Ras signaling activates glycosylphosphatidylinositol (GPI) anchor biosynthesis via the GPI-N-acetylglucosaminyltransferase (GPI-GnT) in Candida albicans.

The ability of Candida albicans to switch between yeast to hyphal form is a property that is primarily associated with the invasion and virulence of this human pathogenic fungus. Several glycosylphosphatidylinositol (GPI)-anchored proteins are expressed only during hyphal morphogenesis. One of the major pathways that controls hyphal morphogenesis is the Ras-signaling pathway. We examine the cross-talk between GPI anchor biosynthesis and Ras signaling in C. albicans. We show that the first step of GPI biosynthesis is activated by Ras in C. albicans This is diametrically opposite to what is reported in Saccharomyces cerevisiae Of the two C. albicans Ras proteins, CaRas1 alone activates GPI-GnT activity; activity is further stimulated by constitutively activated CaRas1. CaRas1 localized to the cytoplasm or endoplasmic reticulum (ER) is sufficient for GPI-GnT activation. Of the six subunits of the GPI-N-acetylglucosaminyltransferase (GPI-GnT) that catalyze the first step of GPI biosynthesis, CaGpi2 is the key player involved in activating Ras signaling and hyphal morphogenesis. Activation of Ras signaling is independent of the catalytic competence of GPI-GnT. This too is unlike what is observed in S. cerevisiae where multiple subunits were identified as inhibiting Ras2. Fluorescence resonance energy transfer (FRET) studies indicate a specific physical interaction between CaRas1 and CaGpi2 in the ER, which would explain the ability of CaRas1 to activate GPI-GnT. CaGpi2, in turn, promotes activation of the Ras-signaling pathway and hyphal morphogenesis. The Cagpi2 mutant is also more susceptible to macrophage-mediated killing, and macrophage cells show better survival when co-cultured with Cagpi2.

An in vivo evaluation of different methods of working length determination.

OBJECTIVE: The purpose of this in vivo study was to compare the ability of digital tactile, digital radiographic and electronic methods to determine reliability in locating the apical constriction. MATERIALS AND METHODS: Informed consent was obtained from patients scheduled for orthodontic extraction. The teeth were anesthetized, isolated and accessed. The canals were negotiated, pulp chamber and canals were irrigated and pulp was extirpated. The working length was then evaluated for each canal by digital tactile sensation, an electronic apex locator (The Root ZX) and digital radiography. The readings were then compared with post-extraction working length measurements. RESULTS: The percentage accuracy indicated that EAL method (Root ZX) shows maximum accuracy, i.e. 99.85% and digital tactile and digital radiographic method (DDR) showed 98.20 and 97.90% accuracy respectively. CLINICAL SIGNIFICANCE: Hence, it can be concluded that the EAL method (Root ZX) produced most reliable results for determining the accurate working length.

SLD with and without ADHD: Comparison of cognitive profiles.

A quarter of children with Specific Learning Disorder (SLD) have a comorbid attention deficit hyperactivity disorder (ADHD), which impacts the testing of intelligence. ADHD is therefore treated before proceeding with the assessment of intelligence. It is expected that the treatment of ADHD will mitigate its effects on intelligence testing. Though this is frequently done in clinical practice, we could not find any study comparing IQ profiles of children having SLD with and without ADHD after treating ADHD to look for any changes between them. Therefore, we planned this study to compare the cognitive profiles of children having SLD with and without ADHD and describe any difference in their profiles. It was a retrospective cross-sectional study. We compared 695 children having SLD with treated ADHD to 721 children having SLD without ADHD on their WISC III profiles. We found that children having SLD with treated ADHD scored significantly lower on Digit Span and Coding. We suggest routine use of Cognitive Working Memory Training, peer tutoring, and task modifications in children having SLD with ADHD along with medicines and remedial education for better outcomes.

Candida auris on Apples: Diversity and Clinical Significance.

Candida auris is a multidrug-resistant nosocomial fungal pathogen. While the marine environment was recently identified as a natural niche for C. auris, the environment(s) that might have contributed to the development and spread of antifungal resistance in C. auris remains a mystery. Because stored fruits are often treated with fungicides to prevent postharvest spoilage, we hypothesized that stored fruits could serve as a possible selective force for and a transmission reservoir of antifungal-resistant isolates of pathogenic yeasts, including C. auris. To test this hypothesis, we screened fruits to study the diversity of pathogenic yeasts and their antifungal susceptibility profiles. Among the 62 screened apples, the surfaces of 8 were positive for C. auris, and all were stored apples. Whole-genome sequencing (WGS) showed that C. auris strains from apples were genetically diverse and exhibited broad phylogenetic distribution among the subclades within clade I. Interestingly, strains from apples had closely related strains from other sources in India, including from patients, hospitals, and marine environments, and from clinical strains from other parts of the world. A broad range of fungicides, including dimethyl inhibitors (DMIs), were detected in stored apples, and all C. auris isolates exhibited reduced sensitivity to DMIs. Interestingly, C. auris was not isolated from freshly picked apples. Together, the results suggest a potentially complex ecology for C. auris with agriculture fungicide application on stored fruits as a significant selective force for drug resistance in clinics. IMPORTANCE In 2019, the U.S. Centers for Disease Control and Prevention classified the multidrug-resistant Candida auris as one of five pathogens posing the most urgent threats to public health. At present, the environment(s) that might have contributed to the development and spread of antifungal resistance in C. auris is unknown. Here, we tested whether fruits could be a source of multidrug-resistant C. auris. We identified genetically diverse C. auris strains with reduced sensitivity to major triazole dimethyl inhibitors fungicides on the surfaces of stored apples. The successful isolation of C. auris from apples here calls for additional investigations into plants as a reservoir of C. auris. Our findings suggest that C. auris in the natural ecosystem may come in contact with agriculture fungicides and that stored fruits could be a significant niche for the selection of azole resistance in C. auris and other human fungal pathogens.

A High Frequency of Candida auris Blood Stream Infections in Coronavirus Disease 2019 Patients Admitted to Intensive Care Units, Northwestern India: A Case Control Study.

BACKGROUND: The ongoing pandemic of coronavirus disease 2019 (COVID-19) has overwhelmed healthcare facilities and raises an important novel concern of nosocomial transmission of Candida species in the intensive care units (ICUs). METHODS: We evaluated the incidence and risk factors for development of candidemia in 2384 COVID-19 patients admitted during August 2020-January 2021 in ICUs of 2 hospitals (Delhi and Jaipur) in India. A 1:2 case-control matching was used to identify COVID-19 patients who did not develop candidemia as controls. RESULTS: A total of 33 patients developed candidemia and accounted for an overall incidence of 1.4% over a median ICU stay of 24 days. A 2-fold increase in the incidence of candidemia in COVID-19 versus non-COVID-19 patients was observed with an incidence rate of 14 and 15/1000 admissions in 2 ICUs. Candida auris was the predominant species (42%) followed by Candida tropicalis. Multivariable regression analysis revealed the use of tocilizumab, duration of ICU stay (24 vs 14 days), and raised ferritin level as an independent predictor for the development of candidemia. Azole resistance was observed in C auris and C tropicalis harboring mutations in the azole target ERG11 gene. Multilocus sequence typing (MLST) identified identical genotypes of C tropicalis in COVID-19 patients, raising concern for nosocomial transmission of resistant strains. CONCLUSIONS: Secondary bacterial infections have been a concern with the use of tocilizumab. In this cohort of critically ill COVID-19 patients, tocilizumab was associated with the development of candidemia. Surveillance of antifungal resistance is warranted to prevent transmission of multidrug-resistant strains of nosocomial yeasts in COVID-19 hospitalized patients.

Environmental Isolation of Candida auris from the Coastal Wetlands of Andaman Islands, India.

Candida auris is a multidrug resistant pathogen that presents a serious global threat to human health. As C. auris is a newly emerged pathogen, several questions regarding its ecological niche remain unexplored. While species closely related to C. auris have been detected in different environmental habitats, little is known about the natural habitat(s) of C. auris Here, we explored the virgin habitats around the very isolated Andaman Islands in the Indian Ocean for evidence of C. auris We sampled coastal wetlands, including rocky shores, sandy beaches, tidal marshes, and mangrove swamps, around the Andaman group of the Andaman & Nicobar Islands, Union Territory, in India. Forty-eight samples of sediment soil and seawater were collected from eight sampling sites representing the heterogeneity of intertidal habitats across the east and west coast of South Andaman district. C. auris was isolated from two of the eight sampling sites, a salt marsh and a sandy beach. Interestingly, both multidrug-susceptible and multidrug-resistant C. auris isolates were found in the sample. Whole-genome sequencing analysis clustered the C. auris isolates into clade I, showing close similarity to other isolates from South Asia. Isolation of C. auris from the tropical coastal environment suggests its association with the marine ecosystem. The fact that viable C. auris was detected in the marine habitat confirms C. auris survival in harsh wetlands. However, the ecological significance of C. auris in salt marsh wetland and sandy beaches to human infections remains to be explored.IMPORTANCECandida auris is a recently emerged multidrug-resistant fungal pathogen capable of causing severe infections in hospitalized patients. Despite its recognition as a human pathogen a decade ago, so far the natural ecological niche(s) of C. auris remains enigmatic. A previous hypothesis suggested that C. auris might be native to wetlands, that its emergence as a human pathogen might have been linked to global warming effects on wetlands, and that its enrichment in that ecological niche was favored by the ability of C. auris for thermal tolerance and salinity tolerance. To understand the mystery of environmental niches of C. auris, we explored the coastal wetland habitat around the very isolated Andaman Islands in the Indian Ocean. C. auris was isolated from the virgin habitats of salt marsh area with no human activity and from a sandy beach. C. auris isolation from the marine wetlands suggests that prior to its recognition as a human pathogen, it existed as an environmental fungus.

Synthesis of amino acid appended indoles: appreciable anti-fungal activity and inhibition of ergosterol biosynthesis as their probable mode of action.

Rationally designed compounds consisting of mono- and di-peptide appendages on bis-indole template were synthesized in appreciable yield. Some of these compounds exhibited significant antifungal activities against Candida albicans with their MIC80 in µg/ml range. However, when used in combination with azoles, the antifungal activities of the azoles were considerably enhanced. The growth inhibition appeared to be specific to the fungal cells and mammalian cells were not affected by these compounds. It is shown that these compounds lower ergosterol levels in the fungal cells and probably act by targeting lanosterol 14α-demethylase, a key enzyme in the sterol biosynthetic pathway of C. albicans. The compounds do not appear to directly act on the fungal cell wall. Hence, the sensitivity of the fungal cells to these compounds cannot be attributed to cell wall damage and consequent accumulation of the compounds in the cell, though defects in cell wall due to defective sterol biosynthesis cannot be completely ruled out.