Antibiogram of S. enterica serovar Typhi and S. enterica serovar Paratyphi A: a multi-centre study from India

WHO South-East Asia Journal of Public Health 2012;1(2):182-188182 Original research Antibiogram of S. enterica serovar Typhi and S. enterica serovar Paratyphi A: a multi-centre study from India Indian Network for Surveillance of Antimicrobial Resistance Groupa Background: Enteric fever continues to be a public health problem in many countries including India. Emergence of the multidrug resistant strains of S. enterica serovar Typhi may render treatment with antibiotics ineffective. A multi-centre surveillance study was, therefore, conducted in India to monitor the time trends in antibiotic susceptibility patterns of S. enterica serovar Typhi and S. enterica serovar Paratyphi A in India. Methods: All S. enterica serovar Typhi and S. enterica serovar Paratyphi A strains isolated from January 2008 to December 2010 in the 15 participating centres were included in the study. Each centre compiled their data in a predefined template which included data of the antimicrobial susceptibility pattern, location of the patient and specimen type. The data in the submitted templates was collated and analysed using a common protocol. Results: A total of 3275 isolates of Salmonellae causing enteric fever were included in the study. There were 2511 S. enterica serovar Typhi and 764 S. enterica serovar Paratyphi A strains during the three-year study period. Resistance to nalidixic acid was seen in 83% of the S. enterica serovar Typhi and 93% of S. enterica serovar Paratyphi A strains. Majority of the strains were susceptible to third generation cephalosporins. Conclusions: Enteric fever in India is caused by S. enterica serovar Typhi and S. enterica serovar Paratyphi A. Nalidixic acid resistance is high among both S. enterica serovar Typhi and S. enterica serovar Paratyphi A. Susceptibility to ampicillin, chloramphenicol and cotrimoxazole is high. Third generation cephalosporins continue to remain susceptible. Key words: S. enterica serovar Typhi , S. enterica serovar Paratyphi A, antibiotics, India a Indian Network for Surveillance of Antimicrobial Resistance Correspondence to Sangeeta Joshi (email: sangeetajo@yahoo.com, insar_india@yahoogroups.com) Introduction Enteric fever continues to remain a public health problem in many countries. In 2000, typhoid fever caused an estimated 21.7 million illnesses and 217 000 deaths, and paratyphoid fever caused an estimated 5.4 million illnesses worldwide.1 In Asia, Crump et al reported a crude incidence of typhoid as 274 per 100 000 persons.1 Salmonella enterica serotype Typhi and Salmonella enterica serotype Paratyphi A are the main causes of enteric fever in India; S. enterica serovar Typhi being predominant.2 In 1999, Sinha et al found the incidence rate to be 9.8 per 1000 person years in an urban slum of North India.3 However, in 2008, Ochlai et al who conducted their surveillance in an urban Book 1.indb 182 05-Jun-2012 2:35:59 PM WHO South-East Asia Journal of Public Health 2012;1(2):182-188 183 Antibiogram of S. typhi and S. paratyphi A in IndiaIndian Network for Surveillance of Antimicrobial Resistance Group slum in Kolkata in eastern India, reported an incidence of 214.2 per 100 000/year.4 After the first reported outbreak of chloramphenicol resistant S. enterica serovar Typhi in 1972, there has been a steady increase in the number of multidrug resistant (MDR) strains of S. enterica serovar Typhi – resistance to ampicillin, chloramphenicol and trimethoprim- sulphamethoxazole - over the next two decades. With the increasing use of fluoroquinolones in the 1990s for treatment of enteric fever, there was a gradual decrease in MDR of S. enterica serovar Typhi with emergence of nalidixic acid resistant strains. A network of microbiology laboratories (Indian Network for Survei l lance of Antimicrobial Resistance) at premier medical colleges and hospitals in India was formed with support from the World Health Organization (Figure 1). The network aims to monitor and Figure 1: INSAR sites India (alphabetically as per institutional name) The sites marked with a red dot indicates those centres whose data is included in the study. All India Institute of Medical Sciences (AIIMS) - New Delhi, Apollo Health City - Hyderabad, BJ medical college (BJMC) - Pune, Chacha Nehru Bal Chikitsalaya (CNBC) - New Delhi, Choithram Hospital and Research Centre - Indore , Christian Medical College (CMC) - Vellore, Fortis Hospital - Mohali, Global Hospitals - Hyderabad, Government Medical College & Hospital (GMC) - Chandigarh, Government medical college (GMC) - Aurangabad, Gujarat Cancer and research Institute (GCRI) - Ahmedabad, Indraprastha Apollo Hospital - New Delhi, Jawaharlal Institute of Postgraduate Medical Education and Research (JIPMER) - Puducherry Manipal Hospital - Bangalore, PD Hinduja National Hospital & MRC (PDNH) - Mumbai, Postgraduate Institute of Medical Education & Research (PGIMER) - Chandigarh, Regional institute of medical sciences (RIMS) - Imphal, Sanjay Gandhi Postgraduate Institute of Medical Sciences (SGPGI) - Lucknow, Stanley medical college (SMC) - Chennai. Book 1.indb 183 05-Jun-2012 2:36:00 PM WHO South-East Asia Journal of Public Health 2012;1(2):182-188184 Antibiogram of S. typhi and S. paratyphi A in India Indian Network for Surveillance of Antimicrobial Resistance Group review the antimicrobial resistance problem in India. Initially, few organisms of public health importance were chosen for monitoring the prevalence and antimicrobial resistance patterns. Therefore, S. enterica serovar Typhi and S. enterica serovar Paratyphi A were chosen from amongst the Gram negative organisms. All participating laboratories shared their antimicrobial susceptibility data and provided technical support to other members. This study provides a national level understanding of the emerging trends of antimicrobial resistance among clinical isolates of S. enterica serovar Typhi and S. enterica serovar Paratyphi A and provides a platform for initiating epidemiological studies of enteric fever. Methods This study was conducted over three years (January 2008 to December 2010) retrospectively. Each centre compiled their susceptibility data for S. enterica serovar Typhi and S. enterica serovar Paratyphi A isolates for the study period in a defined template. The data collection template included patient’s location, source / specimen of isolation and the antibiotic susceptibility profiles. Blood cultures were done by the conventional microbiological techniques or automated systems in the participating centres. The identification of Salmonellae was done by the standard biochemical tests and confirmation was done by serotyping.5 The antibiotic susceptibility testing was performed at different study sites by the Kirby Bauer’s’ disc diffusion technique and / or by MIC testing, using CLSI recommendations.6 One laboratory used the BSAC guidelines for antimicrobial testing and interpretations.7 The antibiotics tested included ampicillin (10μg), co-trimoxazole (1.25/23.75 μg), ciprofloxacin (5 μg), nalidixic acid (30 μg), ceftriaxone (30 μg), and chloramphenicol(30 μg). Inoculum was prepared by making a direct saline suspension of isolated colonies selected from an 18- to 24-hour blood agar plate. Turbidity of the suspension was adjusted to achieve a turbidity equivalent to a 0.5 McFarland standard and five disks were applied on a 100mm Mueller Hinton agar plate as per CLSI guidelines / BSAC. E. coli ATCC 25922 was used as the quality control strain for disc diffusion. Results A total of 3275 isolates of Salmonellae causing enteric fever were included in the study. There were 2511 S. enterica serovar Typhi strains (430 in 2008, 694 in 2009 and 1387 in 2010) and 764 S. enterica serovar Paratyphi A strains (311, 217 and 236 in 2008, 2009 and 2010 respectively) during the study period. These strains were isolated predominantly from blood culture. Few isolates from pus (3), stool (14) and urine (8) also were also included in the study. The antibiotic susceptibility (by CLSI guidelines) for S. enterica serovar Typhi and S. enterica serovar Paratyphi A isolates for the study period are shown in Table 1 and Table 2. Nalidixic acid resistance was high in both S. enterica serovar Typhi and S. enterica serovar Paratyphi A, being 82% and 93% respectively during the three-year period. Third generation cephalosporins were 100% susceptible in 2008. Resistance to third generation cephalosporins was seen in 3% of strains of S. enterica serovar Typhi in 2009 and in 1% S. enterica serovar Paratyphi A in 2010. Multidrug resistance (MDR) to ampicillin, chloramphenicol and cotrimoxazole in S. enterica serovar Typhi was observed in less than 5% isolates. The antibiogram for the isolates reported by BSAC guidelines is summarized in Table 3. Book 1.indb 184 05-Jun-2012 2:36:00 PM WHO South-East Asia Journal of Public Health 2012;1(2):182-188 185 Antibiogram of S. typhi and S. paratyphi A in IndiaIndian Network for Surveillance of Antimicrobial Resistance Group Table 2: Antibiogram of S. enterica serovar Paratyphi A. (% susceptible) Year N Amp Chl Sxt Ctri Cip NA 2008 311 99 99 100 100 100 12 2009 217 91 100 91 100 85 2.9 2010 236 95 100 97 99 71 7.2 Amp-Ampicillin, Chl- Chloramphenicol, Sxt- Cotrimoxazole, Ctri – Ceftriaxone, Cip- Ciprofloxacin, NA- Nalidixic acid Table 3: Antibiogram of S. enterica serovar Typhi and S. enterica serovar Paratyphi A by BSAC guidelines (% susceptible) Year Isolate N Amp Chl Sxt Ctri Cip NA 2009 S. enterica serovar Typhi 43 88 100 100 100 79 77 2010 S. enterica serovar Typhi 55 85 100 100 100 13 9 2009 S. enterica serovar Paratyphi A 22 68 100 100 100 14 9 2010 S. enterica serovar Paratyphi A 18 33 56 44 100 17 17 Amp-Ampicillin, Chl- Chloramphenicol, Sxt- Cotrimoxazole, Ctri – Ceftriaxone, Cip- Ciprofloxacin, NA- Nalidixic acid Table 1: Antibiogram of S. enterica serovar Typhi. (% susceptible) Year N Amp Chl Sxt Ctri Cip NA 2008 430 95 96 96 100 99 23 2009 694 96 97 95 97 75 22 2010 1387 89 95 94 100 59 8.3 Amp-Ampicillin, Chl- Chloramphenicol, Sxt- Cotrimoxazole, Ctri – Ceftriaxone, Cip- Ciprofloxacin, NA- Nalidixic acid Book 1.indb 185 05-Jun-2012 2:36:00 PM WHO South-East Asia Journal of Public Health 2012;1(2):182-188186 Antibiogram of S. typhi and S. paratyphi A in India Indian Network for Surveillance of Antimicrobial Resistance Group Discussion Enteric fever continues to remain an important infection in endemic countries and to travelers to these areas. As observed in the present study, enteric fever in India is mainly caused by S. enterica serovar Typhi and S. enterica serovar Paratyphi A. Nalidixic acid resistance is high among both S. enterica serovar Typhi and S. enterica serovar Paratyphi A. Susceptibility to ampicillin, chloramphenicol and cotrimoxazole - traditional first line drugs for enteric fever treatment is good. Third generation cephalosporins continue to remain susceptible and are a useful choice for the treatment of enteric fever. The major strength of the study is that it had representations from 15 centres which used similar protocols for data collection. However, one limitation of this study is the fact that there was inadequate representation from eastern India. In a study of six years (2000 to 2006) from north India, there were a predominance of S. enterica serovar Typhi (62%) , followed by 38% S. enterica serovar Paratyphi A.8 The study highlighted the emergence of S. enterica serovar Paratyphi A as the predominant serotype in 2003-2004 with resurgence of S. enterica serovar Typhi in the subsequent period in their study. MDR S. enterica serovar Typhi was 10.69% while 13.13% were MDR S. enterica serovar Paratyphi A. Another study from north India also reported high proportion of S. enterica serovar Paratyphi A with nalidixic acid resistance (92.5%).9 Interestingly, 90% of their isolates were sensitive to chloramphenicol. Another study from a tertiary care hospital in Delhi from 1999 to 2004, revealed a gradual decrease in MDR S. enterica serovar Typhi,10 However, MDR S. enterica serovar Paratyphi A increased from 3.5% in 1999 to 11.6% in 2004. The proportion of S. enterica serovar Paratyphi A rose from 20.3% in 1999 to 30.3% in 2004. The gradual increase in prevalence of S. enterica serovar Paratyphi A over the years could probably be due to the introduction of monovalent vaccine effective against S. enterica serovar Typhi only gradually replacing the bivalent TA vaccine.11 A recent study from eastern India showed multidrug resistance in 11.9% and 15.6% of S. enterica serovar Typhi and S. enterica serovar Paratyphi A isolates respectively.2 The percentage of S. enterica serovar Typhi isolations were more than S. enterica serovar Paratyphi A. Similarly, we also had a larger number of S. enterica serovar Typhi isolates (77%) than S. enterica serovar Paratyphi A (23%). Ray et al found that nalidixic acid (NA) susceptibility was a good marker for fluoroquinolone susceptibility but NA resistance had a poor predictive value for ciprofloxacin resistance.12 They suggested that NA resistant isolates should be tested for ciprofloxacin MIC before deciding a change in therapeutic regimen. Nalidixic acid resistant isolates of S. enterica serovar Typhi and S. enterica serovar Paratyphi A had higher MIC’s to fluoroquinolones as compared to the nalidixic acid susceptible ones.13 In a study from Puducherry, south India, there was a high rate of ciprofloxacin resistance (8%) observed in S. enterica serovar Typhi, with 78% isolates being NA resistant.14 They also reported a high rate of MDR S. enterica serovar Typhi (22%). Chitnis et al reported a gradually increasing MIC to ciprofloxacin from <=0.125 mg/L to > 1mg/L among the isolates of S. enterica serovar Typhi over the years 1988 to 2005.15 Concurrently MDR, which was seen in upto 90% isolates in 1990-91 had declined to 5.6% in 2005. In view of the increasing MIC’s of ciprofloxacin, there has been a suggestion to relook at the breakpoints and the zone diameters for reporting ciprofloxacin for Salmonella.16 As per BSAC guidelines 2011 for ciprofloxacin for Salmonella, it is recommended that isolates Book 1.indb 186 05-Jun-2012 2:36:00 PM WHO South-East Asia Journal of Public Health 2012;1(2):182-188 187 Antibiogram of S. typhi and S. paratyphi A in IndiaIndian Network for Surveillance of Antimicrobial Resistance Group with MIC > 0.06 mg/L should be reported as resistant. In the present study, MIC > 0.125mg/L was interpreted as intermediate susceptible/ resistant for the isolates reported by one laboratory as per BSAC guidelines 2010.7 In the present study, S. enterica serovar Typhi susceptibility to ampicillin, chloramphenicol and cotrimoxazole was 93%, 96% and 95% respectively. This is similar to a study done in south India where there was a significant increase in sensitivity to chloramphenicol (86%), ampicillin (84%) and cotrimoxazole (88%). MDR was seen in 12% cases in the same study.17 There is a marked reduction in the reduction of MDR isolates as compared to studies in early 2000.10 The high prevalence of MDR among Salmonella species had led to fluoroquinolones assuming a primary role in the therapy. Some investigators have noted increases in the prevalence of S. enterica serovar Typhi and S. enterica serovar Paratyphi A strains susceptible to traditional first-line antimicrobials coinciding with a switch to fluoroquinolones for the management of enteric fever.18 Azithromycin has been seen to be efficacious for the treatment of typhoid fever;19 however, there are reports of emergence of resistance to azithromycin.20 There are sporadic reports of third generation cephalosporin resistance in Salmonella. Gokul et al. reported an ACC-1 AmpC β- lactamase producing S. enterica serovar Typhi.21 However, third generation cephalosporins still continue to be a good option for the treatment of enteric fever. To conclude, this study demonstrates the re-emergence of susceptibility to ampicillin, chloramphenicol and cotrimoxazole in Salmonella enterica serovar Typhi, a decline in MDR strains and a high resistance to nalidixic acid in India. Third generation cephalosporins seem to be effective therapeutic options. Acknowledgement This work and the INSAR group are supported by the World Health Organization. aIndian Network for Surveillance of Antimicrobial Resistance (INSAR) (alphabetically as per institutional name) All India Institute of Medical Sciences (AIIMS), New Delhi – Arti Kapil Apollo Health City, Hyderabad - Ratna Rao BJ medical college (BJMC), Pune - Anju Kagal Chacha Nehru Bal Chikitsalaya (CNBC), New Delhi - Vikas Manchanda Choithram Hospital and Research Centre, Indore - DS Chitnis Christian Medical College (CMC), Vellore - Veeraraghavan Balaji Fortis Hospital, Mohali - Anita Sharma Global Hospitals, Hyderabad - Ranganathan N Iyer Government Medical College & Hospital (GMC), Chandigarh - Varsha Gupta Indraprastha Apollo Hospital, New Delhi - Raman Sardana Jawaharlal Institute of Postgraduate Medical Education and Research (JIPMER), Puducherry - BN Harish Manipal Hospital, Bangalore - Sangeeta Joshi PD Hinduja National Hospital & MRC (PDNH), Mumbai - Camilla Rodrigues Postgraduate Institute of Medical Education & Research (PGIMER), Chandigarh - Pallab Ray and Vikas Gautam Sanjay Gandhi Postgraduate Institute of Medical Sciences (SGPGI), Lucknow - TN Dhole Book 1.indb 187 05-Jun-2012 2:36:00 PM WHO South-East Asia Journal of Public Health 2012;1(2):182-188188 Antibiogram of S. typhi and S. paratyphi A in India Indian Network for Surveillance of Antimicrobial Resistance Group References Crump JA, Luby SP, Mintz ED. 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