, is more efficient, requiring shorter exposure at a lower temperature than is needed for disinfection by dry heat. When used correctly, steam under pressure is the most efficient means of achieving sterility. The recommended temperature-pressure-exposure time to produce sterilization with an autoclave is 121° C at 15 psi for 15 minutes or 126° C at 20 psi for 10 minutes. Steam heat is also most effective for eliminating resistant protozoal cysts, such as Toxoplasma and coccidia. Hot-air ovens are the most common dry-heat sterilizers, but to be effective, they must provide a consistent heat source. Dry-heat sterilization can be assumed if objects are maintained at 160° ± 10° C for a minimum of 1 hour but preferably 2 hours. Times of microwave oven sterilization of dry materials are similar to those of dry ovens once the sterilization temperature is reached. The only advantage of the microwave is the shorter time it takes to achieve these temperatures. Dry heat is recommended for sterilizing cutting instruments and glassware or items that might be damaged by moisture, such as glass syringes and reusable needles, but its use as a sterilant is generally not recommended because of the availability of other approaches and difficulty assessing adequacy of sterilization.

Radiation

Ionizing or high-energy radiation can be produced by radioactive elements, which are sources of γ-rays, or by a cathode-ray tube that produces x-rays. γ-Rays and x-ray radiation induce ionization of the vital cell components, especially nuclear DNA. Because of the cost and dangers of handling this equipment, this type of microbial control has found practical application chiefly in the industrial field. Pharmaceuticals, plastic disposables, and suture materials are generally sterilized by the manufacturer by means of ionizing radiation. Foodstuffs can be sterilized or disinfected of pathogenic microbes by using ionizing radiation. γ-Irradiation is used for sterilization of foods intended for specific-pathogen free or germ-free animals.⁵¹ Although bacterial sterilization can be achieved, vitamin A levels can be reduced and peroxide levels may increase. Pasteurization similarly reduced vitamin A levels but had no effect on peroxide concentration. Despite its safety, use of any irradiation method has met with resistance from the public. An unfounded misconception exists concerning residual radioactivity in treated foods. Nonionizing, or low-energy, radiation in the form of ultraviolet (UV) light has found practical application in the destruction of airborne organisms. Because low-energy rays do not penetrate well, they are used primarily as surface-active agents. The bactericidal range of UV light is 240 to 280 nm. UV lamps usually produce radiation in the range of 254 nm and work at maximal efficiency at temperatures of 27° C to 40° C. They depend on air convection currents to circulate airborne organisms. Germicidal lamps must be positioned above eye level to prevent retinal burns. For best efficiency, the lamps can also be placed in air conditioning or heating ducts. (See Airborne Contaminants, under Prevention of Nosocomial Infections.)

Chemical Agents

Biocides (germicides) denote chemical agents having antiseptic, disinfectant, or preservative properties. In addition to the environmental control of infection, many of these agents are used to preserve food, pharmaceuticals, and medical supplies. Typically, biocides lack selective toxicity to microorganisms. Viruses in a dried state are less susceptible to disinfectant, and their survival is enhanced by the presence of plasma protein.³⁶⁹ Moisturizing surfaces before application of disinfectants may improve their efficacy. The antimicrobial properties of various chemical disinfectants are summarized in Table 93-2. Table 93-3 describes the uses of the compounds to disinfect hospital equipment. It is difficult to achieve a high level of disinfection for thermometers. At exposure times of 10 to 12 hours to various chemicals, Salmonella has been isolated from contaminated thermometers.³⁹¹ Therefore, it is recommended that disposable thermometer covers be used.

TABLE 93-2

Antimicrobial Properties of Common Classes of Chemical Disinfectants

Class of Disinfectant	Bacteria				Fungi	Viruses
Class of Disinfectant	Gram Positives	Gram Negatives	Acid Fast	Spores	Fungi	Enveloped	Nonenveloped
ALCOHOLS
Ethyl	+	+	+	−	−	+	+/−
Isopropyl	+	+	+	−	−	+	−
HALOGENS
Chlorine (hypochlorite)	+	+	+	+	+	+	+
Iodine	+	+	+	±	+	+	±
Chlorine dioxide	+	+	+	+	+	+	+
ALDEHYDES
Formaldehyde	+	+	+	+	+	+	+
Glutaraldehyde	+	+	+	+	+	+	+
Ortho-phthalaldehyde	+	+	+	+	+	+	+
Phenolics	+	+	+	−	+	+	±
Potassium monopersulfate	+	+	+	+/−	+	+	+
Accelerated hydrogen peroxide	+	+	+	+	+	+	+/−
SURFACE-ACTIVE COMPOUNDS
Quats (cationic)	+	±	−	−	+	±	−
Amphoterics (anionic)	+	±	+	−	+	±	−
Biguanides	+	+	−	−	−	?	?
Ethylene oxide	+	+	+	+	+	+	+

quats, Quaternary ammonium compounds; +, effective; ±, somewhat effective; -, not effective; ?, effectiveness not known.

TABLE 93-3

Treatment Time Required for Chemical Disinfection of Hospital Equipment

Equipment	Type of Disinfection^a	Disinfectant^b	Exposure Time
Objects with smooth hard surfaces	H	17	3–12 hours
	H	18–10	15–18 hours
	H	11	5 hours
	I	1–3, 6, 8–10, 12	30 minutes
	L	1, 4, 7, 13, 14–16	10 minutes
	L	1, 2, 4, 14, 16	5 minutes
Rubber tubing (completely filled) and catheters	H	17	3–12 hours
	I	6, 10, 12	30 minutes
	L	7, 13, 14–16	10 minutes
Polyethylene tubing (completely filled) and catheters	H	17	3–12 hours
	H	8–10	15–18 hours
	I	1, 2, 6, 10, 12	30 minutes
Thermometers	L	1, 7, 13–16	10 minutes
	L	6	10 minutes
	I	2, 5, 10	30 minutes
Lens instruments	H	9–11	10–12 hours
	I	9, 10	30 minutes
	L	7, 13–16	10 minutes
Hinged instruments (free of organic material)	H	9, 10	10–12 hours
	I	2, 8–10	20 minutes
	I	6, 12	30 minutes
	L	8–10	10 minutes
	L	1, 7, 13–16	20 minutes
Inhalation, anesthetic, and endoscopic equipment	H	9, 11	10 hours
	H	17	3–12 hours
	I	2, 10	20 minutes
	L	1, 14–16	20 minutes
	L	10, 11	5 minutes
Housekeeping (floors, furnishings, and walls)	I	3, 6, 12	20 minutes
Housekeeping (floors, furnishings, and walls)	L	4, 7, 13–16	10 minutes

^a H, High-level disinfection (free of all microorganisms; equivalent to sterilization); I, intermediate-level disinfection (free of all vegetative bacteria, fungi bacilli, and most viruses); L, low-level disinfection (free of vegetative bacteria, fungi, and most enveloped viruses).

^b 1, 70%–90% ethyl or isopropyl alcohol; 2, 70%–90% ethyl alcohol; 3, hypochlorite (1000 ppm); 4, hypochlorite (100 ppm); 5, 0.2% iodine + alcohol; 6, iodophors (500 ppm); 7, iodophors (100 ppm); 8, 20% formalin + alcohol; 9, 20% formalin aqueous; 10, 2% activated glutaraldehyde aqueous; 11, 0.13% activated glutaraldehyde + phenate complex; 12, 2% phenolic aqueous; 13, 1% phenolic aqueous; 14, quaternary ammonium compounds; 15, amphoterics; 16, chlorhexidine; 17, ethylene oxide.

Effects of alcohol-containing disinfectants have been extensively studied for feline calicivirus (FCV), a surrogate for human norovirus disinfection. Ethanol (75% by volume), or a commercial hand gel containing at least 62% ethanol, was more effective in reducing the transfer rate of FCV by human hands than handwashing with water or no cleansing measures.²⁸ However, in other studies, the efficacy paralleled higher concentrations (99.5% best) of ethanol being the best and corresponding concentrations of isopropyl alcohol being less effective.²⁰¹ In the same study, hand sanitizers containing 60% ethanol or lower were much less effective. For these reasons, the authors of this chapter (CEG et al) recommend products with at least 70% ethanol. Alcohol-based sanitizers with added disinfectants have shown more efficacy.^196,215

Halogens

These compounds are ineffective or unstable in the presence of organic material, soap, or hard water. Halogens are active against a wide variety of viruses and resistant bacteria, such as Proteus and Pseudomonas. Table 93-4 summarizes the disinfectant activity of chlorine and iodine, which are described in greater detail next.

TABLE 93-4

Disinfectant Activity of Chlorine and Iodine 16,397

Chemicals	Concentration (mg/L)	Temperature (°C)	Time (min)
CHLORINE^a
Unenveloped virus	200	22	10
Gram-negative bacteria	0.1	5	0.16
Bacillus anthracis spores	2.3	22	60
Campylobacter	0.3	25	0.5
Giardia cyst	2.5	60	5
Cryptosporidium oocyst	10	20	720
IODINE
Unenveloped virus	0.5	5	30
Gram-negative bacteria	1.3	2–5	1
Giardia cyst	4	5	120

^a For further information on chlorine bleach, see Table 93-1.

Disinfection

Household bleach, a 5.25% to 6.15% sodium hypochlorite solution diluted to a minimum concentration of 50 ppm (vol/vol), is a common form of chlorine for disinfection of inanimate objects (see Table 93-1). Bleach is sold at a pH of 12 to prolong its shelf life. To increase its germicidal activity, especially against spores, the bleach is diluted with water to increase the available chlorine to change the pH of the solution to 7.0.³²⁰ Increasing the temperature of the solution decreases the exposure time needed. Precautions should be taken, because increasing temperature and decreasing pH of bleach can result in the release of chlorine gas, which is toxic. Chlorine bleach loses its effect in the presence of oil, dirt, or organic debris. Other than aldehydes, peroxymonosulfates, and chlorine dioxide, sodium hypochlorite solutions are one of the few chemicals that will inactivate parvoviruses²³¹ (see Chapters 8 and 9) and kill clostridial spores. For inanimate surfaces, sodium hypochlorite at a concentration of 1000 ppm for 1 minute was effective in inactivating FCV.³⁹⁶ Dilution and acidifying bleach solutions reduces their stability and increases their corrosiveness, and bleach is also deactivated by light. Therefore, bleach should be kept in opaque containers and diluted fresh daily. Because diluted bleach in opaque plastic can lose 50% of its activity over 1 month, tightly closed brown-colored bottles should be used after its dilution. Bleach is corrosive to metals and will discolor fabrics. Because it causes severe tissue damage, bleach solutions are not routinely used for antisepsis. It can be used for emergency antisepsis, but only in very dilute (less than 50 ppm) solutions.

Chlorine dioxide (Endimal, DuPont, USA), a halogen, is superior to chlorine in the destruction of bacteria, including spores and viruses.^321a It has high solubility and no odor, and it is unaffected by pH in the range of 4 to 10 and is nonreactive with ammonia compounds. Unlike bleach, chlorine dioxide is noncorrosive to metals even at high chlorine concentrations and is hypoallergenic and nontoxic to living tissues. It must be generated from on-site synthesis or can be ordered in a stabilized form. Chlorine dioxide has high penetration of environmentally resistant bacterial biofilms and has been used to treat air conditioning cooling systems to remove pathogenic bacteria.

Iodine is only slightly soluble in water; therefore, disinfectant solutions are made by dissolving it in alcohol or combining it with organic compounds. Iodine is sporicidal, fungicidal, protozoacidal, and somewhat virucidal, depending on exposure time and concentration of free iodine. Destruction of bacterial spores requires moist contact for more than 15 minutes. Unlike chlorine, iodine exerts its effect over a wide range of pH.

Iodophors are iodine solutions complexed with surfactants or polymers, which help increase the contact of the iodine with the surface to be disinfected while limiting the concentration of free iodine. Organic matter may reduce their activity, especially with dilute solutions, but the effect is less marked than it is with hypochlorites. Rinsing with alcohol will reduce these solutions’ residual antibacterial activity.

Antisepsis

Iodophors have an advantage over iodine because they are nonstaining and produce minimal tissue damage. Povidone-iodine is a complex of polyvinylpyrrolidone and iodine (Betadine solution, Purdue Frederick, Norwalk, CT). Such iodine compounds have been used for presurgical preparation, topical wound therapy, and joint or body cavity lavage. Solutions of 10% (undiluted) to 1% (1:9) povidone-iodine have been applied for skin and wound disinfection. Dilutions (1:4 to 1:100) of 10% stock solution result in increased bactericidal activity owing to increased concentrations of free iodine compared to the undiluted stock solution. A 1:50 dilution of povidone-iodine is recommended as an ocular surface disinfectant in presurgical situations.267,268 A 7.5% scrub containing an anionic detergent damages tissues and should be used only on intact skin. Bacterial concentrations are reduced on canine skin for up to 1 hour after scrub application.²⁶⁷ Polyhydroxydine is a potent iodine-containing wound and skin antiseptic (Xenodine, Squibb Animal Health Division, Princeton, NJ). It has been effective in treating canine wounds when used undiluted (100%) or as a 1:9 dilution (10%). As long as the iodophor solution maintains its color, it is effective. A concentration of 0.2% povidone-iodine was highly effective in killing greater than 99% of Mycobacterium tuberculosis isolates within 60 seconds.³⁰⁴

Systemic absorption of iodine may result in transient reduction in serum thyroxine or bicarbonate concentrations. Contact dermatitis that persists for several hours may occur in dogs.267–269,306 The skin irritation may lead to inactivation of the iodine through weeping proteins and increased postsurgical infection. Iodophors are also damaging to deeper tissue fibroblasts and must be diluted to 0.001% to be applied as wound or body cavity rinses.²⁰⁶ Concentrations of 0.5% to 1% have been effective but may be too strong for lavage of contaminated peritoneal cavities. Peritoneal lavage with 10% povidone-iodine can be fatal to dogs if 8 mL/kg is infused with intact peritoneum or 2 mL/kg with peritonitis. Concentrations of greater than 0.1% should not be used in joint irrigation.