554
Correspondence
mahophilia to antimicrobial agents, singly and in combination. Antimicrobial Agents and Chemotherapy 16, 833-7. Garcia-Rodriguez, J. A., Garcia Sanchez, J. E., Garcia Garcia, M. I., Garcia Sanchez, E. & Mufioz Bellido, J. L. (1991). Antibiotic susceptibility profile of Xamhomonas maltophilia. "In vitro" activity of /Mactam//Mactamase inhibitor combinations. Diagnostic Microbiology and Infectious Disease 14, in press. Holmes, B., Lapage, S. P. & Easterling, B. G. (1979). Distribution in clinical material and identification of Pseudomonas maltophilia. Journal of Clinical Pathology 32, 66-72. Kroogstad, D. J. & Moellering, R. C. (1986). Antimicrobial combinations. In Antibiotic in Laboratory Medicine, 2nd edn (Lorian, V., Ed.), pp. 537-95. Williams & Wilkins, Baltimore. Lorian, V. (1980). Effects of subminimum inhibitory concentrations of antibiotics on bacteria. In Antibiotic in Laboratory Medicine (Lorian, V., Ed.), pp. 3 4 2 ^ 0 8 . Williams & Wilkins, Baltimore. Metchock, B. & Thornsberry, C. (1989). Susceptibility of Pseudomonas (Xanthomonas) maltophilia to antimicrobial agents. Antimicrobic Newsletter 6, 35-40. Mett, H., Rosta, S., Schacher, B. & Frei, R. (1988). Outer membrane permeability and /Mactamase content in Pseudomonas maltophilia clinical isolates and laboratory mutants. Reviews of Infectious Diseases 10, 765-9. Morrison, A. J., Hoffman, K. K. & Wenzel, R. P. (1986). Associated mortality and clinical characteristics of nosocomial Pseudomonas maltophilia in a university hospital. Journal of Clinical Microbiology 24, 52-5. Muder, R. R., Yu, V. L., Dummer, J. S., Vinson, C. & Lumish, R. M. (1987). Infections caused by Pseudomonas maltophilia. Archives of Internal Medicine 147, 1672-4. National Committee for Clinical Laboratory Standards (1985). Methods for dilution antimicrobial susceptibility tests for bacteria that grow aerobically, approved standards. NCCLS publication M-7 A. Villanova, Pa. Neu, H. C. (1984). Unusual nosocomial infections. Disease-a-Month 30, 1-68. Neu, H. C , Saha, G. & Chin, N. X. (1989). Resistance of Xanthomonas maltophilia to antibiotics and the effect of/Mactamase inhibitors. Diagnostic Microbiology and Infectious Disease 12, 283-5. Saino, Y., Kobayashi, F., Inoue, M. & Mitsuhashi, S. (1982). Purification and properties of inducible penicillin /Mactamase isolated from Pseudomonas maltophilia. Antimicrobial Agents and Chemotherapy 22, 564-70. Schoencknecht, F. D., Sabath, L. D. & Thornsberry, C. (1985). Susceptibility tests: special tests. In Manual of Clinical Microbiology, 2nd edn (Lennette, E. H., Balows, A., Hausler, W. J. & Shadomy, H. J., Eds), pp. 1000-8. American Society for Microbiology, Washington, DC. Swabb, E. A. (1985). Clinical pharmacology of
aztreonam in healthy recipients and patients: a review. Reviews of Infectious Diseases 7, Suppl. 4, 605-12. Wheat, P. F. (1985). Effect of temperature on antimicrobial susceptibilities of Pseudomonas maltophilia. Journal of Clinical Pathology 38, 1055-8. Zuravleff, J. J. & Yu, V. L. (1982). Infections caused by Pseudomonas maltophilia with emphasis on bacteremia: case reports and review of the literature. Reviews of Infectious Diseases 4, 1236-46.
Muitiresistant Salmonella typhi in Bangladesh Sir, There are recent reports of emergence of Salmonella typhi resistant to chloramphenicol, ampicillin, co-trimoxazole and tetracycline from southern and northern parts of India (Gupta, Bhujwala & Srinivas, 1990; Jesudasan & John, 1990). At this Centre, we have also isolated from stool and/or blood cultures of 13 patients, 16 strains of 5. typhi which were resistant to chloramphenicol, ampicillin, cotrimoxazole and tetracycline (Table I), but susceptible to gentamicin, nalidixic acid, ciprofloxacin, pivmecillinam, ceftriaxone and tobramycin by a disk diffusion method (Bauer et al, 1966). All the 16 strains possessed a single plasmid of approximately 110 MDa after extraction and electrophoresis by standard methods (Myers et al., 1976; Birnboim & Doly, 1979). In conjugation experiments (Smith & Guild, 1980) with two 5. typhi strains, using Escherichia colt K-12 (14R525) as the recipient, both strains transferred the resistance to all four drugs with the simultaneous acquisition ofl 10 MDa plasmid by the exconjugants. This confirms the result of the south Indian study that drug resistance is associated with a plasmid (Jesudasan & John, 1990). The MICs of the drugs for donors and ex-conjugants were 500 mg/1 for chloramphenicol, > 2000 mg/1 for ampicillin, 2000 mg/1 for trimethoprim and 125 mg/1 for tetracycline. A S. typhi strain resistant to chloramphenicol, ampicillin, co-trimoxazole, cloxacillin and cephalexin was first reported from a child with enteric fever from Bangladesh in 1986 (Morshed et al., 1986). The simultaneous emergence of muitiresistant strains from various parts of the Indian subcontinent may point to a subcontinent-wide outbreak. It will be interesting to study the clonal origin of the strains from different geographical locations of the subcontinent.
Correspondence Table I. Isolates of S. typhi resistant to four antimicrobial agents'
Number of isolates
Month September, 1989 October November December January, 1990 February March April
May June July August Total
2 11 11 5 17 9 12 12 11 16 13 16 135
Number resistant to four antimicrobial agents
0 0 0 0 0 2 0 0 0 4 7 3 16(11-9%)
"Chloramphenicol, ampicillin, co-trimoxazole and tetracycline. Ampicillin, chloramphenicol and co-trimoxazole are the drugs of choice in the management of enteric fever, and emergence of resistance to these drugs is indeed worrying. Medical practitioners in the subcontinent should be aware of the multiresistant S. typhi when treating patients with enteric fever. M. JOHN ALBERT K. HAIDER S. NAHAR A. K. M. G. KIBRIYA M. A. HOSSAIN International Centre for Diarrhoeal Disease Research, G.P.O. Box 128, Dhaka-1000, Bangladesh References Bauer, A. W., Kirby, W. M. M., Sherris, J. C. & Turck, M. (1966). Antibiotic susceptibility by a standardised single disk method. American Journal of Clinical Pathology 45, 493-6. Birnboim, H. C. & Doly, J. (1979). A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acid Research 7, 1513-23. Gupta, B. L., Bhujwala, R. A. & Sriniwas. (1990). Multiresistant Salmonella typhi in India. Lancet 336, 252. Jesudasan, M.V. & John, T. J. (1990). Multiresistant Salmonella typhi in India. Lancet 336, 252. Meyers, J. A., Sanchez, D., Elwell, L. P. & Falkow, S. (1976). Simple agarose gel electrophoresis method for the identification and characterization of plasmid deoxyribonucleic acid. Journal of Bacteriology 127, 1529-37.
555
Morshed, M. G., Khan, W.A., Khan, N. Z. & Akbar, M. S. (1986). Multiple drug resistant Salmonella typhi in Bangladesh. Journal of Diarrhoeal Diseases Research 4, 241. Smith, M. D. & Guild, W. R. (1980). Improved method for conjugative transfer by filter mating of Streptococcus pneumoniae. Journal of Bacteriology
144, 457-9.
Late quinolone resistance in Pseudomonas aeruginosa Sir, In Dunedin Hospital, antibiotic sensitivity tests are routinely carried out on all potentially significant isolates of Gram-negative bacilli. Using a straight wire, material from one colony (rarely two to three similar colonies where they are very small or particularly difficult to emulsify) on the primary isolation medium is inoculated into Ringer's solution to provide a suspension of around 106 cfu/ml. Using a multipoint inoculating device, this suspension is then inoculated (approximately 104— 10s cfu/drop) onto a variety of media including Iso-Sensitest agar (ISA) plates containing breakpoint concentrations of various antibiotics—5 mg/1 in the case of norfloxacin. After overnight incubation at 37CC, sensitivities are read by recording the presence or absence of growth in comparison with an ISA control plate free of antibiotic. During early 1990, a series of isolates of Pseudomonas aeruginosa from a patient given oral ciprofloxacin (750 mg twice daily) for 12 days in an attempt to clear colonization of cutaneous burns with this microbe prior to skin grafting, revealed unusual norfloxacin susceptibility results. Before the commencement of ciprofloxacin therapy, isolates of P. aeruginosa from the patient appeared susceptible to norfloxacin on routine testing. Unfortunately, ciprofloxacin sensitivities were not requested although the range of P. aeruginosa MICs in Dunedin at that time was around 0-25-0-5 mg/1. Following the cessation of antimicrobial therapy, the grafted burn sites as well as the donor skin site became recolonized with P. aeruginosa which appeared also susceptible to norfloxacin on routine testing. A feature of these apparently susceptible isolates, however, was that if the sensitivity plates were left at room temperature for a further 48-72 h, growth occurred—i.e. isolates revealed "late" resistance. This prompted a more extensive investigation of cells in these "slowly resistant isolates". Subcultures (on blood agar) were prepared
Correspondence
mahophilia to antimicrobial agents, singly and in combination. Antimicrobial Agents and Chemotherapy 16, 833-7. Garcia-Rodriguez, J. A., Garcia Sanchez, J. E., Garcia Garcia, M. I., Garcia Sanchez, E. & Mufioz Bellido, J. L. (1991). Antibiotic susceptibility profile of Xamhomonas maltophilia. "In vitro" activity of /Mactam//Mactamase inhibitor combinations. Diagnostic Microbiology and Infectious Disease 14, in press. Holmes, B., Lapage, S. P. & Easterling, B. G. (1979). Distribution in clinical material and identification of Pseudomonas maltophilia. Journal of Clinical Pathology 32, 66-72. Kroogstad, D. J. & Moellering, R. C. (1986). Antimicrobial combinations. In Antibiotic in Laboratory Medicine, 2nd edn (Lorian, V., Ed.), pp. 537-95. Williams & Wilkins, Baltimore. Lorian, V. (1980). Effects of subminimum inhibitory concentrations of antibiotics on bacteria. In Antibiotic in Laboratory Medicine (Lorian, V., Ed.), pp. 3 4 2 ^ 0 8 . Williams & Wilkins, Baltimore. Metchock, B. & Thornsberry, C. (1989). Susceptibility of Pseudomonas (Xanthomonas) maltophilia to antimicrobial agents. Antimicrobic Newsletter 6, 35-40. Mett, H., Rosta, S., Schacher, B. & Frei, R. (1988). Outer membrane permeability and /Mactamase content in Pseudomonas maltophilia clinical isolates and laboratory mutants. Reviews of Infectious Diseases 10, 765-9. Morrison, A. J., Hoffman, K. K. & Wenzel, R. P. (1986). Associated mortality and clinical characteristics of nosocomial Pseudomonas maltophilia in a university hospital. Journal of Clinical Microbiology 24, 52-5. Muder, R. R., Yu, V. L., Dummer, J. S., Vinson, C. & Lumish, R. M. (1987). Infections caused by Pseudomonas maltophilia. Archives of Internal Medicine 147, 1672-4. National Committee for Clinical Laboratory Standards (1985). Methods for dilution antimicrobial susceptibility tests for bacteria that grow aerobically, approved standards. NCCLS publication M-7 A. Villanova, Pa. Neu, H. C. (1984). Unusual nosocomial infections. Disease-a-Month 30, 1-68. Neu, H. C , Saha, G. & Chin, N. X. (1989). Resistance of Xanthomonas maltophilia to antibiotics and the effect of/Mactamase inhibitors. Diagnostic Microbiology and Infectious Disease 12, 283-5. Saino, Y., Kobayashi, F., Inoue, M. & Mitsuhashi, S. (1982). Purification and properties of inducible penicillin /Mactamase isolated from Pseudomonas maltophilia. Antimicrobial Agents and Chemotherapy 22, 564-70. Schoencknecht, F. D., Sabath, L. D. & Thornsberry, C. (1985). Susceptibility tests: special tests. In Manual of Clinical Microbiology, 2nd edn (Lennette, E. H., Balows, A., Hausler, W. J. & Shadomy, H. J., Eds), pp. 1000-8. American Society for Microbiology, Washington, DC. Swabb, E. A. (1985). Clinical pharmacology of
aztreonam in healthy recipients and patients: a review. Reviews of Infectious Diseases 7, Suppl. 4, 605-12. Wheat, P. F. (1985). Effect of temperature on antimicrobial susceptibilities of Pseudomonas maltophilia. Journal of Clinical Pathology 38, 1055-8. Zuravleff, J. J. & Yu, V. L. (1982). Infections caused by Pseudomonas maltophilia with emphasis on bacteremia: case reports and review of the literature. Reviews of Infectious Diseases 4, 1236-46.
Muitiresistant Salmonella typhi in Bangladesh Sir, There are recent reports of emergence of Salmonella typhi resistant to chloramphenicol, ampicillin, co-trimoxazole and tetracycline from southern and northern parts of India (Gupta, Bhujwala & Srinivas, 1990; Jesudasan & John, 1990). At this Centre, we have also isolated from stool and/or blood cultures of 13 patients, 16 strains of 5. typhi which were resistant to chloramphenicol, ampicillin, cotrimoxazole and tetracycline (Table I), but susceptible to gentamicin, nalidixic acid, ciprofloxacin, pivmecillinam, ceftriaxone and tobramycin by a disk diffusion method (Bauer et al, 1966). All the 16 strains possessed a single plasmid of approximately 110 MDa after extraction and electrophoresis by standard methods (Myers et al., 1976; Birnboim & Doly, 1979). In conjugation experiments (Smith & Guild, 1980) with two 5. typhi strains, using Escherichia colt K-12 (14R525) as the recipient, both strains transferred the resistance to all four drugs with the simultaneous acquisition ofl 10 MDa plasmid by the exconjugants. This confirms the result of the south Indian study that drug resistance is associated with a plasmid (Jesudasan & John, 1990). The MICs of the drugs for donors and ex-conjugants were 500 mg/1 for chloramphenicol, > 2000 mg/1 for ampicillin, 2000 mg/1 for trimethoprim and 125 mg/1 for tetracycline. A S. typhi strain resistant to chloramphenicol, ampicillin, co-trimoxazole, cloxacillin and cephalexin was first reported from a child with enteric fever from Bangladesh in 1986 (Morshed et al., 1986). The simultaneous emergence of muitiresistant strains from various parts of the Indian subcontinent may point to a subcontinent-wide outbreak. It will be interesting to study the clonal origin of the strains from different geographical locations of the subcontinent.
Correspondence Table I. Isolates of S. typhi resistant to four antimicrobial agents'
Number of isolates
Month September, 1989 October November December January, 1990 February March April
May June July August Total
2 11 11 5 17 9 12 12 11 16 13 16 135
Number resistant to four antimicrobial agents
0 0 0 0 0 2 0 0 0 4 7 3 16(11-9%)
"Chloramphenicol, ampicillin, co-trimoxazole and tetracycline. Ampicillin, chloramphenicol and co-trimoxazole are the drugs of choice in the management of enteric fever, and emergence of resistance to these drugs is indeed worrying. Medical practitioners in the subcontinent should be aware of the multiresistant S. typhi when treating patients with enteric fever. M. JOHN ALBERT K. HAIDER S. NAHAR A. K. M. G. KIBRIYA M. A. HOSSAIN International Centre for Diarrhoeal Disease Research, G.P.O. Box 128, Dhaka-1000, Bangladesh References Bauer, A. W., Kirby, W. M. M., Sherris, J. C. & Turck, M. (1966). Antibiotic susceptibility by a standardised single disk method. American Journal of Clinical Pathology 45, 493-6. Birnboim, H. C. & Doly, J. (1979). A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acid Research 7, 1513-23. Gupta, B. L., Bhujwala, R. A. & Sriniwas. (1990). Multiresistant Salmonella typhi in India. Lancet 336, 252. Jesudasan, M.V. & John, T. J. (1990). Multiresistant Salmonella typhi in India. Lancet 336, 252. Meyers, J. A., Sanchez, D., Elwell, L. P. & Falkow, S. (1976). Simple agarose gel electrophoresis method for the identification and characterization of plasmid deoxyribonucleic acid. Journal of Bacteriology 127, 1529-37.
555
Morshed, M. G., Khan, W.A., Khan, N. Z. & Akbar, M. S. (1986). Multiple drug resistant Salmonella typhi in Bangladesh. Journal of Diarrhoeal Diseases Research 4, 241. Smith, M. D. & Guild, W. R. (1980). Improved method for conjugative transfer by filter mating of Streptococcus pneumoniae. Journal of Bacteriology
144, 457-9.
Late quinolone resistance in Pseudomonas aeruginosa Sir, In Dunedin Hospital, antibiotic sensitivity tests are routinely carried out on all potentially significant isolates of Gram-negative bacilli. Using a straight wire, material from one colony (rarely two to three similar colonies where they are very small or particularly difficult to emulsify) on the primary isolation medium is inoculated into Ringer's solution to provide a suspension of around 106 cfu/ml. Using a multipoint inoculating device, this suspension is then inoculated (approximately 104— 10s cfu/drop) onto a variety of media including Iso-Sensitest agar (ISA) plates containing breakpoint concentrations of various antibiotics—5 mg/1 in the case of norfloxacin. After overnight incubation at 37CC, sensitivities are read by recording the presence or absence of growth in comparison with an ISA control plate free of antibiotic. During early 1990, a series of isolates of Pseudomonas aeruginosa from a patient given oral ciprofloxacin (750 mg twice daily) for 12 days in an attempt to clear colonization of cutaneous burns with this microbe prior to skin grafting, revealed unusual norfloxacin susceptibility results. Before the commencement of ciprofloxacin therapy, isolates of P. aeruginosa from the patient appeared susceptible to norfloxacin on routine testing. Unfortunately, ciprofloxacin sensitivities were not requested although the range of P. aeruginosa MICs in Dunedin at that time was around 0-25-0-5 mg/1. Following the cessation of antimicrobial therapy, the grafted burn sites as well as the donor skin site became recolonized with P. aeruginosa which appeared also susceptible to norfloxacin on routine testing. A feature of these apparently susceptible isolates, however, was that if the sensitivity plates were left at room temperature for a further 48-72 h, growth occurred—i.e. isolates revealed "late" resistance. This prompted a more extensive investigation of cells in these "slowly resistant isolates". Subcultures (on blood agar) were prepared
