Fahmi Hasan

Biofilm formation in clinical Candida isolates and its association with virulence

Biofilm formation, an important virulence trait of Candida species was measured in 107 Candida isolates from 32 candidemic patients by XTT [2,3-bis (2-methoxy-4nitro-5-sulfo-phenyl)-2H-tetra-zolium-5-carboxanilide] activity and compared to biofilm formation of Candida isolates from oropharyngeal lesions of 19 AIDS patients. Biofilm formation by XTT varied among species and C. albicans; C. lusitaniae and C. krusei produced more biofilm than the other Candida species. C. tropicalis was the most dominant species isolated from blood followed by C. albicans, and other non-albicans species whereas only C. albicans was recovered from oral lesions. Importantly, though Biofilm formation was variable within a species it was stable in sequential isolates during chronic infection. Sequential isolates exhibited identical Karyotype pattern or RAPD patterns unless patients were co-infected with more than one strain. High biofilm formation was associated with slow growth rate but not with adherence. Murine infection studies demonstrated that, degree of in-vitro biofilm formation was associated with virulence in mice, as mice infected both with no and low biofilm formers survived longer than mice infected with high biofilm former C. albicans (p< or =0.001). We conclude that biofilm formation is a stable but strain specific characteristic that can greatly vary among C. albicans and non-albicans strains, and plays an important role in persistence of infection.

Epidemiology of candidemia in a tertiary care centre of north India: 5-year study.

Background:To determine the distribution of species of Candida and the risk factors associated with candidemia in Indian population for which we conducted a retrospective study for 5 years in a tertiary care centre of North India.Materials and Methods:Blood samples from 7,297 patients aged from 3 days to 85 years, suspected with candidemia, were collected and tested for Candida. The susceptibility patterns toward fluconazole for the year 2005 isolates were tested by micro-dilution assay as described in the CLSI (M27A-2 method).Results:Most of the episodes have been caused by species other than C. albicans. Non-albicans candidemia was 79%–80% in both female and male populations. The most frequent species isolated from 275 patients in 5 years (January 2001–December 2005) was C. tropicalis (35.3%), followed by C. albicans (21.5%), C. parapsilosis (20%), C. glabrata (17.5%), C. krusei (3.3%), C. haemulonii (1.5%), and C. guilliermondii (1%). C. parapsilosis was the predominant in the fifth year of the study (2004–2005). Dose-dependant susceptibility to fluconazole was observed in 5% (n = 3) of the strains. Antifungal resistance was found in 11.7% (n = 7), which includes only C. glabrata.Conclusion:These results were comparable to those derived from other regions of India. C. tropicalis has been reported as the predominant species involved in the cases of candidemia. But in 2005 it has moved toward C. parapsilosis. No true antifungal resistance is reported. Further epidemiological surveillance is needed.

Isolation and Characterization of Senescent Cryptococcus neoformans and Implications for Phenotypic Switching and Pathogenesis in Chronic Cryptococcosis

ABSTRACT Although several virulence factors and associated genes have been identified, the mechanisms that allow Cryptococcus neoformans to adapt during chronic infection and to persist in immunocompromised hosts remain poorly understood. Characterization of senescent cells of C. neoformans demonstrated that these cells exhibit a significantly enlarged cell body and capsule but still cross the blood-brain barrier. C. neoformans cells with advanced generational age are also more resistant to phagocytosis and killing by antifungals, which could promote their selection during chronic disease in humans. Senescent cells of RC-2, a C. neoformans strain that undergoes phenotypic switching, manifest switching rates up to 11-fold higher than those of younger cells. Infection experiments with labeled cells suggest that senescent yeast cells can potentially accumulate in vivo. Mathematical modeling incorporating different switching rates demonstrates how increased switching rates promote the emergence of hypervirulent mucoid variants during chronic infection. Our findings introduce the intriguing concept that senescence in eukaryotic pathogens could be a mechanism of microevolution that may promote pathoadaptation and facilitate evasion of an evolving immune response.

Candida rugosa: a possible emerging cause of candidaemia in trauma patients

IntroductionCandida rugosa appears to be emerging as a distinctive cause of candidaemia in recent years. Candidaemia due to this species is important to recognise because of its decreased susceptibility to azoles.Materials and methodsWe retrospectively evaluated a cluster of C. rugosa candidaemia occurring in critically ill trauma patients from a level I trauma centre of India. During the period from July 2008 to September 2009, a total of 28 blood samples from 19 patients were found to be positive for C. rugosa. Genetic relatedness among 17 C. rugosa isolates were characterised by the random amplified polymorphic DNA (RAPD) assay using M13 primers. These isolates were also characterised for their susceptibility to four antifungal agents, amphotericin B, fluconazole, flucytosine and voriconazole.ResultsIn our study, 21% of C. rugosa isolates were resistant to fluconazole, whereas 100% susceptibility to amphotericin B, flucytosine and voriconazole was noted. Thirteen out of the 19 patients (68.4%) with C. rugosa candidaemia died. Of these, six had received antifungal therapy after confirmation of fungaemia.DiscussionPrior to this cluster, C. rugosa had never been identified as a cause of infection at our centre. Due to the retrospective nature of the evaluation of these cases, the source of this possible outbreak could not be traced. Nevertheless, to the best of our knowledge, this is the largest cluster of cases of C. rugosa candidaemia reported from a single institution in the English literature.

Clinical presentation and survival of retinoblastoma in Indian children

Objective To study the clinical presentation and survival among Indian children with retinoblastoma (RB) and to determine factors predictive of poor outcome. Methods A retrospective review of children newly diagnosed with RB at a tertiary referral centre was undertaken. Demographic and clinical characteristics and treatment outcomes were studied. Results A total of 600 patients (unilateral 67.6%, bilateral 32.4%) was studied. 61% was boys. The median age at presentation was 29 months (18 months vs 36 months in bilateral and unilateral cases, respectively, p<0.001). leukocoria was most common (83%), followed by proptosis (17%). Tumours were intraocular in 72.3% and extraocular in 27.7% cases. In the intraocular group, 78% were advanced Group D or E disease. Metastasis to the central nervous system was noted in 15.7% of extraocular cases. A statistically significant difference was seen between intraocular and extraocular groups in the median age (24 months vs 37.5 months, p<0.001) and median lag period (2.5 months vs 7 months, p<0.001). The Kaplan-Meier survival probability was 83%, 73% and 65% at 1 year, 2 years and 5 years, respectively. On univariate analysis, age >2 years (p=0.002), lag period >6 months (p=0.004) and extraocular stage (p<0.001) were associated with poor outcome. On multivariate analysis, extraocular invasion was predictive of low survival (HR 5.04, p<0.001). Conclusions Delayed presentation is a matter of concern. Improving awareness about the early signs and creating facilities for diagnosing and treating RB at the primary and secondary levels of healthcare are required to reduce mortality and morbidity, and lead to improved outcomes that are comparable with the developed nations.

Five-year profile of candidaemia at an Indian trauma centre: High rates of Candida auris blood stream infections

Candidaemia is a potentially fatal infection with varied distribution of Candida species and their antifungal susceptibility profiles. The recent emergence of Candida auris in invasive candidiasis is a cause for concern. This study describes the profile of candidaemia at an Indian tertiary care hospital and reports the emergence of C. auris. All patients diagnosed with candidaemia between 2012 and 2017 were studied. The isolates were identified using conventional methods, VITEK 2 and MALDI‐TOF MS. The isolates not identified by MALDI‐TOF were sequenced. Antifungal susceptibility testing was done by the CLSI broth microdilution method and VITEK 2. A total of 114 isolates of Candida species were analysed. Candida tropicalis (39.4%) was the most common species, followed by C. auris (17.5%), C. albicans (14%) and C. parapsilosis (11.4%). Notably, Diutina mesorugosa isolates (n = 10) were not identified by MALDI‐TOF and were confirmed by sequencing. Furthermore, 45% (n = 9) C. auris strains exhibited low MICs of FLU (0.05‐4 μg/mL) and the remaining 55% (n = 11) isolates had high MICs ≥ 64 μg/mL. Also, D. mesorugosa exhibited high MICs of FLU (32 μg/mL) in 2 isolates. A high rate of errors in antifungal susceptibility was noted with the VITEK 2 as compared to the CLSI method. Candida auris was the second most prevalent species causing candidaemia warranting infection control practices to be strengthened to prevent its spread.

Magnetic resonance imaging for tumor restaging after chemotherapy in retinoblastoma with optic nerve invasion

ABSTRACT Purpose: Extraocular retinoblastoma with optic nerve invasion is treated by a multimodal protocol consisting of neoadjuvant chemotherapy, enucleation, and adjuvant therapy. This study was conducted to evaluate the performance of magnetic resonance imaging (MRI) used for tumor restaging in these children after systemic chemotherapy administration. Methods: Contrast-enhanced MRI scan of orbits and brain was performed at diagnosis and patients were treated with neoadjuvant chemotherapy. After chemotherapy, MRI scan was repeated for tumor restaging and residual post-laminar thickening and/or enhancement of the affected optic nerve, if any, was recorded. MRI findings were correlated with histopathology in enucleated specimens. The main outcome measures were specificity, sensitivity, and accuracy of MRI in predicting post-laminar invasion after neoadjuvant chemotherapy. Results: A total of 46 eyes (46 patients) were studied. Optic nerve thickening on MRI had a sensitivity, specificity, and accuracy of 100% (95% Confidence Interval (CI): 64.6–100%), 76.9% (95% CI: 61.7–87.4%), and 80.4% (95% CI: 66.8–89.4%), respectively. Optic nerve enhancement had a sensitivity, specificity, and accuracy of 85.7% (95% CI: 48.7–97.4%), 79.5 % (95% CI: 64.5–89.2%), and 80.4% (95% CI: 66.8–89.4%), respectively. Combined thickening and enhancement of the optic nerve had a sensitivity, specificity, and accuracy of 100% (95% CI: 60.9–100%), 82.4% (95% CI: 66.5–91.7%), and 85% (95% CI: 70.9–92.9%), respectively. Conclusion: MRI is a valuable tool for restaging of retinoblastoma and predicting residual optic nerve disease after neoadjuvant chemotherapy. Combined thickening and enhancement on MRI appeared to be a more reliable indicator of post-laminar invasion as compared to thickening or enhancement alone.