Parakriti Gupta’s research while affiliated with Government Medical College & Hospital and other places

Acrophialophora is implicated in superficial and invasive infections, especially in immunosuppressed individuals. The present study was undertaken to provide clinical, microbiological, phylogenetic, and antifungal susceptibility testing (AFST) profile of Acrophialophora isolated from India. All the isolates identified as Acrophialophora species at National Culture Collection for Pathogenic Fungi, Chandigarh, India were revived. Phenotypic and molecular characterization was performed, followed by temperature studies, scanning electron microscopy (SEM) and AFST. We also performed systematic review of all the cases of Acrophialophora species reported till date. A total of nine isolates identified as Acrophialophora species were identified by molecular method as A. fusispora (n = 8) and A. levis (n = 1), from brain abscess (n = 4), respiratory tract (n = 3) and corneal scraping (n = 2). All patients but two had predisposing factors/co-morbidities. Acrophialophora was identified as mere colonizer in one. Temperature studies and SEM divulged variation between both species. Sequencing of the ITS ribosomal DNA and beta-tubulin loci could distinguish species, while the LSU ribosomal DNA locus could not. AFST showed lowest MICs for triazoles and highest for echinocandins. Systematic literature review revealed 16 cases (11 studies), with ocular infections, pulmonary and central nervous system infections, and A. fusispora was common species. All the patients except three responded well. High MICs were noted for fluconazole, micafungin and caspofungin. This is the first study delineating clinical, phenotypic, and genotypic characteristics of Acrophialophora species from India. The study highlights microscopic differences between both species and emphasizes the role of molecular methods in precise identification. Triazoles appear to be the most effective antifungals for managing patients.

Background Cladosporium halotolerans is a saprobic fungus, rarely implicated in human infections. The identification is challenging due to non-specific phenotypic features. Objective To decipher clinical spectrum, microbiological and susceptibility profile of clinical and environmental isolates of Cladosporium halotolerans. Method All the isolates identified as Cladosporium halotolerans deposited in National Culture Collection for Pathogenic Fungi (NCCPF), Postgraduate Institute of Medical Education and Research, Chandigarh, India were revived. Phenotypic and molecular characterization targeting internal transcribed spacer (ITS) region of ribosomal DNA, large subunit of ribosomal DNA (LSU; NL1 and NL4), actin (ACT) and beta-tubulin (TUB) was done. Scanning electron microscopy (SEM) was performed to determine any phenotypic variations. Antifungal susceptibility testing (AFST) was carried out for eight antifungal agents as per CLSI M38 Ed3 guidelines. We also performed systematic literature review of all the cases of Cladosporium halotolerans reported till date. Results A total of four isolates (clinical, n = 3; soil, n = 1) identified as Cladosporium halotolerans were included in the study. The clinical sites were skin, maxillary tissue and nail. All patients were apparently immunocompetent, and history of trauma was recorded in one patient. All patients improved on antifungal therapy. The cultures revealed growth of black mycelial fungus and microscopic examination demonstrated dematiaceous septate hyphae with erect conidiophores and conidia in branched acropetal chains. Based on molecular methods, all the four isolates were identified as C. halotolerans. SEM revealed no variation in length and width of the conidia, conidiophores, ramoconidium and hyphae among the isolates. All molecular targets, such as ITS region, LSU (partially sequenced), ACT and TUB were able to differentiate the isolates. Minimum inhibitory concentrations for antifungals were: triazoles (0.12–2 μg/ml), amphotericin B (4 μg/ml) and echinocandins (2–8 μg/ml). Conclusion We report role of the rarely isolated dematiaceous fungus, C. halotolerans, in causing human infections. The study emphasizes the role of molecular methods in precisely identifying these species. Triazoles are more active against these black fungi compared to polyenes or echinocandins.

The present study describes clinicoepidemiological characteristics of cryptoccoccosis cases from North India. Middle aged HIV positive males were noted to be at highest risk. We report a considerable number of cases in apparently immmunocompetent patients. C. neoformans is the most common species causing the infection in North India. The higher MIC against AMB (drug of choice) needs to be investigated. The administration of 5-FC in the regimen shows better survival and its inclusion in the management protocol needs emphasis particularly in developing countries with high burden like India.

The world is heading towards an era of intractable and impending untreatable N. gonorrhoeae, thereby underlining the significance of rapid and accurate prediction of drug resistance as an indispensable need of the hour. In the present study, we optimized and evaluated a stable isotope labeling-based approach using the MALDI-TOF MS (Matrix-Assisted Laser Desorption/Ionization-Time of Flight Mass Spectrometry) for rapid and reliable detection of ciprofloxacin and azithromycin resistance in N. gonorrhoeae. All the isolates were cultured under three varied condition setups viz. medium supplemented with normal lysine, heavy lysine (isotope), and heavy lysine along with the antibiotics (ciprofloxacin/azithromycin), respectively. After incubation, spectra were acquired using the MALDI-TOF MS which were further screened for unique patterns (media-specific spectra) to differentiate drug-susceptible and resistant isolates. The results of the stable isotope labeling assay were comparable to the results of phenotypic methods used for susceptibility testing.

We describe a 36-year-old post-renal transplant patient diagnosed with a rare dual infection by Cryptococcus neoformans and Lichtheimia ramosa. The case highlights the significance of invasive samples and accurate interpretation of fungal biomarkers in the diagnosis of fungal infections in immunosuppressed patients. The mortality in such cases is high owing to delay in diagnosis, inability to perform surgical management or antifungal resistance.

Purpose Cryptococcosis is one of the most significant systemic fungal infections worldwide. Epidemiological data for pediatric cryptococcosis is very limited. Therefore, we planned this study to determine the burden of cryptococcosis in the pediatric population at our tertiary care center and performed review of literature. Material and methods In this retrospective study, all the patients less than 18 years of age were diagnosed with cryptococcosis, from January 2015–June 2021 were included. Demographic, clinical, and laboratory details of all the patients were noted. Furthermore, PubMed and MedLine databases were comprehensively searched for cases of pediatric cryptococcosis till June 2021. Results Of the total 5420 samples from suspected cryptococcosis cases processed at mycology laboratory, a total of 21 episodes of cryptococcosis (0.39%) were identified in 15 pediatric patients. The majority of the patients were apparently immunocompetent (10/15). Central nervous system (CNS) cryptococcosis was the most common presentation, followed by disseminated disease. All the isolates were identified as Cryptococcus neoformans (formerly referred to as C. neoformans var grubii), except one that was identified as Papiliotrema laurentii (formerly referred to as Cryptococcus laurentii). A standard treatment regimen inclusive of induction and maintenance therapy was provided in only five patients. The literature review revealed a total of 125 studies describing 1134 cases, of which 76.4% are reported from outside Asia, the majority (65.7%) restricted to CNS with C. neoformans as the commonest species. The management profile divulged a significantly higher use of standard drug regimen in Asia as compared to the rest of the world. Mortality of 13.32% was noted worldwide. Conclusion To the best of our knowledge, this is the first clinico-epidemiological study of pediatric cryptococcosis from India and the largest retrospective study worldwide. The rising incidence among immunocompetent individuals, especially in Asia, is a matter of concern. Clinical suspicion and early diagnosis are the cornerstones for the management of cases.

Poster session 2, September 22, 2022, 12:30 PM - 1:30 PM Background Lodderomyces elongisporus, earlier considered as a sexual state of Candida parapsilosis, was described as a distinct species based on ribosomal RNA gene sequencing. Few cases of human infections by this yeast have been described from Mexico, China, Malaysia, Kuwait, Australia, and the USA. We describe here eight cases of fungemia by L. elongisporus from a tertiary care hospital in North India. Methods Clinical characteristics and risk factors associated with L. elongisporus fungaemia were evaluated. Yeast isolated from blood culture (BD BACTEC™ 9240, New Jersey, USA) was identified by matrix-assisted laser desorption/ionization time-of-flight mass spectroscopy (MALDI-TOF MS, Bruker Daltonik GmbH, Bremen, Germany) and sequencing of D1/D2 region of a large subunit of ribosomal DNA. We performed antifungal susceptibility testing for amphotericin B, fluconazole, itraconazole, voriconazole, posaconazole, caspofungin, anidulafungin, and micafungin by the microbroth dilution method recommended by the Clinical and Laboratory Standards Institute (CLSI). Results We report eight cases of fungemia caused by L. elongisporus at our tertiary care center. Of these, three were infants (males) and five were adults (3 males and 2 females). The mean age of adults was 43.4 years. Among the pediatric cases, underlying diseases included congenital heart disease/atrophic kidney (neonate), tracheoesophageal fistula (4 months), and late-onset neonatal sepsis (LONS). Among the adults, underlying illnesses included acute kidney injury (n = 2), superior mesenteric artery thrombosis with bowel gangrene (n = 1), diabetes (n = 1), and central nervous system (CNS) lymphoma (n = 1). The iatrogenic factors included the history of surgery (n = 3), admission to ICU (n = 3), presence of urinary catheter (n = 4), and presence of central venous catheter (n = 1). Seven patients were on broad-spectrum antibiotics. The mean stay in the hospital was 20.38 days. Three of the patients were managed with fluconazole. Six patients improved while one left against medical advice (LAMA) and one expired. The range of minimum inhibitory concentration (MIC) of all the isolates against antifungals was as follows: amphotericin B (0.03-0.25 μg/ml), fluconazole (0.125 μg/ml), voriconazole (0.03 μg/ml), itraconazole (0.03-0.06 μg/ml), posaconazole (0.03-0.25 μg/ml), caspofungin (0.03-0.06 μg/ml), micafungin (0.03-0.06 μg/ml), and anidulafungin (0.03 μg/ml). Conclusion Lodderomyces elongisporus is an emerging pathogenic yeast causing fungemia in patients with comorbidities and undergoing surgery or invasive interventions. Though no antifungal breakpoints exist for this yeast, all the isolates exhibited low MICs to all the tested antifungals.

Poster session 3, September 23, 2022, 12:30 PM - 1:30 PM Objective: Rhytidhysteron spp. are emerging dematiaceous fungi implicated in subcutaneous infections in immunocompromised as well as immunocompetent individuals. Diagnosis of Rhytidhysteron spp is delayed owing to non-sporulation and misidentification. The present study was conducted to characterize Rhytidhysteron spp. isolated from India by molecular techniques. Method: We retrieved a total of eight isolates of Rhytidhysteron spp from our National Culture Collection for Pathogenic Fungi (NCCPF) that were received from various centers across India. Colony morphology on Sabouraud's dextrose agar (SDA) and potato dextrose agar (PDA) was recorded. Molecular identification was performed by sequencing internal transcribed spacer (ITS) and 28 s region of rDNA. Phylogenetic tree was constructed using the Neighbor-Joining method along with sequences of standard strains retrieved from the NCBI database. Results Rhytidhysteron spp was confirmed by sequencing from cutaneous specimens of eight cases. Six patients were immunocompetent while two were post-renal transplant recipients. Phenotypic features on SDA and PDA revealed growth of greyish-black mycelia on the obverse with black pigmentation on reverse after 7 days of incubation. On microscopic examination of lactophenol cotton blue mount, only dematiaceous septate hyphae without any spores were noted. On phylogenetic analysis of 28 s region, four of our isolates were closely clustered with Gloniopsis calami (Fig.1b), whereas two isolates with G. pneumonia and one each with G. percutanea, and Gloniopsis spp. (Fig.1b). On ITS phylogeny, the same four isolates closely clustered with G. calami (Fig.1a) as in 28 s region, one each isolate clustered with G. pneumonia, and G. percutanea while two isolates formed a separate clade with Rhytidhysteron rufulum (Fig.1a). Conclusion This fungus is often difficult to identify due to lack of sporulation making morphological identification challenging. Therefore, molecular sequencing is a must for its identification. However, identification using the sequence of ITS and 28 s rDNA does not clearly correlate. Hence to confirm the identity additional genes needs to be sequenced and analyzed.