Clinical Profile and Epidemiology of Campylobacter Associated Diarrhea Among Children in New Delhi, India

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1 Interntionl Journl of Enteric Pthogens Int J Enteric Pthog August;4(3):e Originl Article doi /ijep Clinicl Profile nd Epidemiology of Cmpylobcter Associted Dirrhe Among Children in New Delhi, Indi Roumi Ghosh 1*, Been Uppl 2, Prbhv Aggrwl 2, Anit Chkrvrti 2, Annd Prksh Dubey 3 1 Deprtment of Microbiology, Institute of Post Grdute Medicl Eduction nd Reserch,West Bengl, Indi 2 Deprtment of Microbiology, Muln Azd Medicl College, New Delhi, Indi 3 Deprtment of Peditrics, Muln Azd Medicl College, New Delhi, Indi *Corresponding Author: Roumi Ghosh, Tel: ; Emil: roumighosh@gmil.com Published Online My 1, 2016 Keywords: Cmpylobcter, Dirrhe, Polymicrobil infection, Peditrics Abstrct Bckground: Cmpylobcter, well-known enteropthogen mong children shows vrible clinicl presenttions. Age groups nd sesonl distribution is dependent on geogrphicl position. Objectives: To explore clinicl mnifesttions nd sesonl vrition of Cmpylobcter infection nd to study its importnce s enteric pthogen mong children. Ptients nd Methods: Two hundred five children ( 12 yers ge) hving cute dirrhe s cses nd 100 children without from dirrhe were tken s control. All the fecl smples were processed for Cmpylobcter species by culture on to modified chrcol cefoperzone deoxycholte gr nd Skirrow s Columbi blood gr medi. Detection of Cmpylobcter specific ntigen in fecl smples ws lso done by enzyme-immuno ssy. Results: A totl of 32 (15.61%) fecl smples of children with dirrhe hd positive results for Cmpylobcter spp. Among them 31.25% cses hd polymicrobil infections. Children below 1 yer were most commonly (18.96%) ffected by the infection. The orgnism ws isolted throughout the yer with higher isoltion rtes during summer nd monsoon months. Wtery dirrhe ws significntly more common in the Cmpylobcter infected cses. Conclusions: Appliction of ntigen ssy increses detection rte of Cmpylobcter enteritis cses, which ws significntly higher thn the control group (P <.05). Specific clinicl profile could not be ssocited with this infection which, indictes need of microbiologicl dignosis of this pthogen for ntibiotic therpy. Received December 22, 2016; Revised Jnury 5, 2016; Accepted Jnury 10, 2016 Bckground Cmpylobcter species re primrily zoonotic, with wide vriety of wild nd domestic nimls, especilly birds implicted s reservoir. They minly cuse foodborne gstroenteritis following ingestion of chicken, rw milk, untreted wter nd contct with pets, especilly household live chickens. Incresing trend of Cmpylobcter infections hve been seen in developed countries for yers nd it ccounts for one of the most common bcteril cuses of dirrhe, with n incidence round 10% mong dirrhe ptients. 1 In recent times there re mny reports from developing countries describing Cmpylobcter jejuni nd Cmpylobcter coli s importnt enteropthogen during first 5 yers of life with isoltion rte rnging from 10% to 46%. 2-5 Though, the epidemiology, clinicl presenttions nd microbil profile re not similr in these two economicl world territories. 6 Appliction of newer, sensitive moleculr dignostic methods beside culture might ply role behind the incresing detection rte of this fstidious, microerophilic orgnism. 7,8 It is present s colonized gstrointestinl flor nd lso s symptomtic crrier commonly in children, but it cuses severe gstroenteritis mong young children <2 yers old, elderly or immunocompromised ptients nd my require ntibiotic therpy. Thus, vrious virulence mrkers hd been studied to prove its pthogenicity. 9 Cliniclly, Cmpylobcter infection is indistinguishble from cute gstrointestinl infections produced by other bcteril pthogens. In some ptients, the dirrhe is miniml nd bdominl crmps nd pins re the predominnt fetures; this cn led to mistken dignosis of cute bdomen nd unnecessry lprotomy. Fluoroquinolones nd mcrolides re min prts of tretment used for this infection; however, mrked increse in fluoroquinolone resistnce nd presence of high level zithromycin resistnce in Cmpylobcter isoltes re becoming thret in In- Copyright 2016 The Author(s); Published by Alborz University of Medicl Sciences. This is n open-ccess rticle distributed under the terms of the Cretive Commons Attribution-NonCommercil 4.0 Interntionl License ( which permits copy nd redistribute the mteril just in noncommercil usges, provided the originl work is properly cited.

2 di. 10,11 The most importnt post-infectious compliction of C. jejuni infection is the Guillin-Brré syndrome tht ffects 1 2 persons per popultions in the United Sttes ech yer. 6 Objectives The present study ws designed to investigte clinicl mnifesttions nd epidemiology of Cmpylobcter infection nd to study the importnce of C. jejuni s enteric pthogen mong children. Ptients nd Methods The study ws conducted with ethicl permission in the Deprtment of Microbiology, Muln Azd Medicl College nd Deprtment of Peditrics, LN hospitl, New Delhi for 2 consecutive yers. The study group included 205 ptients ged 12 yers or below hving cute dirrhe (<14 dys durtion) dmitted in dirrhe wrd of the hospitl. A totl of 100 ge nd sex mtched children without ny gstrointestinl complints were tken s control. After proper counselling, n informed consent ws tken from the prents/gurdins/person ttending the study subject. Detiled personl history, dirrhel episode nd ssocited signs nd symptoms were recorded on pre-designed pro form. Exclusion Criteri Children on ntimicrobil therpy were excluded from the study. Smple Collection nd Trnsport Stool smples were requested from ll ptients nd controls who fulfilled the inclusion criteri. Proper instructions were given regrding collection of specimen i.e. freshly pssed feces to be collected in clen, wide mouth, screw cpped plstic continer nd trnsported to microbiology lbortory within 2 hours of collection. In cse of dely of more thn two hours, smples were trnsported in Cry Blir medium/ buffered glycerol sline. Exmintion of Smple The stool specimen ws processed s follows: Culture: All fecl smples were processed for Cmpylobcter species by direct inocultion nd fter enrichment in BHI broth on modified chrcol cefoperzone deoxycholte gr (CCDA) (Oxoid ) nd Skirrow s Columbi blood gr medi with Cmpylobcter growth supplement nd Cmpylobcter selective supplement (Butzler) (Oxoid ) contining bcitrcin ( IU), cycloheximide (25 mg), colistin sulfte (5000 IU), cephzolin sodium (7.5 mg) nd novobiocin (2.5 mg). The pltes were incubted long with control strin of C. jejuni for 48 hours t 42 C under microerophilic conditions (5% O 2, 5% CO 2, 2% H 2, nd 88% N 2 by volume) generted by AN- OXOMAT AN2OP system. Pltes were exmined fter 48 hours nd in cse of no growth re-exmined fter 72 hours nd then gin fter 7 dys of incubtion. 12 Suspected colonies of Cmpylobcter grown were confirmed by oxidse test, ctlse test, hippurte hydrolysis, hydrolysis of indoxyl cette, growth on 1% glycine nd 1.5% NCl nd susceptibility to cefoperzone (30 ug), nlidixic cid (30 ug) nd cephlothin (30 ug) s per stndrd techniques. 13 All smples were exmined by wet mount for the presence of prsites nd inoculted on severl dignostic medi Such s McConkey s gr, xylose lysine deoxycholte gr, blood gr nd bile slt gr directly nd fter enrichment in selenite F broth nd lkline peptone wter for the isoltion of conventionl enteropthogens. Chrcteristic colonies were identified by bsed on colony chrcteristics, biochemicl rections nd gglutintion test with respective ntiser. Detection of Cmpylobcter Antigen in Stool Smples ProSpecT TM Cmpylobcter Microplte Assy (Oxoid Ltd, UK) ws used for qulittive detection of Cmpylobcter specific ntigen in fecl smples s per mnufcturers instructions. Sttisticl Anlysis All dt obtined ws nlyzed using SPSS sttisticl softwre. Chi-squre test with Ytes correction, Fisher exct test were used to compre the results, wherever pplicble. Results A totl of 32 (15.61%) children with dirrhe hd positive results for Cmpylobcter ntigen mong which 15 smples yielded growths on culture medi. The detection rte from the controls without dirrhe ws 4%. The difference between the isoltion rtes ws sttisticlly significnt (P<0.05). All the isoltes were identified s C. jejuni. Among 32 positive cses, 22 (68.75%) children were infected with C. jejuni s sole pthogen, wheres 10 (31.25%) cses hd polymicrobil infections. Most common pthogen isolted long with C. jejuni ws Vibrio cholere O1 Ogw (15.62%) followed by enteropthogenic Escherichi coli (EPEC) 9.4% nd rotvirus in 3.12% cses. One cse hd triple infection with C. jejuni, EPEC nd rotvirus. Men ge of children with Cmpylobcter infection ws 9 months with pek incidence (18.96%) in children below 1 yer (Tble 1). Mles were more frequently (1.5:1) infected thn femles. Sesonl distribution of Cmpylobcter infection is presented in Figure 1. A totl of 71.87% isoltes were detected during the summer nd monsoon months of which highest recovery ws in the month of July (25%). Wtery dirrhe ws significntly more common thn inflmmtory dirrhe in the Cmpylobcter infected cses (Tble 2). Fever ws observed in 59% cses infected with Cmpylobcter lone, but it ws significntly more (90%) ssocited with mixed infection, P =.0402 (Tble 3). Abdominl pin nd vomiting were eqully common in both the groups (75% nd 90.62%). In contrst, dehydrtion ws observed in 59% cses infected with Cmpylobcter s sole pthogen, but ws much more common mong mixed infection cses (90%), P <.05, which ws 2 Interntionl Journl of Enteric Pthogens Volume 4, Issue 3, August 2016

3 sttisticlly significnt. No. of Cmpylobcter Detected Figure 1. Sesonl Distribution of Cmpylobcter Infection. Discussion A slightly higher isoltion rte (15.61%) ws found in the present study compred to previous studies from Indi where isoltion rte vried from 7%-13.5% mong cute dirrhel cses. 2,14 Vrition in results my be due to the use of different techniques of detection. ELSIA s new method ws used in the present study, while other studies used only culture s method of detection. Studies from neighbouring countries like Bngldesh, Pkistn nd Chin reveled n isoltion rte of 11.8%, 18% nd 25.5% nd very high (62%) prevlence hs been reported from Thilnd. 3,4,15,16 Another feture observed in this study ws the high percentge of mixed infections of C. jejuni with other known enteropthogens, mtching with prior studies (14). Bhdr et l reported Vibrio cholere O1 Ogw s the commonest co-pthogen of Cmpylobcter in Kolkt. 17 High number of co-infection with dirrhegenic E. coli nd rotvirus mong Cmpylobcter positive cses ws found in Vellore. 18 Therefore our study reffirms the previous finding tht polymicrobil infection is common in Cmpylobcter ssocited dirrhe in developing countries. Similr to our study, dominnce of C. jejuni infection mong children less thn 1 yer (18.96%) ws seen in previous studies conducted in Kolkt nd Bngldesh. 17,19 In developing countries, high-level exposure to the orgnism erly in life leding to the grdul development of protective immunity restrict symptomtic infection fter the ge of 2 yers nd durtion of intestinl excretion declines with ge. In order to trce the source of Cmpylobcter infection in infnts, studies pointed out drinking well wter, eting home prepred fruits or vegetbles, exposure to pet with dirrhe, visiting or living in frm, ridden in shopping crt next to met or poultry shop potentil risk fctors. 20,21 Indin studies documented poultry nd cttle s mjor reservoir of this infection. 22,23 Brest feeding, drinking purified wter, wshing hnds fter contct with pets, environmentl seprtion of household livestock, voiding foreign trvel re the protective mesures to stop trnsmission. Dt regrding in exposure to nimls or poultry of the children enrolled in our study ws not vilble; therefore, further risk ssessment could not be crried out. Though Cmpylobcter infection previled throughout the yer, higher rte of isoltion (71.87%) ws seen during the summer nd monsoon months of which highest recovery ws in the month of July (25%). Similr sesonl vrition ws observed in Kolkt. 17 Wtery dirrhe ws found to be significnt- Tble 1. Age Distribution of Cmpylobcter Isoltes Both From Cses nd Control Cses With Acute Dirrhe Controls Without Dirrhe Age (y) No. of Ptients Studied Positive for Cmpylobcter, No. (%) No. of Controls Studied Positive for Cmpylobcter, No. (%) < (18.96) 58 0 (0) (16.22) 19 1 (5.26) (11.11) 13 2 (15.38) > (4) 10 1 (10) Totl (15.61) (4) There ws significnt difference between the isoltion rtes of Cmpylobcter spp. in cses nd controls in totl ws significnt (P =.0023) but between ech ge group ws not significnt (P >.05). Tble 2. Clinicl Presenttion of Cmpylobcter Infection Findings Cmpylobcter Positive Cses, n = 32 (%) Cmpylobcter Negtive Cses, n = 173 (%) Wtery dirrhe 25 (78.12) 66 (38.15) Inflmtory dirrhe 7 (21.87) 35 (20.23) Fever (>100 F) 22 (68.75) 89 (49.13) Abdominl pin 24 (75) 40 (23.12) Vomiting 29 (90.62) 53 (30.63) Dehydrtion 22 (68.75) 52 (30.05) Cough nd coryz 3 (9.37) 27 (15.61) Convulsion 2 (6.25) 19 (10.98) P < Interntionl Journl of Enteric Pthogens Volume 4, Issue 3, August

4 Tble 3. Comprison of Clinicl Findings Between Children With Cmpylobcter Infection Alone nd Cmpylobcter Infection With Other Pthogens Findings Ptients Infected With Cmpylobcter Alone, n=22 (%) Ptients Infected With Cmpylobcter nd Other Pthogens, n=10 (%) Wtery dirrhe 17 (77.27) 8 (80) Inflmmtory dirrhe 5 (22.72) 2 (20) Fever (>100 F) 13 (59.1) 9 (90) Abdominl pin 16 (72.72) 8 (80) Vomiting 19 (86.36) 10 (100) Dehydrtion 13 (59.1) 9 (90) Cough nd coryz 3 (13.64) 0 (0) Convulsion 1 (4.54) 1 (10) P <.05. ly ssocited thn inflmmtory dirrhe in the Cmpylobcter infected cses (71.12% vs %; P <.0001). Though study from Pkistn found blood nd mucus in 90% of dirrhel stools tht yielded C. jejuni, Bhdr et l noticed wtery dirrhe in 97.6% of C. jejuni/coli infected cses. 4,17 In comprtive nlysis to investigte the difference in clinicl presenttion between the ptients infected with Cmpylobcter lone (n = 22) nd those infected with multiple pthogens (n = 10), fever nd dehydrtion ws found to be more common in mixed infections. Though Tribble et l evluted sensitivity nd specificity of vrious clinicl presenttions nd stool chrcteristics s modlity to dignose Cmpylobcter infection, we could not ssocite ny clinicl pttern specific for Cmpylobcter enteritis. 24 In the present study Cmpylobcter comprised significnt percentge of enteropthogens mong children in Indi, which indictes requirement of routine identifiction of this pthogen. Moreover, clinicl fetures could not be used to dignose of cmpylobcteriosis per se becuse of the non-specific nture of the symptoms. Conflict of Interest Disclosures None. References 1. Centers for Disese Control nd Prevention Vitl signs: incidence nd trends of infection with pthogens trnsmitted commonly through food Foodborne Diseses Active Surveillnce Network, 10 U.S. sites, MMWR Morb Mortl Wkly Rep. 2011;60: Jin D, Sinh S, Prsd KN, Pndey CM. Cmpylobcter species nd drug resistnce in north Indin rurl community. Trns R Soc Trop Med Hyg. 2005;99: doi: /j. trstmh Ahmed D, Hoque A, Elhi MS, Endtz HP, Hossin MA. Bcteril etiology of dirrhoel diseses nd ntimicrobil resistnce in Dhk, Bngldesh, Epidemiol Infect. 2012;140: doi: /s Ali AM, Qureshi AH, Rfi S, et l. Frequency of Cmpylobcter jejuni in dirrhoe/dysentery in children in Rwlpindi nd Islmbd. J Pk Med Assoc. 2003;53(11): Wng SC, Chng LY, Hsueh PR, Lu CY, Lee PI, Sho PL. Cmpylobcter enteritis in children in northern Tiwn- - 7-yer experience. J Microbiol Immunol Infect. 2008;41(5): Coker AO, Isokpehi RD, Thoms BN, Amisu KO, Obi CL. Humn cmpylobcteriosis in developing countries. Emerg Infect Dis. 2002;8: doi: /eid Ghosh R, Uppl B, Aggrwl P, Chkrvrti A, Jh AK, Dubey AP. A comprtive study of conventionl nd moleculr techniques in dignosis of cmpylobcter gstroenteritis in children. Ann Clin Lb Sci. 2014;44(1): Kumr A, Kumr S. Comprtive Anlysis of Culturl nd PCR Bsed Assys for Detection of Cmpylobcter spp. in Humn Stool Smples. Proc Ntl Acd Sci Indi Sect B Biol Sci. 2015;85(3): doi: /s Rizl A, Kumr A, Vidyrthi AS. Prevlence of pthogenic genes in Cmpylobcter jejuni isolted from poultry nd humn. Internet J Food Sfety. 2010;12: Ghosh R, Uppl B, Aggrwl P, Chkrvrti A, Jh AK. Incresing ntimicrobil resistnce of Cmpylobcter jejuni isolted from peditric dirrhe cses in tertiry cre hospitl of New Delhi, Indi, J Clin Dign Res. 2013;7: doi: /jcdr/2013/ Mukherjee P, Rmmurthy T, Mitr U, Mukhopdhyy A. Emergence of high-level zithromycin resistnce in Cmpylobcter jejuni isoltes from peditric dirrhe Ptients in Kolkt, Indi. Antimicrob Agents Chemother. 2014;58(7): Fitzgerld C, Nchmkin I. Cmpylobcter nd Arcobcter. Wshington, DC, USA: ASM Press; 2007: Blser MJ, Berkowitz ID, LForce FM, Crvens J, Reller LB, Wng WLL. Cmpylobcter enteritis: clinicl nd epidemiologic fetures. Ann Intern Med. 1979;91(2): Mukherjee P, Rmmurthy T, Bhttchry M, Rjendrn K, Mukhopdhyy A. Cmpylobcter jejuni in Hospitlized Ptients with Dirrhe, Kolkt, Indi. Emerg Infect Dis. 2013;19(7): Desheng L, Zhixin C, Boun W. Age distribution of dirrhoel nd helthy children infected with Cmpylobcter jejuni. J Trop Med Hyg. 1992;95: Tribble DR, Bqr S, Png LW, et l. Dignostic pproch to cute dirrhel illness in militry popultion on trining exercises in Thilnd, region of cmpylobcter hyperendemicity. J Clin Microbiol. 2008;46(4): Bhdr RK, Dutt P, Bhttchry SK, Dutt SK, Pl SC, Nir GB. Cmpylobcter species s cuse of dirrhoe in children in Clcutt. J Infect. 1992;24(1): Ajjmpur SSR, Rjendrn P, Rmni S, et l. Closing the dirrhoe dignostic gp in Indin children by the ppliction of moleculr techniques. J Med Microbiol. 2008;57(11): Albert MJ, Fruque AS, Fruque SM, Sck RB, Mhlnbis D. Cse control study of enteropthogens ssocited with childhood dirrhe in Dhk, Bngldesh. J Clin Microbiol. 4 Interntionl Journl of Enteric Pthogens Volume 4, Issue 3, August 2016

5 1999;37: Fullerton KE, Ingrm LA, Jones TF, et l. Spordic Cmpylobcter infection in infnts: popultion-bsed surveillnce csecontrol study. Peditr Infect Dis J. 2007;26(1): doi: /01.inf Tenkte TD, Stfford RJ. Risk fctors for cmpylobcter infection in infnts nd young children: mtched csecontrol study. Epidemiology nd Infection. 2001;127(3): Rjendrn P, Bbji S, George AT, Rjn DP, Kng G, Ajjmpur SS. Detection nd species identifiction of Cmpylobcter in stool smples of children nd nimls from Vellore, south Indi. Indin J Med Microbiol. 2012;30(1): doi: / Chttopdhyy UK, Rshid M, Sur SK, Pl D. The occurrence of cmpylobcteriosis in domestic nimls nd their hndlers in nd round Clcutt. J Med Microbiol. 2001;50: doi: / Tribble DR, Bqr S, Png LW, et l. Dignostic pproch to cute dirrhel illness in militry popultion on trining exercises in Thilnd, region of cmpylobcter hyperendemicity. J Clin Microbiol. 2008;46(4): Interntionl Journl of Enteric Pthogens Volume 4, Issue 3, August