A study on the multidisciplinary diagnostic approach of leprosy: Can we prevent the recrudescence in the post-elimination Indian scenario?

Introduction: Leprosy also widely known by the name Hansen's disease is a chronic disease caused by Mycobacterium leprae affecting mankind with various clinico-pathological forms. It remained a major public health issue due to associated case load, morbidity and stigma attached to it. India declared elimination of leprosy in the year 2005. However, it is surprising to see that in some parts of the country, the prevalence is still significant. The objective of the study is to describe the spectrum of histopathological profile of leprosy and compare its correlation with clinical diagnosis in this post elimination era. Methods: A 24-months prospective study was conducted with clinically diagnosed leprosy cases in a tertiary care hospital in eastern India. Lesions were graded and the histopathological slides along with its bacteriological index (BI) on slit skin smears where possible was reviewed and analyzed. Agreement of histopathological finding with clinical finding was established. Results: A total of 220 cases were included in the study. On histopathology, borderline category was the most frequently reported with borderline tuberculoid the most common subtype. Most common clinical feature was hypopigmented plaque, followed by erythematous skin lesions, nodules, macules etc. Bacteriological index was studied in 192 slit skin smears. Moderate agreement between clinical and histopathological diagnosis with kappa measure of inter-rater agreement as 0.457 was noted. Conclusion: Clinico-histopathological correlation is pivotal in the accurate diagnosis of leprosy to prevent, treat, and control the resurgence of the disease in the post-elimination era.

Intersectional insights into racism and health: not just a question of identity.

Intersectionality is a useful tool to address health inequalities, by helping us understand and respond to the individual and group effects of converging systems of power. Intersectionality rejects the notion of inequalities being the result of single, distinct factors, and instead focuses on the relationships between overlapping processes that create inequities. In this Series paper, we use an intersectional approach to highlight the intersections of racism, xenophobia, and discrimination with other systems of oppression, how this affects health, and what can be done about it. We present five case studies from different global locations that outline different dimensions of discrimination based on caste, ethnicity and migration status, Indigeneity, religion, and skin colour. Although experiences are diverse, the case studies show commonalities in how discrimination operates to affect health and wellbeing: how historical factors and coloniality shape contemporary experiences of race and racism; how racism leads to separation and hierarchies across shifting lines of identity and privilege; how racism and discrimination are institutionalised at a systems level and are embedded in laws, regulations, practices, and health systems; how discrimination, minoritisation, and exclusion are racialised processes, influenced by visible factors and tacit knowledge; and how racism is a form of structural violence. These insights allow us to begin to articulate starting points for justice-based action that addresses root causes, engages beyond the health sector, and encourages transnational solidarity.

Clinicopathological Spectrum of Syringoma: A Report of 50 Cases From a Tertiary Care Hospital in Eastern India.

Background Syringoma is a benign adnexal neoplasm and is considered safe with very low malignant potential. However, multiple tiny lesions typically affect the face and exposed area, which may cause a cosmetic concern for the patient. After a clinical diagnosis, there are two methods to diagnose syringoma: fine needle aspiration cytology (FNAC) and histopathology. FNAC is generally used for the initial evaluation of syringoma, while histopathology is used as a confirmatory test to diagnose syringoma. In developing and resource-limited settings, the combination of FNAC and histopathology would cause a financial and logistics burden. Objective This study aimed to observe the cytological and histopathological features of cases clinically diagnosed as syringomas in a tertiary care hospital to suggest the use of either FNAC or histopathology for diagnosing syringoma. Materials and Methods This cross-sectional observational study was conducted in the Department of Dermatology and Department of Pathology of a tertiary care hospital in eastern India from November 2021 to April 2022. Any clinically provisionally diagnosed case of syringoma was recruited for the study after obtaining informed consent for voluntary participation. With aseptic precautions, the tissue aspirates and punch biopsy were obtained in the Department of Dermatology and the samples were sent to the Department of Pathology. Cytological and histological examination was conducted by a single expert pathologist. Result A total of 50 cases (36 female, 14 male) with a median age of 23 years (range 10-40 years) were included in the study. A total of 43 cases were presented with papular lesions and seven with nodules. In the majority of the cases (40%), the lesion was in the eyelid followed by 26% in the arm. In FNAC, 22 cases were found to be benign adnexal lesions, 16 were suggestive of syringoma, eight were diagnosed as xanthoma, two were diagnosed as warts, and two cases were inadequate for opinion. Histologically, 42 cases were confirmed as syringoma, six were diagnosed as xanthoma, and two cases were diagnosed as warts. There was a significant difference between diagnosis by FNAC and histopathology (McNemar χ2 = 24.038, p-value = 0.0001). Conclusion We found that FNAC and histopathological diagnosis of syringoma may not be corroborative. Benign adnexal lesions are difficult to categorize by FNAC. Histopathological examination of clinically diagnosed cases of syringoma is of help for definitive diagnosis. Hence, FNAC may be avoided for saving time and discomfort for the patients and clinically diagnosed cases may be diagnosed by histopathological examination.

Regulation of conidiation and antagonistic properties of the soil-borne plant beneficial fungus Trichoderma virens by a novel proline-, glycine-, tyrosine-rich protein and a GPI-anchored cell wall protein.

Trichoderma spp. are widely used as commercial biofungicides, and most commercial formulations are conidia based. Identification of genes that regulate conidiation would thus be of help in genetic reprogramming of these species to optimize sporulation. In this study, we constructed an SSH (suppression subtractive hybridization) library from RNA samples of the wild type strain and a non-conidiating mutant, M7, grown under constant illumination for 2 days. We identified several genes that are underexpressed in the mutant. Some of these genes are related to secondary metabolism, and a few could be associated with conidiation. Genes coding for the following proteins, among others, were identified: O-methyl transferase, ATPase, alpha/beta-hydrolase, WD repeat containing protein, dehydrogenase, thioesterase, translationally controlled tumour protein, and a proline-glycine-tyrosine-rich protein (PGYRP) that has been annotated in T.reesei as a signalling protein. Two of these genes, encoding Pgy1, a novel PGYRP, and Ecm33, a GPI-anchored cell wall protein, were further studied in detail by generation of deletion mutants. We demonstrate here that both these genes not only regulate radial growth and conidiation in Trichoderma virens, but are also involved in antagonism against soil-borne wide host range plant pathogens. Furthermore, deletion of ecm33 affected hydrophobicity and cell wall integrity.

Leukocytoclastic vasculitis secondary to clozapine.

Leukocytoclastic vasculitis (LCV) may be secondary to drugs, underlying infection, collagen vascular disorders, or malignancy. Drug-induced vasculitis contributes to 10% of vasculitic skin lesions cases usually developing within 7-21 days of treatment initiation. The present case highlights a report of LCV in a 59-year-old male with a history of paranoid schizophrenia on clozapine therapy. The report upsurges the need to promote awareness and expedite diagnosis and treatment of drug-induced LCVs.

Etoricoxib-induced toxic epidermal necrolysis: A fatal case report.

Cyclooxygenase inhibitors were developed in the quest of enhanced analgesic efficacy devoid of gastric side effects. High usage of etoricoxib by prescription as well as self-administered routes has led to increasing reports of side effects and adverse reactions including dermatologic reactions in 0.1%-0.3% of cases. The present report enumerates a case of toxic epidermal necrolysis induced by etoricoxib.

Fsr1, a striatin homologue, forms an endomembrane-associated complex that regulates virulence in the maize pathogen Fusarium verticillioides.

Fsr1, a homologue of mammalian striatin, containing multiple protein-binding domains and a coiled-coil (CC) domain, is critical for Fusarium verticillioides virulence. In mammals, striatin interacts with multiple proteins to form a STRIPAK (striatin-interacting phosphatase and kinase) complex that regulates a variety of developmental processes and cellular mechanisms. In this study, we identified the homologue of a key mammalian STRIPAK component STRIP1/2 (striatin-interacting proteins 1 and 2) in F. verticillioides, FvStp1, which interacts with Fsr1 in vivo. Gene deletion analysis indicates that FvStp1 is critical for F. verticillioides stalk rot virulence. In addition, we identified three proteins, designated FvCyp1, FvScp1 and FvSel1, which interact with the Fsr1 CC domain via a yeast two-hybrid screen. Importantly, FvCyp1, FvScp1 and FvSel1 co-localize to endomembrane structures, each having a preferred localization in the cell, and they are all required for F. verticillioides stalk rot virulence. Moreover, these proteins are necessary for the correct localization of Fsr1 to the endoplasmic reticulum (ER) and nuclear envelope. Thus, we identified several novel components in the STRIPAK complex that regulates F. verticillioides virulence, and propose that the correct organization and localization of Fsr1 are critical for STRIPAK complex function.

Simultaneous knockdown of uPA and MMP9 can reduce breast cancer progression by increasing cell-cell adhesion and modulating EMT genes.

In cancer progression, proteolytic enzymes like serine proteases and metalloproteinases degrade the basement membrane enabling the tumor cells to invade the adjacent tissues. Thus, invasion and metastasis are augmented by these enzymes. Simultaneous silencing of uPA and MMP9 in breast cancer cells decreased the wound healing, migratory, invasive and adhesive capacity of the cells. After simultaneous down regulation, cells were seen to be arrested in the cell cycle. There was a remarkable increase in the expression of cell to cell adhesion molecule E-cadherin, and decrease in Vimentin and Snail expression. In addition, there was a significant decrease in the expression of the stem cell marker Oct-4. In the breast tumor samples it has been observed that, tumors, expressing higher level of uPA and MMP9, express less amount of E-cadherin. It has also been observed that few tumors also show, Vimentin positive in the ductal epithelial area. Thus, our model can help for checking the aggressive tumor invasion by blocking of uPA and MMP9. Our present observations also give the concept of the presence of aggressive epithelial cells with mesenchymal nature in the tumor micro-environment, altering the expression of EMT genes.

Trichoderma-plant-pathogen interactions: advances in genetics of biological control.

Trichoderma spp. are widely used in agriculture as biofungicides. Induction of plant defense and mycoparasitism (killing of one fungus by another) are considered to be the most important mechanisms of Trichoderma-mediated biological control. Understanding these mechanisms at the molecular level would help in developing strains with superior biocontrol properties. In this article, we review our current understanding of the genetics of interactions of Trichoderma with plants and plant pathogens.

Regulators of G-protein signalling in Fusarium verticillioides mediate differential host-pathogen responses on nonviable versus viable maize kernels.

GBB1, a heterotrimeric G-protein ß-subunit gene, was shown to be a key regulator of fumonisin B(1) (FB(1) ) biosynthesis in the maize pathogen Fusarium verticillioides. In this study, we performed functional analyses of genes that encode putative RGS (regulators of G-protein signalling) proteins and PhLPs (phosducin-like proteins) in F. verticillioides. These proteins are known to regulate heterotrimeric G-protein activity by altering the intrinsic guanosine triphosphatase (GTPase) activity, which, in turn, influences the signalling mechanisms that control fungal growth, virulence and secondary metabolism. Our aim was to isolate and characterize gene(s) that are under the transcriptional control of GBB1, and to test the hypothesis that these genes are directly associated with FB(1) regulation and fungal development in F. verticillioides on maize kernels. We first identified eight genes (two PhLPs and six RGSs) in the F. verticillioides genome, and a subsequent transcriptional expression study revealed that three RGS genes were up-regulated in the gbb1 deletion (Δgbb1) mutant and one RGS gene was up-regulated in the wild-type. To characterize their function, we generated knockout mutants using a homologous recombination strategy. When grown on autoclaved nonviable kernels, two mutants (ΔflbA2 and ΔrgsB) produced significantly higher levels of FB(1) compared with the wild-type progenitor, suggesting that the two mutated genes are negative regulators of FB(1) biosynthesis. ΔflbA2 also showed a severe curly conidia germination pattern, which was contradictory to that observed in the Δgbb1 strain. Strikingly, when these mutants were grown on live maize kernels, we observed contrasting FB(1) and conidiation phenotypes in fungal mutants, which strongly suggests that these G-protein regulators have an impact on how F. verticillioides responds to host/environmental factors. Our data also provide evidence that fungal G-protein signalling is important for modulating the ethylene biosynthetic pathway in maize kernels.

Comparative genome sequence analysis underscores mycoparasitism as the ancestral life style of Trichoderma.

BACKGROUND: Mycoparasitism, a lifestyle where one fungus is parasitic on another fungus, has special relevance when the prey is a plant pathogen, providing a strategy for biological control of pests for plant protection. Probably, the most studied biocontrol agents are species of the genus Hypocrea/Trichoderma. RESULTS: Here we report an analysis of the genome sequences of the two biocontrol species Trichoderma atroviride (teleomorph Hypocrea atroviridis) and Trichoderma virens (formerly Gliocladium virens, teleomorph Hypocrea virens), and a comparison with Trichoderma reesei (teleomorph Hypocrea jecorina). These three Trichoderma species display a remarkable conservation of gene order (78 to 96%), and a lack of active mobile elements probably due to repeat-induced point mutation. Several gene families are expanded in the two mycoparasitic species relative to T. reesei or other ascomycetes, and are overrepresented in non-syntenic genome regions. A phylogenetic analysis shows that T. reesei and T. virens are derived relative to T. atroviride. The mycoparasitism-specific genes thus arose in a common Trichoderma ancestor but were subsequently lost in T. reesei. CONCLUSIONS: The data offer a better understanding of mycoparasitism, and thus enforce the development of improved biocontrol strains for efficient and environmentally friendly protection of plants.

Overlapping and distinct functions of two Trichoderma virens MAP kinases in cell-wall integrity, antagonistic properties and repression of conidiation.

We have studied the functions of the Trichoderma virens TmkB, a homologue of the yeast cell-wall integrity MAP kinase Slt2, using gene knockout. The functions of TmkB were compared to those of the pathogenicity MAP kinase homologue (TmkA). Like the tmkA loss-of-function mutants, tmkB mutants exhibited reduced radial growth and constitutive conidiation in dark as well as in liquid shake cultures. The tmkB mutants, in contrast to tmkA mutants, had cell-wall integrity defects, as shown by autolysis of the mycelia and increased sensitivity to cell-wall degrading enzymes. Interestingly, the tmkB mutants were not autolytic on the synthetic Vogels minimal medium. The tmkB mutants had attenuated ability to overgrow the plant pathogen Sclerotium rolfsii, while retaining the ability to overgrow Rhizoctonia solani and Pythium spp., a phenotype also exhibited by the tmkA mutants. This first functional analysis of a cell-wall integrity MAPK in Trichoderma spp., a group of economically important fungi, shows the importance of this signaling pathway in biocontrol. Common phenotypes of the TmkA and TmkB pathways suggest that the two MAPKs may share some substrates, perhaps subunits of key transcription factors, thus dependent on two phosphorylation events for their activity.

Comparative genomics reveals mobile pathogenicity chromosomes in Fusarium.

Fusarium species are among the most important phytopathogenic and toxigenic fungi. To understand the molecular underpinnings of pathogenicity in the genus Fusarium, we compared the genomes of three phenotypically diverse species: Fusarium graminearum, Fusarium verticillioides and Fusarium oxysporum f. sp. lycopersici. Our analysis revealed lineage-specific (LS) genomic regions in F. oxysporum that include four entire chromosomes and account for more than one-quarter of the genome. LS regions are rich in transposons and genes with distinct evolutionary profiles but related to pathogenicity, indicative of horizontal acquisition. Experimentally, we demonstrate the transfer of two LS chromosomes between strains of F. oxysporum, converting a non-pathogenic strain into a pathogen. Transfer of LS chromosomes between otherwise genetically isolated strains explains the polyphyletic origin of host specificity and the emergence of new pathogenic lineages in F. oxysporum. These findings put the evolution of fungal pathogenicity into a new perspective.

cAMP signalling is involved in growth, germination, mycoparasitism and secondary metabolism in Trichoderma virens.

An adenylate-cyclase-encoding gene, tac1, of Trichoderma virens, a soil fungus used in the biocontrol of plant pathogens, has been cloned and sequenced. The tac1 ORF spanned 7032 bp, encoding a protein of 2153 aa, which shared an identity of 65 % with the adenylate cyclase of Colletotrichum lagenarium. Deletion of tac1, through double-crossover homologous recombination, lowered the intracellular cAMP levels to below the detection limit. The mutants showed only 5-6 % of the wild-type growth rate on agar, but grew normally in shake culture. The mutants did not sporulate in darkness, and the spores failed to germinate in water. In the confrontation assay, the mutants did not overgrow the test plant pathogens Sclerotium rolfsii, Rhizoctonia solani and Pythium sp. Against Pythium sp., the mutants produced a clear zone of inhibition in the confrontation assay. HPLC analysis and bioassay showed reduced secondary metabolite production in the mutants. Using suppression subtractive hybridization (SSH), the genes that were underexpressed in the mutants were identified. Based on an array of 53 SSH library clones, 11 clones were identified as strongly downregulated in the Deltatac1 mutants; of these 11 clones, nine sequences were homologous to secondary metabolism-related gene sequences. Therefore, cAMP signalling positively regulates secondary metabolism in T. virens. This is believed to be the first direct genetic study on the role of cAMP signalling in a Trichoderma sp. Tac1 is also believed to be the first regulatory protein to be identified in T. virens that is involved in growth, germination, mycoparasitism and secondary metabolism.

A secondary metabolite biosynthesis cluster in Trichoderma virens: evidence from analysis of genes underexpressed in a mutant defective in morphogenesis and antibiotic production.

A transcriptional comparison of wild type and a secondary metabolite deficient Trichoderma virens mutant resulted in the identification of six genes similar to those involved in secondary metabolism in other fungi, including four cytochrome P450 genes, one O-methyl transferase and one terpene cylase. Four of the genes (three cytochrome P450s and the cyclase) are located as a cluster. Transcript levels of three of the P450 genes, the O-methyl transferase and the terpene cyclase were measured. These genes are underexpressed in the mutant, which lacks the major secondary metabolites produced by this strain, viridin and viridiol. Expression levels of clones from the differential library with similarity to fungal trehalose synthase and a hydrophobin were also underexpressed in the mutant, while a heat shock protein hsp98 homolog was not. Based on the gene expression pattern and associated secondary metabolite profile, along with similarity to other secondary metabolism pathways in related fungi, we predict that the cluster is associated with the production of a terpene. The terpene could be viridin. This is the first report on cloning of secondary metabolism related genes from T. virens, and of their organization in a cluster, in this biocontrol fungus.