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A Bacteriologic Evaluation of Some Simple Sterilizing Methods
A BACTERIOLOGIC EVALUATION OF SOME SIMPLE STERILIZING METHODS By H
enry
A . B a r t e l s , B .S ., D .D .S., and E
V
A R I O U S methods have been em ployed for the destruction o f micro organisms. T h e ideal method still is the use o f moist heat for suitable periods of time, either as steam under pressure (the autoclave) or boiling water (the instrument sterilizer). Th e clinical emergency m ay find such meth ods impractical, owing to the time factor involved, unless the dental office possesses a sufficient number of instruments and maintains them in an aseptic condition. T h e cleaning of root canals m ay be such an emergency. Prevention of extension of infection through the apex requires, in addition to a skilful technic, the em ployment of a number of previously sterilized files or the frequent resteriliza tion o f the several files in use. This study was undertaken to determine whether some simple methods advocated for sterilization possessed definite value. R e sults with Staphylococcus aureus indi cated no superiority o f one method over another. It was therefore decided to em ploy a more critical criterion, namely the destruction of B. subtilis spores, as a means o f determining the efficiency of ( i ) the flam e; (2) alcohol and the flam e; (3 ) formalin-alcohol and the flam e; (4) benzene, phenol, alcohol and boiling water, each used thirty seconds, and (5) the Flaherty molten metal sterilizer. From the Department of Oral Pathology, School of Dental and Oral Surgery, Columbia University. Read before the International Association for Dental Research, N ew York, N. Y ., M arch 14 , 194a. Jour. A .D .A ., V ol. 29, August 1, 19 42
l iz a b e t h
R
ic e ,
A.B., N ew York, N . Y .
METHOD
Suspensions of B. subtilis spores were prepared by emulsifying the growth of agar slants, several weeks old, in a sterile physiologic sodium chloride solution. The presence o f spores was determined by microscopic examination. Small iron screws were immersed in these suspen sions for at least ten minutes and then removed and allowed to dry about a half hour. O nly plain iron screws were used, since either cadmium coated or brass screws exert in themselves a bacterio static action. Screws were employed be cause of their low cost and relatively large surface area. Th e treated screws were then individ ually exposed to the sterilizing pro cedures mentioned. Subsequently, they were dropped, by means of sterile for ceps, into tubes of broth and incubated for forty-eight hours. Turbidity of the broth was no indication of growth since oxidation of the screws caused cloudiness of the broth. A ll broth tubes were there fore subcultured to agar plates. Tests were run on different days with different suspensions of B. subtilis spores. T h e five sterilizing methods were always tested in series at the same time so that all would be exposed to the same technical condi tions and each would serve as a control for’the other. Passage Through the Flam e.— Screws covered with B. subtilis spores were passed through the flame three, six, ten, fifteen and twenty times respectively. Since the beaks of the forceps cover a small area of the screw and possibly could protect some spores thereon, the 139 8
Bartels
and
R
ic e —
E
v a l u a t io n o f
beaks were always heated until they be came red before the screw was grasped and passed through the flame. Th e re sults are given in the accompanying table. Alcohol and- the Flam e.— Treated screws were immersed in 95 per cent ethyl alcohol and removed immediately, and the alcohol was ignited, the results are indicated in the table. Formalin, Alcohol and the Flam e.— Sanderson,1 in 1922, found that a m ix ture of one part formalin (40 per cent formaldehyde) and 3 parts 95 per cent
S t e r il iz in g M
1399
eth o d s
in tests with B. subtilis spores, as shown in the table. Flaherty M olten M etal Sterilizer.— Spore-covered screws were immersed in mineral oil and then submerged in molten alloy (melting point 1 7 5 0 C .) for periods of five or ten seconds. T h e min eral oil was employed only as an aid in preventing adherence o f the alloy to the screws. Tests were performed with the alloy heated to various temperatures ranging between 18 0 ° C. and 30 0 ° C., as shown in the table.
E ffec t of S im ple St e r iliz in g M ethods on
B.
S u btilts S po res
Results Method
Number Tests
Efficiency Per Cent
Growth
No Growth 0 0 0 1
17
5 85
Times Flamed
20
20 20
3 3 4 19 3
95 % alcohol and flame
30
27
3
10
40 % formalin, 1 part, 95 % alcohol, 3 parts
25
2
23
92
26
22
4
15.4
68
6
62
91.2
3 6 10
15
Benzene, phenol alcohol, boiling water, each 30 seconds Flaherty molten metal sterilizer
alcohol when ignited caused destruction of spore-bearing bacilli present on instru ments in more than 90 per cent of tests. Results previously reported by one of us,2 as well as our present results, corroborate Sanderson’s findings, as shown in the table. Benzene, Phenol, Alcohol and Boiling Water.— Treated screws were immersed for thirty seconds consecutively in each of the following solutions : benzene, con centrated phenol, 95 per cent alcohol and boiling water. This method is em ployed at a local hospital for sterilizing delicate instruments used in eye surgery. It was found b y us to be satisfactory in destroying non-spore formers such as Staphylococcus aureus, but of little value
3 3 4
0 0 0
c o n c l u s io n
Five simple methods of sterilization were evaluated. M oist heat is still con sidered the most efficient sterilizing agent. However, where conditions render its use impractical, we suggest the em ployment of either formalin-alcohol and the flame or the Flaherty molten metal sterilizer. b ib l io g r a p h y
1. S a n d e r s o n , E. A .: Sterilization of Sur gical Instruments. /. Lab. & Clin. M ed., 7 : 360, M arch 19 22. 2. B a r t e l s , H. A .: Relation of Bacteriology to Clinical Dentistry: Modern Methods of Sterilization in Dental Office /. D. Education, 5 : 3 1 2 , April 19 4 1.
enry
A . B a r t e l s , B .S ., D .D .S., and E
V
A R I O U S methods have been em ployed for the destruction o f micro organisms. T h e ideal method still is the use o f moist heat for suitable periods of time, either as steam under pressure (the autoclave) or boiling water (the instrument sterilizer). Th e clinical emergency m ay find such meth ods impractical, owing to the time factor involved, unless the dental office possesses a sufficient number of instruments and maintains them in an aseptic condition. T h e cleaning of root canals m ay be such an emergency. Prevention of extension of infection through the apex requires, in addition to a skilful technic, the em ployment of a number of previously sterilized files or the frequent resteriliza tion o f the several files in use. This study was undertaken to determine whether some simple methods advocated for sterilization possessed definite value. R e sults with Staphylococcus aureus indi cated no superiority o f one method over another. It was therefore decided to em ploy a more critical criterion, namely the destruction of B. subtilis spores, as a means o f determining the efficiency of ( i ) the flam e; (2) alcohol and the flam e; (3 ) formalin-alcohol and the flam e; (4) benzene, phenol, alcohol and boiling water, each used thirty seconds, and (5) the Flaherty molten metal sterilizer. From the Department of Oral Pathology, School of Dental and Oral Surgery, Columbia University. Read before the International Association for Dental Research, N ew York, N. Y ., M arch 14 , 194a. Jour. A .D .A ., V ol. 29, August 1, 19 42
l iz a b e t h
R
ic e ,
A.B., N ew York, N . Y .
METHOD
Suspensions of B. subtilis spores were prepared by emulsifying the growth of agar slants, several weeks old, in a sterile physiologic sodium chloride solution. The presence o f spores was determined by microscopic examination. Small iron screws were immersed in these suspen sions for at least ten minutes and then removed and allowed to dry about a half hour. O nly plain iron screws were used, since either cadmium coated or brass screws exert in themselves a bacterio static action. Screws were employed be cause of their low cost and relatively large surface area. Th e treated screws were then individ ually exposed to the sterilizing pro cedures mentioned. Subsequently, they were dropped, by means of sterile for ceps, into tubes of broth and incubated for forty-eight hours. Turbidity of the broth was no indication of growth since oxidation of the screws caused cloudiness of the broth. A ll broth tubes were there fore subcultured to agar plates. Tests were run on different days with different suspensions of B. subtilis spores. T h e five sterilizing methods were always tested in series at the same time so that all would be exposed to the same technical condi tions and each would serve as a control for’the other. Passage Through the Flam e.— Screws covered with B. subtilis spores were passed through the flame three, six, ten, fifteen and twenty times respectively. Since the beaks of the forceps cover a small area of the screw and possibly could protect some spores thereon, the 139 8
Bartels
and
R
ic e —
E
v a l u a t io n o f
beaks were always heated until they be came red before the screw was grasped and passed through the flame. Th e re sults are given in the accompanying table. Alcohol and- the Flam e.— Treated screws were immersed in 95 per cent ethyl alcohol and removed immediately, and the alcohol was ignited, the results are indicated in the table. Formalin, Alcohol and the Flam e.— Sanderson,1 in 1922, found that a m ix ture of one part formalin (40 per cent formaldehyde) and 3 parts 95 per cent
S t e r il iz in g M
1399
eth o d s
in tests with B. subtilis spores, as shown in the table. Flaherty M olten M etal Sterilizer.— Spore-covered screws were immersed in mineral oil and then submerged in molten alloy (melting point 1 7 5 0 C .) for periods of five or ten seconds. T h e min eral oil was employed only as an aid in preventing adherence o f the alloy to the screws. Tests were performed with the alloy heated to various temperatures ranging between 18 0 ° C. and 30 0 ° C., as shown in the table.
E ffec t of S im ple St e r iliz in g M ethods on
B.
S u btilts S po res
Results Method
Number Tests
Efficiency Per Cent
Growth
No Growth 0 0 0 1
17
5 85
Times Flamed
20
20 20
3 3 4 19 3
95 % alcohol and flame
30
27
3
10
40 % formalin, 1 part, 95 % alcohol, 3 parts
25
2
23
92
26
22
4
15.4
68
6
62
91.2
3 6 10
15
Benzene, phenol alcohol, boiling water, each 30 seconds Flaherty molten metal sterilizer
alcohol when ignited caused destruction of spore-bearing bacilli present on instru ments in more than 90 per cent of tests. Results previously reported by one of us,2 as well as our present results, corroborate Sanderson’s findings, as shown in the table. Benzene, Phenol, Alcohol and Boiling Water.— Treated screws were immersed for thirty seconds consecutively in each of the following solutions : benzene, con centrated phenol, 95 per cent alcohol and boiling water. This method is em ployed at a local hospital for sterilizing delicate instruments used in eye surgery. It was found b y us to be satisfactory in destroying non-spore formers such as Staphylococcus aureus, but of little value
3 3 4
0 0 0
c o n c l u s io n
Five simple methods of sterilization were evaluated. M oist heat is still con sidered the most efficient sterilizing agent. However, where conditions render its use impractical, we suggest the em ployment of either formalin-alcohol and the flame or the Flaherty molten metal sterilizer. b ib l io g r a p h y
1. S a n d e r s o n , E. A .: Sterilization of Sur gical Instruments. /. Lab. & Clin. M ed., 7 : 360, M arch 19 22. 2. B a r t e l s , H. A .: Relation of Bacteriology to Clinical Dentistry: Modern Methods of Sterilization in Dental Office /. D. Education, 5 : 3 1 2 , April 19 4 1.