Journal of Antimicrobial Chemotherapy (1977) 3, 331-337
The in vitro activity of 15 penicillins and mecillinam against Neisseria gonorrhoeae
Barry A. Watts, Ian Phillips*
and Martin W. Stoate Department of Medical Electronics, St Thomas' Hospital, lx>ndon SE1 7EH, England
Minimum inhibitory concentrations (MIC's) of 15 penicillins and mecillinam were determined on solid media for 92 strains of Neisseria gonorrhoeae. Regression lines for MIC's of each antibiotic against those of benzylpenicillin were calculated. The order of activity against benzylpenicillin-insensitive strains was ampicillin, amoxycillin, azidocillin, ticarcillin carbenicillin, sulfocillin, phenoxymethylpenicillin, methicillin, phenethicillin, mecillinam, propicillin,flucloxacillin,nafcillin, cloxacillin and dicloxacillin. The regressions for flucloxacillin, cloxacillin and dicloxacillin were indistinguishable as were those of azidocillin, ticarcillin and carbenicillin. Only ampicillin and amoxycillin showed greater activity than benzylpenicillin against relatively resistant strains. Introduction The availability of several penicillins whose activity against Neisseria gonorrhoeae has not been fully assessed, such as dicloxacillin, flucloxacillin, propicillin, azidocillin (Sjoberg, Ekstrom & Forogren, 1968), sulfocillin (Tsuchiya, Oishi, Iwagishi & Iwahi, 1971), ticarcillin (Sutherland, Burnett & Rolinson, 1971), and the amidinopenicillin mecillinam (Lund & Tybring, 1972), prompted us to conduct this study of the activity of these against N. gonorrhoeae compared with that of benzylpenicillin, and to reassess the activity of ampicillin, phenethicillin, phenoxymethylpenicillin, nafcillin, methicillin and carbenicillin. Materials and methods Antibiotics The following antibiotics were provided as powders of stated potency. Benzylpenicillin sodium, amoxycillin trihydrate, ampicillin trihydrate, azidocillin potassium, carbenicillin *To whom reprint requests should be sent. 331
Downloaded from http://jac.oxfordjournals.org/ at Université Laval on June 28, 2015
Department of Microbiology, St Thomas's Hospital Medical School, London SE1 7EH, England
332
Antibiotic
B. A. Watts, I. Phillips and M. W. Stoate Side-chain structure
Benzylpenlcillin
Antibiotic
Side-chain structure
Corbenlclllin COONo
Dicloxocillin
Ticarcillin COONa
Cloxocillin
Flucloxacillin
Azidocillin
Vcc =/
II II
I
Sulfocillin SOjNa
Penicillin V
f>O-CH 2 -CO-
Propicillin Methicillin
Nafcillin
Ampicillln
Amoxycillin
Mecillinom NH2
Figure 1. Side chain structure of the penicillins used in this study.
sodium, cloxacillin sodium, flucloxacillin, mecillinam, methicillin sodium, phenoxymethylpenicillin potassium, phenethicillin potassium, propicillin potassium, ticarcillin. Sodium nafcillin and sulfocillin were provided as powders of unstated potency and assumed to be pure. Figure 1 shows the side chain structure of these antibiotics. Strains o/N. gonorrhoeae All strains of N. gonorrhoeae were isolated in the Venereal Diseases Department of St Thomas' Hospital on a selective medium containing vancomycin, colistin, nystatin and trimethoprim (Phillips, Humphrey, Middleton & Nicol, 1972). They were identified by their positive oxidase reaction and by characteristic staining with specific anti-gonococcal fluorescent antibody (Difco). Sugar fermentation reactions were determined for strains exhibiting weak fluorescence. A total of 92 strains was selected on the basis of the results of penicillin disc tests, so that roughly half of them would be relatively resistant to benzylpenicillin. Determination of minimum inhibitory concentrations (MIC's) The gonococci were maintained on lysed blood agar plates (Oxoid Columbia agar base, CM 331; plus 6 % saponin-lysed horse blood). A few colonies from these were suspended
Downloaded from http://jac.oxfordjournals.org/ at Université Laval on June 28, 2015
Phenethiciltln
\=/
Penicillins and N. gonorrhoeae
333
in 2-5 ml volumes of nutrient broth (Southern Group Laboratories) plus 6% lysed horse blood, and incubated at 37°C in 10% CO 2 for about 6 h. These broths were inoculated with a multiple inoculator, to deliver approximately 104 colony forming units (cfu) on Diagnostic Sensitivity Test agar (Oxoid CM 261) containing 6% lysed horse blood and antibiotics in suitable concentrations. The Oxford Staphylococcus was included as a Table I. Distribution of sensitivity for 92 strains of Neisseria gonorrhoeae against 15 penicillins and mecillinam Number of strains with MIC (mg/1) of: 1 2 4 0008 0016 0 0 3 0 0 6 012: 5 0-25 0-5 Benzylpenicillin
11
4
21 28
25 33 19 10
4
17* 10 16* 10 4 4 2 0 20* 8 3 21* 7 21* 42
2* 3* 1 3 25
11
10 27 4* 37 14
16 13 10 4 0
5 12* 15 6 25 0 1
14
16
12 28 5 28 12 10 17* 9 1 21 7 18 6 14 5 28 8* 0 16 20 7 6 8 4 2 8 15* 2 12* 14
16
32
8 16 3 15
8 10 7 3*
3 5 3
13 10
9
4
4
18 3 20 2 1 3 19 1 5 6 2 3 26 11 9 21 12 7 18 1 7 10 5 8 13 11 16 4 13 8 9 30 1 21
12
*MIC for Oxford Staphylococcus. Table n . Regression coefficients, slopes, intercepts and variance of slopes and intercepts of regression line depicted in Figure 2 Antibiotic Nafcillin Dicloxacillin Cloxacillin Flucloxacillin Propicillin Phenethicillin Methicillin Penicillin V Sulfocillin Azidocillin Carbenicillin Ticaicillin Amoxycillin Ampicillin Mecillinam
Regression coefficient
b
Variance ofi
c (intercept)
Variance
(slope)
0-88 0-90 0-88 0-88 0-88 0-93 0-94 0-94 0-93 0-95 0-94 0-94 0-88 0-92 0-94
0-814 0-727 0-72 0-751 0-787 0-805 0-881 0-859 0-916 0-895 0-839 0-850 1-403 1-528 1-29
00021 00013 00017 00019 00021 00011 00011 00011 00014 00011 00010 0-0011 00066 00051 00023
-2-935 -2-789 -2-598 -2-62 -213 -1-885 -1-73 -1-328 -1078 -0-789 -0-847 -0-821 0-378 0-994 -3-412
00067 00054* 00069* 00068* 00080 00053 00047 00061 00068 00081** 0-0086* * 00091** 0-039 0035 00039
• and • * regressions not statistically different.
ofc
Downloaded from http://jac.oxfordjournals.org/ at Université Laval on June 28, 2015
Amoxycillin Ampicillin 1* Azidocillin 4 Carbenicillin Cloxacillin Dicloxacillin Flucloxacillin Mecillinam Methicillin Nafcillin Phenethicillin Phenoxymethyl penicillin Propicillin Sulfocillin Ticarcillin 5
28*
8
334
B. A. Watts, I. Phillips and M. W. Stoate
control. The plates were read after overnight incubation at 37°C in 10% CO2. The MIC was taken as the concentration of antibiotic that completely inhibited growth of the organism. MIC's of each antibiotic were compared with those of benzylpenicillin for each strain, and regression lines drawn by the method of least square analysis. Statistical computations were carried out on a Varian 620 computer (Statistical appendix). A B
2
A Ampicillin B Amoxycillin I C Sulfocillin D Phenoxymethylpenicillin E Methicillin 05 F Meciilinam G Nafcillin _ BP Benzylpenicillin
BP
C
D
E F
025
2
0062 0031 0016 0008 0004
BP
0001
DC EAB
001
G
F 01 I Antibiotic MIC's (mg/l)
10
BP
o»
E
2 " A B 1. C D E 05 - F G H 0-25 BP
Azidocillin Ticorcillin Corbenicillin Phenethicillin Propicillin Flucloxacillin Cloxacillin Dicloxocillin Benzylpenicillin
/
ABC
D
J^
/ /
/
jf
' ' '/ / / / / / / / / / / / /
/ / / /
to
0 125
5
nzylperilcillir
100
///y ///y
E
FG H
/ /
/ / / /
/
/ ,///*/ / / /
0062 0031 -
w
o
0016 0008 n.nni 0 001 BCA
0 EGFH 001
01
10
100
Antibiotic MIC's (mg/l)
Figure 2. Regression lines for MIC's of benzylpenicillin against other penicillins and meciilinam.
Downloaded from http://jac.oxfordjournals.org/ at Université Laval on June 28, 2015
0125
Penicillins and N. gonorrhoeae
335
Results The distribution of MIC's obtained for the antibiotics, listed after benzylpenicillin in alphabetical order, is shown in Table I. Of the 92 strains studied 45 were relatively resistant to benzylpenicillin (MIC JJO-1 25 mg/1). Only four strains had MIC's as high as 1 mg/1. Regression lines of benzylpenicillin MIC's against MIC's of the other antibiotics are shown in Figure 2. Table II gives the regression coefficients, slopes and intercepts for these lines. From statistical analysis of these slopes and intercepts two groups were identified in which the antibiotics could not be considered significantly different. These were first, cloxacillin, dicloxacillin and flucloxacillin, and second, carbenicillin, ticarcillin and azidocillin. All other regressions differed significantly in slope, intercept or both. Discussion Downloaded from http://jac.oxfordjournals.org/ at Université Laval on June 28, 2015
Our results confirm that there are considerable differences in the inhibitory activity on N. gonorrhoeae of the various penicillins. From the regression lines, strains inhibited by benzylpenicillin 1 mg/1 would have the following theoretical MIC's to the other penicillins: ampicillin 0-5 mg/1; amoxycillin 1 mg/1; azidocillin, ticarcillin, carbenicillin 2 mg/1; sulfocillin, phenoxymethyl-penicillin 4 mg/1; methicillin, phenethicillin 8 mg/1; propicillin 16 mg/1; flucloxacillin, nafcillin, cloxacillin, dicloxacillin 32 mg/1. Mecillinam would have a MIC of 16 mg/1. The slopes of most of the regression lines were very similar, being slightly less than one, indicating that the antibiotics were relatively less active than benzylpenicillin against penicillin-resistant strains compared with fully sensitive strains. The exceptions to this were ampicillin, amoxycillin and mecillinam which had slopes greater than one and so were relatively more active against penicillin-resistant than against sensitive strains. Penicillins of possible clinical importance are those that show in vitro effectiveness against strains of N. gonorrhoeae that are relatively insensitive to benzylpenicillin, as failures of treatment are likely to occur with strains such as these. Our results, in agreement with those of 0degaard (1962) and Reyn & Bentzon (1968), showed that only ampicillin and amoxycillin are more active weight-for-weight than benzylpenicillin against penicillin insensitive strains. Unlike Martin, Lester, Kellogg & Thayer (1969) we did not find carbenicillin to have greater activity against penicillin insensitive strains. Carbenicillin, azidocillin and ticarcillin all showed equal activity to that of benzylpenicillin against sensitive strains but were less active against insensitive strains. Like Thayer & Axnick (1963) and Reyn & Bentzon (1968) respectively we found nafcillin and methicillin relatively ineffective. Similarly, the activity of the cloxacillin group was similar to that reported for oxacillin by these two groups of workers. We have been able to find only one report of sulfocillin MIC's for gonococci—one strain with an MIC of 0-39 mg/1, which resembles our relatively resistant strains. Finally our results for phenoxymethyl- and phenoxyethylpenicillin were very similar to those reported by Thayer and Axnick (1963). Chemotherapeutic effectiveness depends on many properties of antibiotics besides their in vitro effectiveness, but clinical trials have already shown the usefulness of ampicillin and amoxycillin (Morton, 1975), the two penicillins in this study to show good activity. Trials have also shown phenoxymethyl penicillin (Love & Weir, 1956; Shiel, 1956; Willcox, 1964), phenethicillin (Hilton, 1961; Allison, 1962; Willcox, 1964), and cloxacillin (Willcox, 1964) to be relatively ineffective. There would therefore appear to be
336
B. A. Watts, I. Phillips and M. W. Stoate
a reasonable relationship between MIC's and clinical effectiveness and on this basis ticarcillin, carbenicillin, azidocillin and sulfocillin might be worthy of clinical trials. Acknowledgements We thank the following pharmaceutical companies for their supplies of antibiotics, Beecham Research Laboratories (amoxycillin, ampicillin, azidocillin, benzylpenicillin, carbenicillin, cloxacillin, dicloxacillin, flucloxacillin, methicillin, phenethicillin, propicillin and ticarcillin). Wyeth Laboratories Inc. for nafcillin; Leo Laboratories Ltd for mecillinam; Lilly Research Centre Ltd for phenoxymethyl penicillin and Takeda Chemical Industries Ltd for sulfocillin. We also thank Mr T. Cowell, Head of the Department of Medical Electronics, St Thomas' Hospital for his assistance. This work was made possible by a DHSS grant.
Lund, F. & Tybring, L. 6P-Amidino-penicillanic acids—a new group of antibiotics. Nature, New Biology 236: 135-6 (1972). Martin, J. E., Lester, A., Kellogg, D. S. & Thayer, J. D. In vitro susceptibility of Neisseria gonorrhotae to nine antimicrobial agents. Applied Microbiology 18: 21-3 (1969). Morton, R. S. In Recent Advances in Sexually Transmitted Disease, Vol. 1, (R. S. Morton and J. R. W. Harris, Eds). Churchill Livingstone, Edinburgh, London and New York (1975. pp. 69-87. 0degaard, K. Penicillins in gonorrhoea. British Medical Journal i: 645 (1962). Phillips, I., Humphrey, D., Middleton, A. & Nicol, C. Diagnosis of gonorrhoea by culture on a selective medium containing vancomycin, colistin, nystatin and trimethoprim (VCNT): a comparison with Gram-staining and immunofluorescence. British Journal of Venereal Diseases 4: 287-92 (1972). Phillips, I., King, A., Warren, C , Watts, B. & Stoate, M. W. The activity of penicillin and eight cephalosporins on Neisseria gonorrhoeae. Journal of Antimicrobial Chemotherapy 2: 31-9 (1976). Reyn, A. & Bentzon, M. W. A study of the relationships between the sensitivities of Neisseria gonorrhoeae to sodium penicillin G, four semisynthetic penicillins, spiramycin and fusidic acid. British Journal of Venereal Diseases 44: 140-150 (1968). Sjoberg, B., Ekstrdm, B. & Forsgren, U. a-Azidobenzylpenicillin 1. Chemistry, bacteriology and experimental chemotherapy. Antimicrobial Agents and Chemotherapy—1967. 560-7 (1968). Sutherland, R., Burnett, J. & Rolinson, G. N. a-Carboxy-3-thienyl-methylpenicillin. Antimicrobial Agents and Chemotherapy—1970: 390-395 (1971). Thayer, J. D. and Axnick, N. W. Susceptibility of gonococci to ten penicillins. Antimicrobial Agents and Chemotherapy—1963: 427-9 (1964). Tsuchiya, K., Oishi, T., Iwagishi, C. & Iwahi, T. In vitro antibacterial activity of disodium asulfobenzylpenicillin. Journal of Antibiotics 24: 607-19 (1971). Willcox, R. R. The newer penicillins in the treatment of gonorrhoea. Postgraduate Medical Journal 40 (Suppl.): 202-7 (1964). Statistical appendix M. W. Stoate Firstly an analysis of variance (Kempthorne, 1952) was performed on the MIC results for the 16 antibiotics. This showed that there was a significant difference between the antibiotics at the 0-1 % level: thus all antibiotics could not be considered to be the same.
Downloaded from http://jac.oxfordjournals.org/ at Université Laval on June 28, 2015
References
Penicillins and N. gonorrhoeae
337
Regression analysis was then performed on the data, by the method previously described (Phillips, King, Warren, Watts & Stoate, 1976) to test these differences. T-tests (Beaumont, 1972) were also performed on the population means and these confirmed the results obtained by the above method. References Beaumont, G. P. Elementary Mathematical Statistics. McGraw Hill, New York (1972), pp. 224-227. Kempthorne, O. The Design and Analysis of Experiments. Wiley, New York (1952), pp. 32-33, Phillips, I., King, A., Warren, C , Watts, B. & Stoate, M. W. The activity of penicillin and eight cephalosporins on Neisseria gonorrhoeae. Journal of Antimicrobial Chemotherapy 2: 31-9 (1976). Downloaded from http://jac.oxfordjournals.org/ at Université Laval on June 28, 2015
(Manuscript accepted 22 October, 1976)
The in vitro activity of 15 penicillins and mecillinam against Neisseria gonorrhoeae
Barry A. Watts, Ian Phillips*
and Martin W. Stoate Department of Medical Electronics, St Thomas' Hospital, lx>ndon SE1 7EH, England
Minimum inhibitory concentrations (MIC's) of 15 penicillins and mecillinam were determined on solid media for 92 strains of Neisseria gonorrhoeae. Regression lines for MIC's of each antibiotic against those of benzylpenicillin were calculated. The order of activity against benzylpenicillin-insensitive strains was ampicillin, amoxycillin, azidocillin, ticarcillin carbenicillin, sulfocillin, phenoxymethylpenicillin, methicillin, phenethicillin, mecillinam, propicillin,flucloxacillin,nafcillin, cloxacillin and dicloxacillin. The regressions for flucloxacillin, cloxacillin and dicloxacillin were indistinguishable as were those of azidocillin, ticarcillin and carbenicillin. Only ampicillin and amoxycillin showed greater activity than benzylpenicillin against relatively resistant strains. Introduction The availability of several penicillins whose activity against Neisseria gonorrhoeae has not been fully assessed, such as dicloxacillin, flucloxacillin, propicillin, azidocillin (Sjoberg, Ekstrom & Forogren, 1968), sulfocillin (Tsuchiya, Oishi, Iwagishi & Iwahi, 1971), ticarcillin (Sutherland, Burnett & Rolinson, 1971), and the amidinopenicillin mecillinam (Lund & Tybring, 1972), prompted us to conduct this study of the activity of these against N. gonorrhoeae compared with that of benzylpenicillin, and to reassess the activity of ampicillin, phenethicillin, phenoxymethylpenicillin, nafcillin, methicillin and carbenicillin. Materials and methods Antibiotics The following antibiotics were provided as powders of stated potency. Benzylpenicillin sodium, amoxycillin trihydrate, ampicillin trihydrate, azidocillin potassium, carbenicillin *To whom reprint requests should be sent. 331
Downloaded from http://jac.oxfordjournals.org/ at Université Laval on June 28, 2015
Department of Microbiology, St Thomas's Hospital Medical School, London SE1 7EH, England
332
Antibiotic
B. A. Watts, I. Phillips and M. W. Stoate Side-chain structure
Benzylpenlcillin
Antibiotic
Side-chain structure
Corbenlclllin COONo
Dicloxocillin
Ticarcillin COONa
Cloxocillin
Flucloxacillin
Azidocillin
Vcc =/
II II
I
Sulfocillin SOjNa
Penicillin V
f>O-CH 2 -CO-
Propicillin Methicillin
Nafcillin
Ampicillln
Amoxycillin
Mecillinom NH2
Figure 1. Side chain structure of the penicillins used in this study.
sodium, cloxacillin sodium, flucloxacillin, mecillinam, methicillin sodium, phenoxymethylpenicillin potassium, phenethicillin potassium, propicillin potassium, ticarcillin. Sodium nafcillin and sulfocillin were provided as powders of unstated potency and assumed to be pure. Figure 1 shows the side chain structure of these antibiotics. Strains o/N. gonorrhoeae All strains of N. gonorrhoeae were isolated in the Venereal Diseases Department of St Thomas' Hospital on a selective medium containing vancomycin, colistin, nystatin and trimethoprim (Phillips, Humphrey, Middleton & Nicol, 1972). They were identified by their positive oxidase reaction and by characteristic staining with specific anti-gonococcal fluorescent antibody (Difco). Sugar fermentation reactions were determined for strains exhibiting weak fluorescence. A total of 92 strains was selected on the basis of the results of penicillin disc tests, so that roughly half of them would be relatively resistant to benzylpenicillin. Determination of minimum inhibitory concentrations (MIC's) The gonococci were maintained on lysed blood agar plates (Oxoid Columbia agar base, CM 331; plus 6 % saponin-lysed horse blood). A few colonies from these were suspended
Downloaded from http://jac.oxfordjournals.org/ at Université Laval on June 28, 2015
Phenethiciltln
\=/
Penicillins and N. gonorrhoeae
333
in 2-5 ml volumes of nutrient broth (Southern Group Laboratories) plus 6% lysed horse blood, and incubated at 37°C in 10% CO 2 for about 6 h. These broths were inoculated with a multiple inoculator, to deliver approximately 104 colony forming units (cfu) on Diagnostic Sensitivity Test agar (Oxoid CM 261) containing 6% lysed horse blood and antibiotics in suitable concentrations. The Oxford Staphylococcus was included as a Table I. Distribution of sensitivity for 92 strains of Neisseria gonorrhoeae against 15 penicillins and mecillinam Number of strains with MIC (mg/1) of: 1 2 4 0008 0016 0 0 3 0 0 6 012: 5 0-25 0-5 Benzylpenicillin
11
4
21 28
25 33 19 10
4
17* 10 16* 10 4 4 2 0 20* 8 3 21* 7 21* 42
2* 3* 1 3 25
11
10 27 4* 37 14
16 13 10 4 0
5 12* 15 6 25 0 1
14
16
12 28 5 28 12 10 17* 9 1 21 7 18 6 14 5 28 8* 0 16 20 7 6 8 4 2 8 15* 2 12* 14
16
32
8 16 3 15
8 10 7 3*
3 5 3
13 10
9
4
4
18 3 20 2 1 3 19 1 5 6 2 3 26 11 9 21 12 7 18 1 7 10 5 8 13 11 16 4 13 8 9 30 1 21
12
*MIC for Oxford Staphylococcus. Table n . Regression coefficients, slopes, intercepts and variance of slopes and intercepts of regression line depicted in Figure 2 Antibiotic Nafcillin Dicloxacillin Cloxacillin Flucloxacillin Propicillin Phenethicillin Methicillin Penicillin V Sulfocillin Azidocillin Carbenicillin Ticaicillin Amoxycillin Ampicillin Mecillinam
Regression coefficient
b
Variance ofi
c (intercept)
Variance
(slope)
0-88 0-90 0-88 0-88 0-88 0-93 0-94 0-94 0-93 0-95 0-94 0-94 0-88 0-92 0-94
0-814 0-727 0-72 0-751 0-787 0-805 0-881 0-859 0-916 0-895 0-839 0-850 1-403 1-528 1-29
00021 00013 00017 00019 00021 00011 00011 00011 00014 00011 00010 0-0011 00066 00051 00023
-2-935 -2-789 -2-598 -2-62 -213 -1-885 -1-73 -1-328 -1078 -0-789 -0-847 -0-821 0-378 0-994 -3-412
00067 00054* 00069* 00068* 00080 00053 00047 00061 00068 00081** 0-0086* * 00091** 0-039 0035 00039
• and • * regressions not statistically different.
ofc
Downloaded from http://jac.oxfordjournals.org/ at Université Laval on June 28, 2015
Amoxycillin Ampicillin 1* Azidocillin 4 Carbenicillin Cloxacillin Dicloxacillin Flucloxacillin Mecillinam Methicillin Nafcillin Phenethicillin Phenoxymethyl penicillin Propicillin Sulfocillin Ticarcillin 5
28*
8
334
B. A. Watts, I. Phillips and M. W. Stoate
control. The plates were read after overnight incubation at 37°C in 10% CO2. The MIC was taken as the concentration of antibiotic that completely inhibited growth of the organism. MIC's of each antibiotic were compared with those of benzylpenicillin for each strain, and regression lines drawn by the method of least square analysis. Statistical computations were carried out on a Varian 620 computer (Statistical appendix). A B
2
A Ampicillin B Amoxycillin I C Sulfocillin D Phenoxymethylpenicillin E Methicillin 05 F Meciilinam G Nafcillin _ BP Benzylpenicillin
BP
C
D
E F
025
2
0062 0031 0016 0008 0004
BP
0001
DC EAB
001
G
F 01 I Antibiotic MIC's (mg/l)
10
BP
o»
E
2 " A B 1. C D E 05 - F G H 0-25 BP
Azidocillin Ticorcillin Corbenicillin Phenethicillin Propicillin Flucloxacillin Cloxacillin Dicloxocillin Benzylpenicillin
/
ABC
D
J^
/ /
/
jf
' ' '/ / / / / / / / / / / / /
/ / / /
to
0 125
5
nzylperilcillir
100
///y ///y
E
FG H
/ /
/ / / /
/
/ ,///*/ / / /
0062 0031 -
w
o
0016 0008 n.nni 0 001 BCA
0 EGFH 001
01
10
100
Antibiotic MIC's (mg/l)
Figure 2. Regression lines for MIC's of benzylpenicillin against other penicillins and meciilinam.
Downloaded from http://jac.oxfordjournals.org/ at Université Laval on June 28, 2015
0125
Penicillins and N. gonorrhoeae
335
Results The distribution of MIC's obtained for the antibiotics, listed after benzylpenicillin in alphabetical order, is shown in Table I. Of the 92 strains studied 45 were relatively resistant to benzylpenicillin (MIC JJO-1 25 mg/1). Only four strains had MIC's as high as 1 mg/1. Regression lines of benzylpenicillin MIC's against MIC's of the other antibiotics are shown in Figure 2. Table II gives the regression coefficients, slopes and intercepts for these lines. From statistical analysis of these slopes and intercepts two groups were identified in which the antibiotics could not be considered significantly different. These were first, cloxacillin, dicloxacillin and flucloxacillin, and second, carbenicillin, ticarcillin and azidocillin. All other regressions differed significantly in slope, intercept or both. Discussion Downloaded from http://jac.oxfordjournals.org/ at Université Laval on June 28, 2015
Our results confirm that there are considerable differences in the inhibitory activity on N. gonorrhoeae of the various penicillins. From the regression lines, strains inhibited by benzylpenicillin 1 mg/1 would have the following theoretical MIC's to the other penicillins: ampicillin 0-5 mg/1; amoxycillin 1 mg/1; azidocillin, ticarcillin, carbenicillin 2 mg/1; sulfocillin, phenoxymethyl-penicillin 4 mg/1; methicillin, phenethicillin 8 mg/1; propicillin 16 mg/1; flucloxacillin, nafcillin, cloxacillin, dicloxacillin 32 mg/1. Mecillinam would have a MIC of 16 mg/1. The slopes of most of the regression lines were very similar, being slightly less than one, indicating that the antibiotics were relatively less active than benzylpenicillin against penicillin-resistant strains compared with fully sensitive strains. The exceptions to this were ampicillin, amoxycillin and mecillinam which had slopes greater than one and so were relatively more active against penicillin-resistant than against sensitive strains. Penicillins of possible clinical importance are those that show in vitro effectiveness against strains of N. gonorrhoeae that are relatively insensitive to benzylpenicillin, as failures of treatment are likely to occur with strains such as these. Our results, in agreement with those of 0degaard (1962) and Reyn & Bentzon (1968), showed that only ampicillin and amoxycillin are more active weight-for-weight than benzylpenicillin against penicillin insensitive strains. Unlike Martin, Lester, Kellogg & Thayer (1969) we did not find carbenicillin to have greater activity against penicillin insensitive strains. Carbenicillin, azidocillin and ticarcillin all showed equal activity to that of benzylpenicillin against sensitive strains but were less active against insensitive strains. Like Thayer & Axnick (1963) and Reyn & Bentzon (1968) respectively we found nafcillin and methicillin relatively ineffective. Similarly, the activity of the cloxacillin group was similar to that reported for oxacillin by these two groups of workers. We have been able to find only one report of sulfocillin MIC's for gonococci—one strain with an MIC of 0-39 mg/1, which resembles our relatively resistant strains. Finally our results for phenoxymethyl- and phenoxyethylpenicillin were very similar to those reported by Thayer and Axnick (1963). Chemotherapeutic effectiveness depends on many properties of antibiotics besides their in vitro effectiveness, but clinical trials have already shown the usefulness of ampicillin and amoxycillin (Morton, 1975), the two penicillins in this study to show good activity. Trials have also shown phenoxymethyl penicillin (Love & Weir, 1956; Shiel, 1956; Willcox, 1964), phenethicillin (Hilton, 1961; Allison, 1962; Willcox, 1964), and cloxacillin (Willcox, 1964) to be relatively ineffective. There would therefore appear to be
336
B. A. Watts, I. Phillips and M. W. Stoate
a reasonable relationship between MIC's and clinical effectiveness and on this basis ticarcillin, carbenicillin, azidocillin and sulfocillin might be worthy of clinical trials. Acknowledgements We thank the following pharmaceutical companies for their supplies of antibiotics, Beecham Research Laboratories (amoxycillin, ampicillin, azidocillin, benzylpenicillin, carbenicillin, cloxacillin, dicloxacillin, flucloxacillin, methicillin, phenethicillin, propicillin and ticarcillin). Wyeth Laboratories Inc. for nafcillin; Leo Laboratories Ltd for mecillinam; Lilly Research Centre Ltd for phenoxymethyl penicillin and Takeda Chemical Industries Ltd for sulfocillin. We also thank Mr T. Cowell, Head of the Department of Medical Electronics, St Thomas' Hospital for his assistance. This work was made possible by a DHSS grant.
Lund, F. & Tybring, L. 6P-Amidino-penicillanic acids—a new group of antibiotics. Nature, New Biology 236: 135-6 (1972). Martin, J. E., Lester, A., Kellogg, D. S. & Thayer, J. D. In vitro susceptibility of Neisseria gonorrhotae to nine antimicrobial agents. Applied Microbiology 18: 21-3 (1969). Morton, R. S. In Recent Advances in Sexually Transmitted Disease, Vol. 1, (R. S. Morton and J. R. W. Harris, Eds). Churchill Livingstone, Edinburgh, London and New York (1975. pp. 69-87. 0degaard, K. Penicillins in gonorrhoea. British Medical Journal i: 645 (1962). Phillips, I., Humphrey, D., Middleton, A. & Nicol, C. Diagnosis of gonorrhoea by culture on a selective medium containing vancomycin, colistin, nystatin and trimethoprim (VCNT): a comparison with Gram-staining and immunofluorescence. British Journal of Venereal Diseases 4: 287-92 (1972). Phillips, I., King, A., Warren, C , Watts, B. & Stoate, M. W. The activity of penicillin and eight cephalosporins on Neisseria gonorrhoeae. Journal of Antimicrobial Chemotherapy 2: 31-9 (1976). Reyn, A. & Bentzon, M. W. A study of the relationships between the sensitivities of Neisseria gonorrhoeae to sodium penicillin G, four semisynthetic penicillins, spiramycin and fusidic acid. British Journal of Venereal Diseases 44: 140-150 (1968). Sjoberg, B., Ekstrdm, B. & Forsgren, U. a-Azidobenzylpenicillin 1. Chemistry, bacteriology and experimental chemotherapy. Antimicrobial Agents and Chemotherapy—1967. 560-7 (1968). Sutherland, R., Burnett, J. & Rolinson, G. N. a-Carboxy-3-thienyl-methylpenicillin. Antimicrobial Agents and Chemotherapy—1970: 390-395 (1971). Thayer, J. D. and Axnick, N. W. Susceptibility of gonococci to ten penicillins. Antimicrobial Agents and Chemotherapy—1963: 427-9 (1964). Tsuchiya, K., Oishi, T., Iwagishi, C. & Iwahi, T. In vitro antibacterial activity of disodium asulfobenzylpenicillin. Journal of Antibiotics 24: 607-19 (1971). Willcox, R. R. The newer penicillins in the treatment of gonorrhoea. Postgraduate Medical Journal 40 (Suppl.): 202-7 (1964). Statistical appendix M. W. Stoate Firstly an analysis of variance (Kempthorne, 1952) was performed on the MIC results for the 16 antibiotics. This showed that there was a significant difference between the antibiotics at the 0-1 % level: thus all antibiotics could not be considered to be the same.
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References
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Regression analysis was then performed on the data, by the method previously described (Phillips, King, Warren, Watts & Stoate, 1976) to test these differences. T-tests (Beaumont, 1972) were also performed on the population means and these confirmed the results obtained by the above method. References Beaumont, G. P. Elementary Mathematical Statistics. McGraw Hill, New York (1972), pp. 224-227. Kempthorne, O. The Design and Analysis of Experiments. Wiley, New York (1952), pp. 32-33, Phillips, I., King, A., Warren, C , Watts, B. & Stoate, M. W. The activity of penicillin and eight cephalosporins on Neisseria gonorrhoeae. Journal of Antimicrobial Chemotherapy 2: 31-9 (1976). Downloaded from http://jac.oxfordjournals.org/ at Université Laval on June 28, 2015
(Manuscript accepted 22 October, 1976)
