Int.J.Curr.Microbiol.App.Sci (2014) 3(9) 78-84 78

Original Research ArticleEvaluation of microbiological quality of ice creams marketed inKolhapur city, Maharashtra, India A.S.Jadhav* and P.D.Raut Department of Environmental Science, Shivaji University, Kolhapur, India *Corresponding author A B S T R A C TIntroductionIcecreamisthemajordairyproductwhich isoneofthefavoritefooditemsinlarge segmentsofthepopulation.Itisa nutritionallyenrichedfrozendairyproduct consumedbyallagegroupsparticularly childrenmostlyduringsummer(Sharifet al., 2005). Ice cream is a product of mixture consistingmilk,sweeteningandstabilizing agentstogetherwithflavoringandcoloring matter(Graff-Johnson,1974).Different ingredients like milk, cream, evaporated orcondensedmilk,driedmilk,coloring material,flavors,fruits,nuts,sweetening agents, egg products and stabilizers are used inicecreampreparation.Also,thereare many parameters which should be controlled accurately during processing to obtain a high qualityicecreamwiththerequiredtaste, flavor,viscosity,consistencyand appearance(Yamanetal.,2006).Duetoits composition,itcanharbormanypotent pathogens.Handlingandstorageconditions ISSN: 2319-7706 Volume 3 Number 9 (2014) pp. 78-84 http://www.ijcmas.com Ke ywor dsIcecream, Microbiological quality, Escherichia coli, Salmonella, Staphylococcus aureus. Icecreamisafavoritefooditemcommonlyconsumedduringsummerinallage groups.Duetoitscomposition,itcanharbormanypotentpathogens.Mostice creamsbecomecontaminatedwithmicrobesduringproduction,transit,and preservation.Suchcontaminatedfoodproductcanberesponsibleforfoodborne infectionsinchildren,elderlypeopleandimmune-suppressedpatients.Therefore, thestudywasconductedtoevaluatethebacteriologicalqualityoficecreamssold indifferentareasofKolhapurcity,Maharashtra.Randomlycollectedicecream sampleswereanalysedforTotalViableCount(cfu/g),Coliformcount(per100 ml),detectionofE.coli,Shigellaspecies,SalmonellaspeciesandStaphylococcus aureus.Allsamplesshowedpositivegrowthofbacteriarangingfrom1.2x103 to 8.0 x 107 cfu/g. The study revealed that all the samples showed positive growth of coliformbacteriawhile40%icecreamsampleswerecontaminatedbytheE.coli, 33%samplesshowedpositivegrowthofSalmonella,40%showedgrowthof Staphylococcusaureusand53%Shigellaspecies.Itindicatesthelackof maintenanceofhygienicconditionsduringpreparation,preservationorservingof ice cream. Hence, the present study is undertaken to assess potential of health risk to public. Int.J.Curr.Microbiol.App.Sci (2014) 3(9) 78-84 79 aresomeofthesevariablesbecausethey affectthephysical,chemicalandmicrobial qualityofproduct(MarshallandArbuckle, 1996,Warke et al., 2000, Champagne et al., 1994, Daniels et al. 2002). Most ice creams becomecontaminatedwithmicrobesduring production,transitandpreservation.The possibilityofacquiringfoodbornediseases byinfant,children,elderlypeopleand immune-suppressedpatientsismoredueto contaminatedicecreams.Duringlastfew decades,ithasbeenseenthatconsumption ofcontaminatedicecreamsareresponsible fordiseaseoutbreaksinmanycountries fromAsia,Europe,andNorthAmerica (Chug,1996;Djureticetal.,1997;Digrak andOzcelik,1991).Sincetheconsumption ofice-creamishigheramongchildrenof vulnerableagegroups,thereisaneedto maintainahighmicrobiologicalsafety standard (Champagne et al., 1994).It is a known fact that relatively low storage temperatureandpasteurizationstepsduring processingoficecreamsareconsideredto eliminatemostofthepathogenic microorganisms.However,theadditionof contaminated ingredients, improper handling andtheuseofimproperstorage temperaturesarethefactorsthatcontribute tothepotentialhazardsofthefinished productsespeciallyduringprocessingafter pasteurizationprocesses(Vermaetal., 1992). Bacteriologicalqualityoficecreamreflects hygienicpracticeinproductionandisan indicationofgoodpracticeinproduction.It isanindicationoffoodwhichdetermines thebacteriologicalqualityoficecreams (AmbilyandBeena,2012).Hence,the present study was taken up to determine the bacteriologicalqualityoficecream marketedinKolhapurcityandtoassessthe potentialofthesefrozenproductstopose risk to public health. Materials and MethodsTherearedifferenttypesoficecreams availableinmarket.Thepresentstudyaims todeterminethemicrobiologicalqualityof ice-creamsmarketedinKolhapurcity, Maharashtra,India.Themicrobiological testsconductedfortheicecreamsamples wereTotalViableCount,Coliformscount anddetectionofpathogenicmicro organisms. Forstudypurpose,icecreamsampleswere divided in three categories like branded, non branded and self-made ice creams. These ice creamsampleswerecollectedfrompopular icecreamshopsaswellasroadvendorice creams from local places like public gardens andicecreamparlors.Thesampleswere analyzedinthemonthofNovember,2013 andinFebruary,2013.Theicecream sampleswerecollectedandanalyzedtwice. Duringtheanalysissampleswerecollected andpreservedatlowertemperaturefor further analysis.TotalViableCountsweredeterminedby incorporating10-folddilutedsamplesinto moltenNutrientAgarplates.Theplates wereallowedtoset andthenincubatedat 37Cfor48h.Coliformcountsoftheice cream samples were determined by Multiple TubeFermentationTechnique(Fengetal., 2007).Thetubesshowingacidandgas productions were streaked on Mac Conkey s agarplatetoobtaindiscretecolonies. Typicallactosefermentingcolonies(n=5) were picked up and identified as Escherichia colibystandardbiochemicalreactions (Colleeetal.,1996).Inaddition,the sampleswerealsoscreenedforenteric pathogenslikeSalmonellaspecies,Shigella speciesandStaphylococcusaureus.The sampledilutionswerespreadonselective anddifferentialmediabyspreadplate method.Then,theplateswereincubatedat Int.J.Curr.Microbiol.App.Sci (2014) 3(9) 78-84 80 37C for 24 hr. After incubation plates were observed for presence of pathogenic bacteria Results and DiscussionAccordingtoIndianFoodSafetyStandards andRegulations(2011),TotalVolatile Count(TVC)oficecreamshouldnot exceed 2, 50,000 cfu/g. In the present study, it was observed that the TVC for all the self madeicecreamswas(2x105 to7x105 cfu/g) and for branded (1.2 x 103 to 7.0 x103 cfu/g).However,noneoftheroadvendor icecreamsamplesshowedTVCwithinthe standards(4.3x107 to8.0x107 cfu/g).A studywasconductedregarding bacteriologicalqualityoflocalmadeopen scoopicecreamssoldbystreethawkersin differentareasofJalandharcity,Punjab whichshowedheavycontaminationof bacteriarangingfrom0.1109 to10.2109 cfu/g(Kumaretal.,2011).Themicrobial countsinicecreamsamplesmayhave resulted from inadequate processing, such as initial improper cooling of the hot ice cream mix,whichmayleadtomultiplicationof microorganismspresentinicecream immediatelyafterpasteurization(Ojokoh, 2006). It is also indicated that high microbial count occurrence in virtually sterile mix may occurwhencooledslowlyatatemperature conducivetosporegermination(Beck, 1973).Thepresenceofcoliformorganismsis takenasanindicationthatotherpathogenic organismsmayalsopresentinthesample (Trabulsietal.,2002).Duringthepresent studyallthesamplesshowedpresenceof coliformbacteria.Themaximumcoliform countwas400cfu/gmintheroadvender s ice cream sample and minimum count was 9 cfu/gminthebrandedsamples.Coliforms beingnon-sporeformersshouldbe susceptibletopasteurization.Theirpost pasteurizationpresenceinicecreams indicatesfaultyheatprocessduring preparation.Also,theotherreasonsfor contaminationmayeithercomefromwater, lackofpersonalhygieneoftheicecream manufacturer, utensils used for ice cream. It wasobservedthatcoliformbacteriawere presentinallsamples.Similar,findings werealsoreportedbyAnuranjinietal., (2008)alsoreportedtheincidenceofhigh coliformcountinicecreamsmarketedin Mangalore town.Further,theisolationandconfirmationof E.coliwasdonebyIMVICtest.Itwas observedthatabout40%samplesshowed presenceofE.coliwithmaximumE.coli count(4cfu/gm)inroadvendericecream sample. AstudyconductedbyAhmedand Shakoori(2002)reported640-683E.coli colonies per 100 ml of drinking water. Such watercanbethemajorsourceof contaminationwhenusedforpreparationof icecreams.Oncetheicecreambecome contaminated,freezingtemperaturelater couldnotmaketheproductsafer(Jay, 1996). About40%icecreamsamplesshowed presenceofStaphylococcusspeciesmostof themwerepurchasedfromroadvendorsin thecity.Themaximumoccurrenceofthese organismswasinroadvendersicecream samples i.e. 7 cfu/gm. It is assumed that the possiblesourcesofthisorganisminice creamhands,skinandclothingofhandlers whereitiscommonlyfound;(Hobbsand Golbert,1982).Thereareothersourceslike coughing,talkingandsneezingwhich produce droplets. These droplets could settle onicecreamduringtransportation,storage and retailing (Ojokoh, 2006).Moreover,Salmonellaspecies(1 8cfu/g) andShigellaspecies(1 5cfu/g)werealso isolatedfromsomeicecreamsamples.It wasobservedthatabout40%samples Int.J.Curr.Microbiol.App.Sci (2014) 3(9) 78-84 81 showedpresenceofSalmonellaspecies while53%samplesshowedpresenceof Shigella species.Salmonella is still the most importantagentcausingacutefoodborne diseases(Tood,1997).Consumptionofice creamcontaminatedwithenteropathogenic bacteriasuchasSalmonellahasbeenthe causeofseveraldiseaseoutbreaks (Hennessy et al., 1996).Contaminatedicecreamscouldbe responsibleforfoodpoisoninginmasses. Therefore,thepresenceofpossible pathogenicorganismsintheanalyzedice creamsamplesshouldbeviewedwith concern by the consumers. Not only bacteria but some viruses and fungal species are also responsible for spreading diseases. It should alsobenotedthatsomespeciesof Aspergillusareknowntoproducepowerful mycotoxinswhichareharmfultoman,thus their occurrence in ice cream is undesirable.Itisneededtocontinuouslymonitorthe foodproducts.Sanitaryeducationthrough trainingandworkshopstoproducers, distributorsandretailersduringprocessing andsaleoftheproductcanbethepossible remedy to avoid such problems.Table.1 Collection of different categories of ice creams from Kolhapur cityS. No.Roadvendorice-creams (X) (n= 6) Selfmadeicecreams from parlors (Y)(n= 5) Brandedicecreams (Z) (n= 4) 1(X1)(Y1)(Z1) 2(X2)(Y2)(Z2) 3(X3)(Y3)(Z3) 4(X4)(Y4)(Z4) 5 (X5)( Y5)---- 6(X6) ------- Total samples=15

Table.2 Media used for isolation of microorganisms S. No. ParametersMedia used 1.Total Plate CountNutrient agar 2.Coliform GroupMac Conkeys broth 3.Salmonella speciesBismuth Sulphite agar 4.Staphylococcus aureus Mannitol salt agar 5.Shigella species.Deoxycholate citrate agar 6.E. coliEMB agar Int.J.Curr.Microbiol.App.Sci (2014) 3(9) 78-84 82

Table.3 Types of microorganisms and their percentage (%) occurrence in ice cream samples S. No.TypesofmicroorganismsPercentage (%) 1. Coliform Count 100 2. Staphylococcalspecies (cfu/g) 40 3. Salmonellaspecies(cfu/g) 33 4. E. coli (cfu/g) 40 5. Shigella Species(cfu/g) 53 Table.4 Bacteriological quality of ice creams marketed in Kolhapur City,Maharashtra, IndiaS. No. Typeofice cream TVC (cfu/g) Coliforms Count(Per100 ml ) Staphylococcalspecies(cfu/g) Salmonella species(cfu/g) E. coli (cfu/g) Shigella species (cfu/g) 1. Road vendor ice-creams (X1) 8.0 x 107 160482-- 2. Road vendor ice-creams (X2) 7 .1 x 107 61--342 3. Road vendor ice-creams (X3) 4.8 x 107 245--34 4. Road vendor ice-creams (X4) 6.5 x 107 1807----2 5. Road vendor ice-creams( X5) 5.8 x 107 350-----45 6. Road vendor ice-creams (X6) 4.3 x 107 4003731 7. Self made ice creams (Y1) 2 x 105 24--5---- 8. Self made ice creams (Y2) 7 x 105 10-------- 9. Self made ice creams (Y3) 3 x 105 12------4 10. Self made ice creams (Y4) 2 x 105 17-------2 11. Self made ice creams (Y5) 3.5 x 105 94------ 12. Branded ice creams (Z1) 1.2 x103 10-------- 13. Branded ice creams (Z2) 4.4 x103 12----2-- 14. Branded ice creams( Z3) 7.0 x103 63------ 15. Branded ice creams (Z4) 2.8 x103 5--1--1 Int.J.Curr.Microbiol.App.Sci (2014) 3(9) 78-84 83 AcknowledgementAuthorsarethankfultoDepartmentof Environmental Science, Shivaji University KolhapurandauthoritiesofShivaji University,Kolhapur,Maharashtra,for providingfacilitiestocarryoutthe analysis work.References Ahmed, K., Shakoori, A.R. (2002). Vibrio choleraeElTor,Ogawa01asthe mainetiologicalagentofthetwo majoroutbreaksofgastroenteritisin the Northern areas of Pakistan, Gilgit. Pakistan J. Zool., 34: 73 80. Ambily,R.,Beena,A.K.,(2012).Bacteriologicalqualityofice-cream marketedinThrissurtown,Kerala, India,VeterinaryWorld,5(12):738741. Anuranjini,C.,SebastianGeethu,B., Dhanashree,B.(2008).Bacteriological analysis of ice creams fromMangalore,SouthIndia.Indian J. Med. Res., 127: 91 92. Beck,E.(1973).Hygienicaspectofthe productionandmarketingofsoftice cream(II).BackerandKonditor,27: 297 298. Champagne,C.P.,Laing,R.R.Roy,D., Mafu,A.A.,(1994).Psycrotrophsin dairy products: Their effects and their control. Crit. Rev. Food Sci. Nutr., 34: 1 30. Collee,J.G., Miles,R.S.,Watt,B.(1996). Tests for the identification of bacteria. In: Collee J.G., Fraser A.G., Marmion B.P.,SimmonsA,(Eds).Mackie& McCartneypracticalmedical microbiology,14th ed.London: Churchill Livingstone, p. 131 149. Feng,P.,Weagant,S.D.,Grant,M.A. (2007).U.S.FoodandDrug Administration.Enumerationof Escherichiacoliandthecoliform bacteria. In: Bacteriological analytical manual online. Chapter 4.FoodSafetyandStandardsAuthorityof India.(2011).Foodsafetyand Standards(Foodproductstandards and food additives) regulation, part II: 498. Graff-Johnson,C.D.,(1974).Themicro biologicalexaminationoficecream manufacturedinGhana.GhanaJ. Agri. Sci., 7: 227 230. Hennessy,T.W.,HedbergC.W.,Slutsker L., White K.W., Verma, G.S., Besser-Wiek,M.S.,Moen,M.E.,John, F.B.S.,Coleman,W.W.,Edmonson, L.M.,MacDonald,K.L.,Osterholm, M.T.(1996).Anationaloutbreakof Salmonella enteritidis infections from icecream.N.Engl.J.Med.,334: 1281 1286. Hobbs,B.C.,Golbert,J.R.(1982).Food PoisoningandFoodHygiene.4th Edn.,EdwardArnoldLimited, London, p. 366. Jay,J.M.(1996).ModernFood Microbiology,4th Edn.,Champman and Hall Inc, New York. ChugK.(1996).Salmonellaoutbreak fromicecream.IndianPediatr.,33:976 977.Kumar,H.,Wadhwa,G.,Palaha,R., Gandhi,R,Singh,S.(2011). Microbiologicalqualityanalysisof icecreamssoldbystreethawkers:A casestudyofJalandharcity,India. Int. J. Food Safety.,13: 164 169.Digrak,M.,Ozcelik,S.,(1991).The microbiologicalqualityoficecream soldinElazigcity.Food,16:195200. Marshall,R.T., Arbuckle W.S. (1996). Ice Cream,5th Edn.,Chapman&Hall, New YorkDaniels,N.A.,Mackinnon,L.,Rowe S.M.,BeanN.H.,GriffinP.M.,Mead Int.J.Curr.Microbiol.App.Sci (2014) 3(9) 78-84 84 P.S.(2002).Foodbornedisease outbreaksinUnitedStatesschools. Pediatric. Infect. Dis. J., 21: 623 628.Ojokoh,A.O.(2006).Microbiological examinationoficecreamsoldin Akure,Pakistan.J.Nutrition,5(6): 536 538. Sharif,N.,Ghanghesh,K.S.,Gnan, Y.A.S.,Rahouma,A.(2005). Bacteriological quality of ice cream in Tripoli-Libya.FoodCont.,17:637641. Djuretic,T.,Wall,P.,Nichols,G.(1997). Generaloutbreaksofinfectious intestinal disease associated with milk anddairyproductsinEnglandand Wales:1992to1996.Commun.Dis. Rep. Rev. CDR Rev., 7: R41 R45. Tood,E.C.(1997).Epidemiologyoffood bornediseases:aworldwidereview. WorldhealthStatQ.,50(1 2): 3050. Trabulsi,L.R.,Keller,R.,Tardelli Gromes,T.A.(2002).Typicalanda typical enteropathogenic Escherichia coli. Emerging Infect. Dis., 8(5): 508513. Verma,G.S.,(1992).Bacteriaqualityof someingredientsusedinthe manufactureofice-cream.IndianJ. Anem. Res., 6: 27 50. Warke,R.A.,Kamat,M.,Kamat,P., Thomas,P.(2000).Incidenceof pathogenicpsychrotrophsinice creamssoldinsomeretailoutletsin Mumbai,India.FoodControl,11: 77 83.Yaman, H., Elmali M., Ulukanli Z., Tuzcu M.,Genctave,K.(2006).Microbial qualityoficecreamsoldopenlyby retailoutletsinTurkey.Revue.Med. Vet., 157: 457 462.