Rachel Chandy

Tobacco Agar, a New Medium for Differentiating Candida dubliniensis from Candida albicans

ABSTRACT Isolates of Candida dubliniensis may be misidentified as Candida albicans in microbiological laboratories if only the germ tube and/or the chlamydospore test is used for identification to the species level. In this study, we have evaluated the efficacy of tobacco agar for the differentiation of C. dubliniensis from C. albicans. On this medium at 28°C, all 30 C. dubliniensis isolates produced yellowish-brown colonies with hyphal fringes and abundant chlamydospores, whereas 54 C. albicans isolates formed smooth, white-to-cream-colored colonies with no chlamydospore production. This medium provides a simple tool for presumptive differentiation of C. dubliniensis from C. albicans.

Outbreak of Fungemia among Neonates Caused by Candida haemulonii Resistant to Amphotericin B, Itraconazole, and Fluconazole

ABSTRACT The first outbreak of Candida haemulonii fungemia is described. The seven isolates from the blood of four neonates were identified by DNA sequencing of the ribosomal DNA. They were all resistant to amphotericin B, fluconazole, and itraconazole. This report highlights the emergence of C. haemulonii as an opportunistic pathogen in immunocompromised patients.

Prevalence of Candida dubliniensis among germ tube-positive Candida isolates in a maternity hospital in Kuwait

In this study, 1644 germ tube‐positive Candida isolates from a maternity hospital was prospectively examined for the prevalence of Candida dubliniensis. Candida species were isolated from different clinical specimens, but majority (>90%) of them came from high vaginal swabs and urine specimens. The phenotypic and molecular identification methods for C. dubliniensis included production of rough colonies and chlamydospores on simplified sunflower seed agar, determination of assimilation profile by Vitek 2 yeast identification system, specific amplification of rDNA of internally transcribed spacer (ITS)‐2 region by semi‐nested PCR and direct DNA sequencing of the ITS‐1 and ITS‐2 regions. Three germ tube‐positive Candida isolates were identified as C. dubliniensis with an overall prevalence of 0.2%. Of these, two came from urine specimens and one from a vaginal swab. None of the C. dubliniensis isolates showed resistance against fluconazole, voriconazole and amphotericin B. The study reinforces the usefulness of sunflower seed agar in presumptive identification of C. dubliniensis and confirms the prevailing view that this species forms only a minor constituent of Candida species occurring in vagina or other anatomic sites of non‐HIV/AIDS‐infected individuals.

Candida albicans strain carriage in patients and nursing staff of an intensive care unit: a study of morphotypes and resistotypes

Candida albicans carriage of patients and nursing staff of an intensive care unit (ICU) was studied over an 8‐month period. Swabs were taken at weekly intervals from multiple sites from patients. None of the patients had clinical Candida infection at the time of the first sampling. The hands and mouth of the nursing staff were sampled at fortnightly intervals. Of the 68 patients investigated for varying periods, 37 (54%) yielded C. albicans from one or more body sites, resulting in the isolation of 269 strains. Nosocomial acquisition of C. albicans was recorded in seven (19%) patients. The frequency of C. albicans isolation increased with extended stay in ICU. Sixteen of the 180 samples taken from hands and mouth of nursing staff, yielded C. albicans, 12 of which came from the mouth. Morphotyping of 88 randomly selected strains of C. albicans originating from 31 patients yielded 34 morphotypes. There appeared to be no preference for any morphotype to colonize a particular anatomic site. Based on the susceptibility results, nine resistotypes were recognized. No correlation was apparent between any specific morphotype and resistotype patterns. The differences in morphotype and resistogram patterns of C. albicans isolates originating from same patients over a period of time suggest that some of the patients were colonized with more than one strain. Similarities in the morphotype and resistotype patterns of C. albicans strains isolated from patients and nursing staff tend to suggest possibility of exogenous acquisition.

Cryptococcus randhawai sp. nov., a novel anamorphic basidiomycetous yeast isolated from tree trunk hollow of Ficus religiosa (peepal tree) from New Delhi, India.

A novel anamorphic Cryptococcus species is described, which was isolated in New Delhi (India) from decaying wood of a tree trunk hollow of Ficus religiosa. On the basis of sequence analysis of the D1/D2 domains of the 26S rRNA gene and the internally transcribed spacer (ITS)-1 and ITS-2 region sequences, the isolate belonged to the Cryptococcus albidus cluster (Filobasidiales, Tremellomycetes) and was closely related to Cryptococcus saitoi, Cryptococcus cerealis and Cryptococcus friedmannii with 98% sequence identity. Phenotypically, the species differed from C. saitoi with respect to growth temperature (up to 37oC), presence of a thin capsule, ability to grow in the absence of vitamins, and inability to assimilate citrate and ethylamine. With respect to C. friedmannii, it differed in growth temperature, ability to assimilate lactose, raffinose, l-rhamnose, myo-inositol, and inability to utilize citrate. Furthermore, our isolate also differed from C. cerealis in growth temperature, presence of capsule and inability to assimilate l-sorbose. In view of the above phenotypic differences and unique rDNA sequences, we consider that our isolate represents a new species of Cryptococcus, and therefore, a new species, Cryptococcus randhawai is proposed for this taxon. The type strain J11/2002 has been deposited in the culture collection of the Centraalbureau voor Schimmelcultures (CBS10160) and CABI Biosciences (IMI 393306).

Candida dubliniensis: an appraisal of its clinical significance as a bloodstream pathogen.

A nine-year prospective study (2002–2010) on the prevalence of Candida dubliniensis among Candida bloodstream isolates is presented. The germ tube positive isolates were provisionally identified as C. dubliniensis by presence of fringed and rough colonies on sunflower seed agar. Subsequently, their identity was confirmed by Vitek2 Yeast identification system and/or by amplification and sequencing of the ITS region of rDNA. In all, 368 isolates were identified as C. dubliniensis; 67.1% came from respiratory specimens, 11.7% from oral swabs, 9.2% from urine, 3.8% from blood, 2.7% from vaginal swabs and 5.4% from other sources. All C. dubliniensis isolates tested by Etest were susceptible to voriconazole and amphotericin B. Resistance to fluconazole (≥8 µg/ml) was observed in 2.5% of C. dubliniensis isolates, 7 of which occurred between 2008–2010. Of note was the diagnosis of C. dubliniensis candidemia in 14 patients, 11 of them occurring between 2008–2010. None of the bloodstream isolate was resistant to fluconazole, while a solitary isolate showed increased MIC to 5-flucytosine (>32 µg/ml) and belonged to genotype 4. A review of literature since 1999 revealed 28 additional cases of C. dubliniensis candidemia, and 167 isolates identified from blood cultures since 1982. In conclusion, this study highlights a greater role of C. dubliniensis in bloodstream infections than hitherto recognized.

Mucor circinelloides as a Cause of Invasive Maxillofacial Zygomycosis: an Emerging Dimorphic Pathogen with Reduced Susceptibility to Posaconazole

ABSTRACT A case of maxillofacial zygomycosis caused by Mucor circinelloides, identified by phenotypic and molecular methods and treated successfully with liposomal amphotericin B (AmBisome) and surgical debridement, is described. The isolate was resistant to posaconazole. This report underscores the importance of prior susceptibility testing of zygomycetes to guide therapy with the most effective antifungal agent for an improved prognosis.

Aspergillus and other moulds in the air of Kuwait

A one-year survey was carried out to study the aerial prevalence of Aspergillus species and other moulds in the outdoor and indoor environments of Kuwait. Petri plates containing rose-Bengal medium were exposed for 20 minutes twice a month using a six-stage Andersen air sampler at the pre-determined sites. The exposed plates were incubated at 28 °C ± 1 °C up to 5 days and colonies were enumerated and identified by colonial and microscopic morphology. The data revealed that Aspergillus species were the predominant component (27.7%) of the outdoor aerospora of Kuwait and A. fumigatus alone accounted for 21.3% of the total aspergilli. In contrast, Cladosporium species formed the major component of the indoor aerospora (22.8%), followed by Aspergillus species (20.9%), Penicillium species (14.6%), and Bipolaris species (10.6%). A comparison of the fungi recorded in the outdoor and in the indoor air revealed that Aspergillus, Alternaria and Fusarium were significantly higher in the outdoor environment, whereas Cladosporium, Penicillium, and Bipolaris were significantly higher in the indoor environment. The relative prevalence of Aspergillus species and other moulds in the outdoor and indoor air of Kuwait was as follows: A. fumigatus 5.9 and 9.8%, A. flavus 4.9 and 3.9%, other aspergilli 16.8 and 7.0%, Alternaria species 19.8 and 7.9%, Cladosporium species 13.7 and 22.8%, Penicillium species 7.6 and 14.6%, and other moulds 31.2 and 34.1%, respectively. During the study, 25 different genera were identified, indicating a wide diversity in the spectrum of local fungal aerospora. The study provides useful information on the prevalence of allergenic fungi in the outdoor and indoor environments of Kuwait.

Nocardia asteroides in the soil of Kuwait.

A pilot study was undertaken to determine the occurrence and distribution of pathogenic nocardiae in Kuwaiti soil. A total of 102 soil samples collected from two localities were investigated by the paraffin bait technique. Nocardia asteroides was the only species isolated from 42 (41%) soil samples. None of the isolates fulfilled the criteria required for identification of N. farcinica or N. nova. Thirty one (73.8%) isolates showed equivalent growth at 45 °C and 35 °C, 17 (40.4%) isolates utilized acetamide for carbon and nitrogen requirements and 3 (7.1%) isolates showed delayed arylsulphatase activity. Only a solitary isolate was resistant to cefamandole. Soil samples originating from the Kuwait University Campus Shuwaikh, which were rich in humus/organic matter, were more productive for N. asteroides (67%) than the samples which were devoid of it but were mixed with crude oil (39%). Sand samples that lacked organic matter and crude oil samples were least productive of N. asteroides. These preliminary findings do not suggest that massive oil contamination of soil in the Ahmadi oil field area during the Gulf war promoted the natural occurrence of N. asteroides. However, isolation of N. asteroides in as many as 41% of the soil sample is a significant observation warranting further epidemiologic studies including its possible role in the operation desert storm sickness syndrome. This is the first report on the natural occurrence of N. asteroides in Kuwait.

Performance comparison of phenotypic and molecular methods for detection and differentiation of Candida albicans and Candida dubliniensis

BackgroundCandida albicans is the most pathogenic Candida species but shares many phenotypic features with Candida dubliniensis and may, therefore, be misidentified in clinical microbiology laboratories. Candidemia cases due to C. dubliniensis are increasingly being reported in recent years. Accurate identification is warranted since mortality rates are highest for C. albicans infections, however, C. dubliniensis has the propensity to develop resistance against azoles more easily. We developed a duplex PCR assay for rapid detection and differentiation of C. albicans from C. dubliniensis for resource-poor settings equipped with basic PCR technology and compared its performance with three phenotypic methods.MethodsDuplex PCR was performed on 122 germ tube positive and 12 germ tube negative isolates of Candida species previously identified by assimilation profiles on Vitek 2 ID-YST system. Typical morphologic characteristics on simplified sunflower seed agar (SSA), and reaction with a commercial (Bichro-Dubli) latex agglutination test were also performed. The assay was further applied on 239 clinical yeast and yeast-like fungi and results were confirmed by DNA sequencing of internal transcribed spacer (ITS) region of rDNA.ResultsThe results of duplex PCR assay for 122 germ tube positive and 12 germ tube negative isolates of Candida species were comparable to their identification by Vitek 2 ID-YST system, colony characteristics on SSA and latex agglutination test. Application of duplex PCR also correctly identified all 148 C. albicans and 50 C. dubliniensis strains among 239 yeast-like fungi.ConclusionsThe data show that both, duplex PCR and Bichro-Dubli are reliable tests for rapid (within few hours) identification of clinical yeast isolates as C. dubliniensis or C. albicans. However, duplex PCR may be applied directly on clinical yeast isolates for their identification as C. dubliniensis or C. albicans as it does not require prior testing for germ tube formation or latex Candida agglutination.