JOURNAL OF CLINICAL MICROBIOLOGY, Aug. 1976, P. 196-200 Copyright © 1976 American Society for Microbiology
Vol. 4, No. 2 Printed in U.S.A.
Phage Typing Set for E1, E2, E3, and E4 Salmonellae M. GERSHMAN Departments ofMicrobiology and Animal and Veterinary Sciences, University ofMaine, Orono, Maine 04473
Received for publication 7 May 1976
Eighteen common serotypes representative of group E,, E2, E3, and E4 Salmonella were characterized using a single set of phages.
Salmonellae are commonly implicated in cases of food poisoning and constitute one of the major problems confronting health authorities everywhere. An estimated two to four million people are infected annually in the United States alone (5). Outbreaks of salmonellosis, however, can be controlled, if not prevented, if the presence of the organism can be established. Many outbreaks of this disease have been resolved by locating the source of an infection by serologically identifying the species involved (2). However, if a particular species is fairly common and widely distributed, it becomes difficult, if not impossible, to assess its role in an outbreak. One of the most important properties of bacteriophage is its host specificity and its accompanying ability to distinguish types among apparently identical serotypes. Epidemiological use has been made of this property, and a number of commonly occurring Salmonella serotypes are now routinely characterized throughout the world by phage typing (1). In the course of our activities to develop phage typing sets for some of the more important salmonellae, it was observed that one single set of phages could be used to delineate a variety of Salmonella C, and C, serotypes (4). Given this revelation, we proceeded to develop a single set of phages for typing Salmonella serotypes representative of serological groups E1, E2, E3, and E4. Using a collection of Salmonella anatum, S. binza, and S. senftenberg phages developed in our laboratory (3; Gershman, J. Milk Food Technol., in press), we were able to type 412 of the 420 Salmonella El, E2, E3, and E4 cultures examined at random. The lytic pattern of these phages is noted in Table 1.
Isolates to be typed were lightly inoculated into 3 ml of nutrient broth and incubated at 37°C for 1.5 h or until turbidity was barely detectable. A small quantity of the broth culture was then flooded onto a nutrient agar plate, allowed to dry for approximately 15 min, and then spotted with phages by using a 1-ml syringe with a 26-gauge needle. All of the phages employed in this study were used at a routine test dilution of not less than 10-3. The plates were incubated overnight at 37°C and read the next day. The cultures were examined with the aid of an X10 aplantic hand lens and viewed through the bottom of the plate. Phage activity was recorded on the basis of the reactions described in Table 1. The serotypes used in this study and the ensuing patterns are listed in Table 2. The convenience inherent in this expanded typing set appears to be epidemiologically significant. Appreciation is expressed to Jacqueline Hunter for her most valuable laboratory assistance, and to Billie 0. Blackburn of the National Animal Disease Laboratory, Ames, Iowa, and L. Le Minor of the Pasteur Institute, Paris, France, for their assistance in securing cultures for this project.
LITERATURE CITED 1. Anderson, E. S. 1964. Phage typing ofSalmonella other than S. typhi, p. 89-110. In Van Oye (ed.), The world problem of salmonellosis. Junk, The Hague. 2. Gayler, G. E., R. A. MacCready, J. P. Reardon, and B. F. McKernan. 1955. An outbreak of Salmonellosis traced to watermelon. Public Health Rep. 70:311-313. 3. Gershman, M. 1974. A phage typing system for Salmonella anatum. Avian Dis. 18:565-568. 4. Gershman, M. 1976. Phage typing set for group C, and C2 Salmonellae. J. Clin. Microbiol. 3:214-217. 5. Okey, C. H. 1967. Salmonellosis. J. Maine Med. Assoc. 58:85-94.
196
NOTES
VOL. 4, 1976
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197
198
J. CLIN. MICROBIOL.
NOTES
TABLE 2. Reactions of test strains at routine test dilutions a Type strain
Salmonella group EI S. amsterdam S. amsterdam S. amsterdam S. amsterdam S. amsterdam S. amsterdam S. amsterdam S. amsterdam S. amsterdam S. amsterdam S. amsterdam S. amsterdam
S. anatum S. anatum S. anatum S. anatum S. anatum S. anatum S. anatum S. anatum S. anatum S. anatum S. anatum S. anatum S. anatum S. anatum S. anatum
Pattern
Culture no.
1
2 3 4 5 6 7 8 9 10 11
12 6 7 8 9 10 11
12 13 14 15 16 17 18 19 20
1/6/7/13/16 6/7/13/16 1/6/7/16 1/6/7/13/17/18 1/6/7/13 1/7/13 1/6/7/13/16/17/18 1/2/3/6/7/16/18 6/7/13 7/13 1/6/7/10/16 7/13/16/17
1/2/3/4/6/7/8/9/10/12/15/16/18 1
1/4 2/4/6 6/17/18 1/2/4/5/6/7/8/9/10/12/13/15/16/18 1/2/4/5 1/2/4/5/6/7/9/10/13/16/18 7/13 1/2/3/4/5/6/7/8/9/10/12/13/15/16/18 1/2/4/5/6/9/10/13/16/17/18 1/2/3/4/6/7/9/10/11/13/16/18 1/2/3/4/6/7/8/9/10/12/13/15/16/18 1/2/3/4/5/6/7/9/10/13/14/16/18 1/2/3/4/5/6/7/8/9/10/12/13/14/15/16/18
S. give S. give S. give S. give S. give S. give S. give S. give S. give
2 3 4 5 6 7 8 9
1/6/7/13 1/4/6/7/9/10/16 1/2/4/5/6/7/9/10/12/15/16/17/18 6/7/13 6/7/13/17/18 1/2/4/6/7/9/10/12/15/16/17/18 1/2/3/4/5/6/7/9/12/13/14/16/17/18 6/7 2/6/7/13
S. lexington
1
6
S. meleagridis
1
1/4/7/9/10/13/16
S. muenster muenster muenster muenster muenster muenster muenster muenster
1
S. S. S. S. S. S. S.
2 3 4 5 6 7 8
1/2/4/5/6/7/9/10/16 1/2/4/5/6/7/9/10/13/16 1/2/3/4/6/7/9/10/13/16/18 2/3/5/9/10/13/16 1/2/3/4/5/7/9/10/13/16 1/2/4/5/7/9/10/13/16 2/3/4/5/7/9/10/13/16 1/2/3/4/5/6/7/8/9/10/12/13/14/15
S. uganda
1
1/2/4/5/6/7/8/9/10/12/15/16/17/18
S. westhampton
1
16/18
1
NOTES
VOL. 4, 1976
TABLE 2 -Continued Type strain
Salmonella group E2 S. binza S. binza S. binza S. binza S. binza S. binza S. binza S. binza S. binza S. binza S. binza S. binza S. binza S. binza S. binza S. binza S. drypool S. drypool S. drypool S. drypool S. drypool S. drypool S. drypool S. drypool S. drypool S. drypool
Pattern
Culture no.
7 8 9 10 11
12 13 14 15 16 17 18 19
20 21 22 1
2 3 4 5 6 7 8 9 10
1/4/6/8/12 1/4/6/8/9/10 1/4/5/8/11/12 1/4/6/7/9/10/11 1/4/6/8/11 1/4/6/8 1/4/6/8/9/11 1/2/3/4/9/12 1/4/6/10/11 1/4/6/8/9/10/11 1/4/5/6/8/9/10/11/12/15/16 1/2/3/4/6/7/8/9/10/11/12/13/14/15/16 1/4/6/8/9/10/11/12/15/16 1/4/5/6/8/9/10/12/14/15/16 1/4/6/9/10/11/15/16 11
6/7/16 1/6/7/16 1/6/7/16/17 7 1/6/7 6/7/17 7/17 1/6/7/10/16 2/3/5/6/7 1/7/16
S. halmstad S. halmstad
2
1/18 1/12/15
S. manila
1
6
S. newington S. newington S. newington S. newington S. newington S. newington S. newington S. newington S. newington S. newington
1
1 2 3 4 5 6 7 8 9 10
1/4/6/7/8/9/10/16 4/7 1/6/7/10/18 1/4/6/7/9/10/16/18 1/4/6/7/8/9/10/16/17
1/4/5/6/7/9/10/16/17/18 1/4/6/9/10/16/18 1/4/5/6/7/9/10/14/16 1/6/7/17 1/4/6/9/10/16
Salmonella group E3 S. illinois S. illinois S. illinois S. illinois S. illinois S. illinois S. illinois
1 2 3 4 5 6 7
1/6 1/6/16 1/4/6/7/9/10/16 1/6/7/10/16 6
minneapolis minneapolis minneapolis minneapolis
1 2 3 4
1/4/6/7/8/9/10/12/15/16/18 6/7
S. S. S. S.
1/2/3/4/5/6/7/8/9/10/14/15/16 1/4/6/9/10/16
1/2/3/4/5/6/7/9/10/14/15/16/17/18 1/4/5/6/7/8/9/10/14/15/16/18
199
200
J. CLIN. MICROBIOL.
NOTES
TABLE 2-Continued Type strain
S. thomasville S. thomasville S. thomasville S. thomasville S. thomasville S. thomasville S. thomasville S. thomasville S. thomasville S. thomasville
Salmonella group E4 S. senftenberg S. senftenberg S. senftenberg S. senftenberg S. senftenberg S. senftenberg S. senftenberg S. senftenberg S. senftenberg
Culture no.
Pattern
1 2 3 4 5 6 7 8 9 10
1/2/3/4/5/6/8/9/10/12/15/16 1/2/4/5/6/7/8/9/10/11/12/13/14/15/16/18 6 6/7/11/16/17 1/4/6/11/16 1/4/5/6/8/9/10/11/14/15/16 1/4/5/9/10/11/14/15/16 1/6/11/16 1/4/6/7/9/10/11/15/16/17/18 1/6/11/14/15/16/17/18
7 8 9 10 11 12 13 14 15
1/6/17/18 1/6/13/16/17 1/6/13/15/16/17/18 1/6/16 1/6/13/16 1/13 1/6/13/15/17 6 1/6/8/13/15/16/17
S. taksony S. taksony S. taksony Only strong reactions (+ + + or above)
1 2 3
are recorded.
1/4/6/9/10 1/4/9/10/16 1/13
Vol. 4, No. 2 Printed in U.S.A.
Phage Typing Set for E1, E2, E3, and E4 Salmonellae M. GERSHMAN Departments ofMicrobiology and Animal and Veterinary Sciences, University ofMaine, Orono, Maine 04473
Received for publication 7 May 1976
Eighteen common serotypes representative of group E,, E2, E3, and E4 Salmonella were characterized using a single set of phages.
Salmonellae are commonly implicated in cases of food poisoning and constitute one of the major problems confronting health authorities everywhere. An estimated two to four million people are infected annually in the United States alone (5). Outbreaks of salmonellosis, however, can be controlled, if not prevented, if the presence of the organism can be established. Many outbreaks of this disease have been resolved by locating the source of an infection by serologically identifying the species involved (2). However, if a particular species is fairly common and widely distributed, it becomes difficult, if not impossible, to assess its role in an outbreak. One of the most important properties of bacteriophage is its host specificity and its accompanying ability to distinguish types among apparently identical serotypes. Epidemiological use has been made of this property, and a number of commonly occurring Salmonella serotypes are now routinely characterized throughout the world by phage typing (1). In the course of our activities to develop phage typing sets for some of the more important salmonellae, it was observed that one single set of phages could be used to delineate a variety of Salmonella C, and C, serotypes (4). Given this revelation, we proceeded to develop a single set of phages for typing Salmonella serotypes representative of serological groups E1, E2, E3, and E4. Using a collection of Salmonella anatum, S. binza, and S. senftenberg phages developed in our laboratory (3; Gershman, J. Milk Food Technol., in press), we were able to type 412 of the 420 Salmonella El, E2, E3, and E4 cultures examined at random. The lytic pattern of these phages is noted in Table 1.
Isolates to be typed were lightly inoculated into 3 ml of nutrient broth and incubated at 37°C for 1.5 h or until turbidity was barely detectable. A small quantity of the broth culture was then flooded onto a nutrient agar plate, allowed to dry for approximately 15 min, and then spotted with phages by using a 1-ml syringe with a 26-gauge needle. All of the phages employed in this study were used at a routine test dilution of not less than 10-3. The plates were incubated overnight at 37°C and read the next day. The cultures were examined with the aid of an X10 aplantic hand lens and viewed through the bottom of the plate. Phage activity was recorded on the basis of the reactions described in Table 1. The serotypes used in this study and the ensuing patterns are listed in Table 2. The convenience inherent in this expanded typing set appears to be epidemiologically significant. Appreciation is expressed to Jacqueline Hunter for her most valuable laboratory assistance, and to Billie 0. Blackburn of the National Animal Disease Laboratory, Ames, Iowa, and L. Le Minor of the Pasteur Institute, Paris, France, for their assistance in securing cultures for this project.
LITERATURE CITED 1. Anderson, E. S. 1964. Phage typing ofSalmonella other than S. typhi, p. 89-110. In Van Oye (ed.), The world problem of salmonellosis. Junk, The Hague. 2. Gayler, G. E., R. A. MacCready, J. P. Reardon, and B. F. McKernan. 1955. An outbreak of Salmonellosis traced to watermelon. Public Health Rep. 70:311-313. 3. Gershman, M. 1974. A phage typing system for Salmonella anatum. Avian Dis. 18:565-568. 4. Gershman, M. 1976. Phage typing set for group C, and C2 Salmonellae. J. Clin. Microbiol. 3:214-217. 5. Okey, C. H. 1967. Salmonellosis. J. Maine Med. Assoc. 58:85-94.
196
NOTES
VOL. 4, 1976
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197
198
J. CLIN. MICROBIOL.
NOTES
TABLE 2. Reactions of test strains at routine test dilutions a Type strain
Salmonella group EI S. amsterdam S. amsterdam S. amsterdam S. amsterdam S. amsterdam S. amsterdam S. amsterdam S. amsterdam S. amsterdam S. amsterdam S. amsterdam S. amsterdam
S. anatum S. anatum S. anatum S. anatum S. anatum S. anatum S. anatum S. anatum S. anatum S. anatum S. anatum S. anatum S. anatum S. anatum S. anatum
Pattern
Culture no.
1
2 3 4 5 6 7 8 9 10 11
12 6 7 8 9 10 11
12 13 14 15 16 17 18 19 20
1/6/7/13/16 6/7/13/16 1/6/7/16 1/6/7/13/17/18 1/6/7/13 1/7/13 1/6/7/13/16/17/18 1/2/3/6/7/16/18 6/7/13 7/13 1/6/7/10/16 7/13/16/17
1/2/3/4/6/7/8/9/10/12/15/16/18 1
1/4 2/4/6 6/17/18 1/2/4/5/6/7/8/9/10/12/13/15/16/18 1/2/4/5 1/2/4/5/6/7/9/10/13/16/18 7/13 1/2/3/4/5/6/7/8/9/10/12/13/15/16/18 1/2/4/5/6/9/10/13/16/17/18 1/2/3/4/6/7/9/10/11/13/16/18 1/2/3/4/6/7/8/9/10/12/13/15/16/18 1/2/3/4/5/6/7/9/10/13/14/16/18 1/2/3/4/5/6/7/8/9/10/12/13/14/15/16/18
S. give S. give S. give S. give S. give S. give S. give S. give S. give
2 3 4 5 6 7 8 9
1/6/7/13 1/4/6/7/9/10/16 1/2/4/5/6/7/9/10/12/15/16/17/18 6/7/13 6/7/13/17/18 1/2/4/6/7/9/10/12/15/16/17/18 1/2/3/4/5/6/7/9/12/13/14/16/17/18 6/7 2/6/7/13
S. lexington
1
6
S. meleagridis
1
1/4/7/9/10/13/16
S. muenster muenster muenster muenster muenster muenster muenster muenster
1
S. S. S. S. S. S. S.
2 3 4 5 6 7 8
1/2/4/5/6/7/9/10/16 1/2/4/5/6/7/9/10/13/16 1/2/3/4/6/7/9/10/13/16/18 2/3/5/9/10/13/16 1/2/3/4/5/7/9/10/13/16 1/2/4/5/7/9/10/13/16 2/3/4/5/7/9/10/13/16 1/2/3/4/5/6/7/8/9/10/12/13/14/15
S. uganda
1
1/2/4/5/6/7/8/9/10/12/15/16/17/18
S. westhampton
1
16/18
1
NOTES
VOL. 4, 1976
TABLE 2 -Continued Type strain
Salmonella group E2 S. binza S. binza S. binza S. binza S. binza S. binza S. binza S. binza S. binza S. binza S. binza S. binza S. binza S. binza S. binza S. binza S. drypool S. drypool S. drypool S. drypool S. drypool S. drypool S. drypool S. drypool S. drypool S. drypool
Pattern
Culture no.
7 8 9 10 11
12 13 14 15 16 17 18 19
20 21 22 1
2 3 4 5 6 7 8 9 10
1/4/6/8/12 1/4/6/8/9/10 1/4/5/8/11/12 1/4/6/7/9/10/11 1/4/6/8/11 1/4/6/8 1/4/6/8/9/11 1/2/3/4/9/12 1/4/6/10/11 1/4/6/8/9/10/11 1/4/5/6/8/9/10/11/12/15/16 1/2/3/4/6/7/8/9/10/11/12/13/14/15/16 1/4/6/8/9/10/11/12/15/16 1/4/5/6/8/9/10/12/14/15/16 1/4/6/9/10/11/15/16 11
6/7/16 1/6/7/16 1/6/7/16/17 7 1/6/7 6/7/17 7/17 1/6/7/10/16 2/3/5/6/7 1/7/16
S. halmstad S. halmstad
2
1/18 1/12/15
S. manila
1
6
S. newington S. newington S. newington S. newington S. newington S. newington S. newington S. newington S. newington S. newington
1
1 2 3 4 5 6 7 8 9 10
1/4/6/7/8/9/10/16 4/7 1/6/7/10/18 1/4/6/7/9/10/16/18 1/4/6/7/8/9/10/16/17
1/4/5/6/7/9/10/16/17/18 1/4/6/9/10/16/18 1/4/5/6/7/9/10/14/16 1/6/7/17 1/4/6/9/10/16
Salmonella group E3 S. illinois S. illinois S. illinois S. illinois S. illinois S. illinois S. illinois
1 2 3 4 5 6 7
1/6 1/6/16 1/4/6/7/9/10/16 1/6/7/10/16 6
minneapolis minneapolis minneapolis minneapolis
1 2 3 4
1/4/6/7/8/9/10/12/15/16/18 6/7
S. S. S. S.
1/2/3/4/5/6/7/8/9/10/14/15/16 1/4/6/9/10/16
1/2/3/4/5/6/7/9/10/14/15/16/17/18 1/4/5/6/7/8/9/10/14/15/16/18
199
200
J. CLIN. MICROBIOL.
NOTES
TABLE 2-Continued Type strain
S. thomasville S. thomasville S. thomasville S. thomasville S. thomasville S. thomasville S. thomasville S. thomasville S. thomasville S. thomasville
Salmonella group E4 S. senftenberg S. senftenberg S. senftenberg S. senftenberg S. senftenberg S. senftenberg S. senftenberg S. senftenberg S. senftenberg
Culture no.
Pattern
1 2 3 4 5 6 7 8 9 10
1/2/3/4/5/6/8/9/10/12/15/16 1/2/4/5/6/7/8/9/10/11/12/13/14/15/16/18 6 6/7/11/16/17 1/4/6/11/16 1/4/5/6/8/9/10/11/14/15/16 1/4/5/9/10/11/14/15/16 1/6/11/16 1/4/6/7/9/10/11/15/16/17/18 1/6/11/14/15/16/17/18
7 8 9 10 11 12 13 14 15
1/6/17/18 1/6/13/16/17 1/6/13/15/16/17/18 1/6/16 1/6/13/16 1/13 1/6/13/15/17 6 1/6/8/13/15/16/17
S. taksony S. taksony S. taksony Only strong reactions (+ + + or above)
1 2 3
are recorded.
1/4/6/9/10 1/4/9/10/16 1/13
