Department of Microbiology, AIMS, B.G.Nagara,
Bowring & L.C.Hospital Bangalore, India.
Dr. D.Vijaya 130, IInd main road Emailfirstname.lastname@example.org Sheshadripurum, Bangalore 560020, Karnataka. Phone: 080-23446667; 9482009120
Introduction: CHROM agar for speciation of Candida is a differential culture medium which facilitates the isolation and identification of some clinically important species.
Materials and Methods: A total of 102 Candida species were isolated from various clinical specimens (100) including stool, sputum, nasal, oral, ear swabs, diabetic foot, skin and nail scrapings. Speciation of Candida was done using CHROM agar and conventional method simultaneously.
Observations: Non albicans candida (NAC) predominated (54.1%) over Candida albicans (45.9%). Non albicans candida spp isolated were C.tropicalis (35.29%), C.krusei (10.78%), C.parapsilosis (7.84%), and C. dubliniensis (0.9%). Antifungal susceptibility testing was done using antimycotic sensitivity testing by disc diffusion method. Isolates were 100% sensitive toketoconazole, clotrimazole, nystatin and amphotericin B. 87.5% of C.krusei, 36% C.tropicalis, 6.25% ,C. albicans were resistant to itraconazole. 25% C.krusei and 8% C.tropicalis were resistant of fluconazole. C.dubliniensis was resistant to itraconozole only.
Conclusion:The advantage of using CHROM agar is that it facilitates the isolation and identification of Candida to species level. The performance of CHROM agar exactly paralleled that of conventional methods. Use of this medium is rapid, technically simple and cost effective compared to time consuming technically demanding expensive conventional method.
CHRO Magar serves as a primary isolation and differentiation medium for clinical specimens that could allow mycology laboratories to rapidly identify Candida spp, enabling clinicians to choose appropriate antifungal agents,thus decreasing patients morbidity and mortality.
cite this article :
Vijaya D., Harsha T.R., Nagaratnamma T.. CANDIDA SPECIATION USING CHROM AGAR. Journal of Clinical and Diagnostic Research [serial online] 2011 August [cited: 2014 Mar 9 ]; 5:755-757. Available from http://www.jcdr.net/back_issues.asp?issn=0973-709x&year=2011&month=August&volume=5&issue=4&page=755-757&id=1482
Candida albicans historically has been predominant cause of Candidiasis.In 1980s C.albicans accounted for greater than 80% of all candidial isolates recovered from nosocomial yeast infections (1). More recently non albicans candida (NAC)species has been recovered with increasing frequency.
Several brands of chromogenic media have been developed to produce rapid yeast identification.These media contain chromogenic substrates that react with enzymes secreted by microorganisms producing colonies with various pigmentation. These enzymes are species specific, allowing organisms to be identified to the species level by their colour and colony characteristics (1). It is necessary to identify Candida to species level as many NAC have decreased susceptibility to antifungal agents. The present study was undertaken to evaluate the advantages of CHROM agar over conventional method for speciation of Candida isolates and their susceptibility to antifungal agents by disc diffusion method.
Material and Methods
A total of 102 Candida spp isolated from randomly selected 100 clinical specimens which included stool, sputum samples, nasal, oral and ear swabs, diabetic foot, skin and nail scrapings formed the study group. The study was conducted from Jan 2008 to Dec 2008,in the department of Microbiology, Bowring and L.C. Hospital, Bangalore.
CHROM agar was prepared as per the instruction manual (HI media India). Candida spp isolated were inoculated simultaneously to CHROM agar plates and SDA tubes. These were incubatedat 37ºC for 48 hours. Species were identified on CHROM agar by morphology and colour of the colony. Growth on SDA were speciated by standard methods using germ tube, corn meal agar, slide culture, sugar fermentation and assimilation test. Ten faecal samples and two oral swabs from AIDS patients were directly inoculated onto SDA and CHROMagar.
A variety of species specific colony were seen.
Appearances of Candida spp on CHROM agar were as follows: • C.albicans – blue green • C.tropicalis – dark blue gray centre with pink halo • C. krusei – pink large rough spreading colonies with pale edge. • C.parapsilisis – pale cream coloured colonies. • C.dubliniensis – dark green colonies.
Antifungal susceptibility was performed by disc diffusion method using antimycotic sensitiyity test agar. Discs used were amphotericin B (100 units), fluconazole (10mcg), clotrimazole (10mcg), nystatin (100mcg), itraconazole (10mcg), ketaconanole (10 mcg) was measured as for the instruction manual (HiMedia). ATCC strain of C.albicans was used as control.
(Table/Fig 1) shows Candida spp isolated in various clinical samples. All isolates of candida grew on CHROMagar after 48 hr of incubation at 370c.
C.dubliniensis isolated from faecal sample of AIDS patient grew only on CHROMagar.C.dubliniensis was further identified by germ tube, chlamydospore formation and no growth at 450C.
A mixed growth of C.tropicalis +C.parapsilosis, C.albicans + C.tropicalis were isolated in two sputum samples.
Isolates were 100% sensitive to amphotericin B, clotrimazole, nystatin and ketaconozole 87.5% of C.krusei, 36% C.tropicalis, 6.5% C.albicans were resistant to itraconozole.25% of C.krusei, 28% C.tropicalis showed resistant to fluconozole. C.dubliniensis was resistant to itraconozole only.
In the present study NAC (55.8%) was isolated at a higher rate than C.albicans as reported by other workers. Candida spp isolated by various workers is shown in (Table/Fig 2)(2)(3)(4)(5)(6)(7)(8)(9). In our 1997 study C.albicans predominated [10 ]. Among the stool samples received 90% were from antibiotic associated diarrhoea. C.dubliniensis isolated on CHROM agar did not grow on SDA. Isolation of this species has been reported by Raut (11). Using CHROMagar species like C.dubiliniensis can be isolated as in the present study. Mixed growth of Candida spp found in faecal specimens also has been reported by Frank (6).
(Table/Fig 3) shows the antifungal susceptibility as reported by various workers (12)(13)(14). Variability found in antifungal susceptibility testing using disc diffusion methods as shown in [Table/Fig-3], as it is dependent on the disc potency, inoculums size, temperature and duration of incubation.
The advantages of CHROMagar are easy to prepare ie boiling and dispencing in Petri plates, facilitates the rapid isolation and identification of yeast species. CHROM agar facilitates identification between yeast spp from specimens containing mixture of yeast spp as in the present study and do not affect the viability on subsequent subcultures (6). CHROM agar has the advantage of rapid identification of Candida species, technically simple, rapid and cost effective compared to technically demanding time consuming andThough the results on CHROM agar exactly paralled that of conventional method, it is superior to SDA in terms of suppressing the bacterial growth. Use of CHROM agar medium would allow mycology laboratories to identify rapidly, clinically important Candida Spp while potentially decreasing laboratory cost. Moreimportantly this capability will also enable clinicians to choose appropriate anti fungal agents, thus decreasing patient’s morbidity and mortality (Table/Fig 4), (Table/Fig 5),(Table/Fig 6). expensive conventional method.
This study was supported by Medical Education and Research Trust, Bangalore. Authors are grateful to the trustee of MERT Bangalore.
Lymn L. H., Duane R. H., Eliriton K. M. Dooley D. Direct isolation of candida spp from blood cultures on the chromogenic medium CHROM agar candida. J Clin Microbiol. 2003;41:6:2629-26322] Shivaprakasha S,Radhakrishan K,Karim PMS.Candida spp other Candida albicans: A major cause of fungemia in a terciary care centre. Indian J Med Microbiol 2007;25:4:405-407.
Capoor MR, Rawat D,Nair D,Deb M, Aggarwal P.Evaluation of glucosemethlene blue Muellar Hinton agar for E test minimum inhibitory concentration determination in candida spp. Indian J Med Microbiol 2007;25:4:432-433
Pfeller MA,Messer SA,,Hollis RJ et al. Trends in species distribution and susceptibility of fluconozole among blood stream isolates of Candida species in the United States. Diag Microbiol Infect Dis 1999;33:4:311-7