Resistance of Salmonella serotypes to chloramphenicol*

BRIEF COMMUNICATIONS Resistance of Salmonella serotypes to chloramphenicol * S. K. SETHI,' S. ANAND,2 AJAIB SINGH,3 & D. V. VADEHRA Abstract A total of 704 Salmonella strains isolated from various sources were tested for their resistance to chloramphenicol. The results indicate that 36 out of 46 serotypes, comprising 671 strains (95.4 °O), were sensitive to chloramphenicol up to a level of 10 Mg per ml of medium. Most of the strains were sensitive at concentrations between 2 ug and 4 Mg of chlor- amphenicol per ml of medium. None of the strains isolated from animals, including poultry, was resist- ant. The results indicate that in India resistance of Salmonella strains to chloramphenicol has not increased over the years though this may have occurred in other parts of the world. The WHO Expert Committee on Antibiotics rec- ommended in its second report (1) that a survey of resistance of Salmonella typhi to chloramphenicol should be conducted in different parts of the world. Several workers in India (2-5) and elsewhere (8-12) have since obtained conflicting results regarding the frequency of chloramphenicol resistant Salmonella strains. In this paper we report results obtained with 704 Salmonella strains isolated from various sources and tested for their resistance to chloramphenicol. Materials and methods Salmonella strains. The strains used in the present studies were identified as salmonellae at the National Salmonella and Escherichia Centre, Kasauli, India, by biochemical, serological, and culture methods (6). A total of 704 Salmonella strains belonging to 46 serotypes were tested for sensitivity to chlor- amphenicol. The strains were isolated from diseased and apparently healthy humans and animals (includ- ing poultry) and from meat, foods, sewage, water, and other sources. Most of the strains were isolated in different parts of India but some strains isolated * From the Department of Microbiology, Panjab Uni- versity, Chandigarh, India. Reader in Medical Microbiology. 'Research student. 3Lecturer in Pathogenic Bacteriology. ' Professor of Microbiology. in Burma were sent to the Salmonella and Escheri- chia Centre for identification and serotyping. Antibiotic susceptibility test. Standard chloram- phenicol powder was dissolved in fresh distilled water and diluted to a stock concentration of 100 ,tg/ml. A fresh solution of antibiotic was used for each experiment. Medium. The medium used was modified Muller- Hinton broth and Muller-Hinton agar described by Otaya et al. (7). Various volumes of antibiotic solution at the stock concentration were added to flasks containing 200 ml of agar medium that had been previously melted and cooled to 44°C in a water bath. The concentration of antibiotic varied from 0.5 ,ug per ml of agar medium. The medium was then poured into Petri dishes (20 ml per plate) and allowed to set at room temperature (20c-25oC). Each plate was divided into eight segments for inoculation of eight strains. Similar agar plates without antibiotic served as controls. Inoculation and reading of plates. Tubes contain- ing 3 ml of broth were inoculated with a loopful of bacterial growth taken from a 24-h agar slant and incubated at 37'C for 18 hours. A standard inoculating wire loop was used to transfer a loopful of broth culture to a segment of an agar plate with and without added antibiotic. Inoculum for all strains was applied as spots of approximately equal size. The plates were incubated at 37^C for 18 hours. Organisms showing no growth were classified as sensitive while those showing confluent growth, as observed in the control plates, were considered resist- ant to the particular concentration of antibiotic incorporated in the medium. Results and discussion Reported resistance of Salnionella serotypes to chloramphenicol varies considerably. In India, Agar- wal (2) reported that 9% of S. typhi strains were resistant to chloramphenicol up to a concentration of 16 jug/ml while Murti et al. (3) found that up to 20% of Salmonella tested were resistant. In Nigeria, Njoku-Obi & Njoku-Obi (13) reported chlor- 3527 - 353 BULL. WORLD HEALTH ORGAN., Vol. 54, 1976 354 BRIEF COMMUNICATIONS t 50 z 40 z 30 0 2 0 z 10 0 05 1.0 2.0 4.0 6.0 8.0 100 CONCENTRATION OF CHLORAMPHENICOL ORAmt) Fig. 1. Chloramphenicol sensitivity pattern in 704 Salmonella strains belonging to 46 serotypes. amphenicol resistance in 25% of Salmonella sero- types. Our study shows that of 704 Salmonella strains, 671 (95.40 ) were sensitive to chlorampheni- col at concentrations ranging from 0.5 ,ug to 10 pg per ml of medium. The sensitivity pattern for dif- ferent concentrations of the antibiotic is shown in Fig. 1. Of the 46 serotypes tested, 36 were completely sensitive to chloramphenicol. Thirty-three strains that showed complete resistance included 29 clinical isolates from patients with symptoms of enteric fever, gasteroenteritis, and meningitis. The main serotypes manifesting these syndromes were S. typhi, S. paratyphi A, S. paratyphi B, S. virchow, S. new- port, S. pulloruim, and S. enteritidis. The other four resistant strains, isolated from water and sewage, belonged to the serotypes S. alachua and S. london. These results indicate a much lower resistance of Salmonella serotypes to chloramphenicol than that found in some of the earlier studies (2, 3, 13) but are in agreement with those of other studies made in India (4, 5, 14), the Netherlands (15), Singapore (17), and the USA (16). S. typhi is the major sero- type found in India, and in the present study com- prised 23.9% of the 704 strains tested. Only 1.700 of these S. typhi strains showed chloramphenicol resistance. S. paratyphi A is relatively infrequent in India and only 1 out of 18 strains was resistant to chloramphenicol, while in S. paratyphi B, which is extremely rare in India, 3 out of 18 strains tested were resistant. All these strains were isolated in Burma. Maximum resistance (100%0) was observed in S. alachua strains isolated from water and sewage. ACKNOWLEDGEMENTS The authors wish to thank the Director, Central Research Institute, Kasauli, India, for providing Salmo- nella strains. The chloramphenicol was kindly supplied by Parke-Davis (India) Ltd., Bombay. REFERENCES 1. WHO Technical Report Series, No. 210, 1961 (Standardization of methods for conducting microbic sensitivity tests: second report of the Expert Com- mittee on Antibiotics). 2. AGARWAL, S.C. Bulletin of the World Health Organ- ization, 27: 331-335 (1962). 3. MURTI, B. R. ET AL. Journal of clinical pathology, 15: 544-551 (1962). 4. SHARMA, K. B. & BHATIA, S. L. Indian journal of medical research, 54: 819-822 (1966) 5. 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