Hemolymph enables communication between organs in insects and ensures necessary coordination and homeostasis. Its composition can provide important information about the physiological state of an insect and can have diagnostic significance, which might be particularly important in the case of harmful insects subjected to biological control. Galleria mellonella Linnaeus 1758 (Lepidoptera: Pyralidae) is a global pest to honey bee colonies. The hemolymph of its larvae was examined after infection with the soil fungus Conidiobolus coronatus (Constantin) Batko 1964 (Entomophthorales). It was found that after one hour of contact with the fungus, the volume of the hemolymph increased while its total protein content decreased. In larvae with a high pathogen load, just before death, hemolymph volume decreased to nearly initial levels, while total protein content and synthesis (incorporation of 35S-labeled methionine) increased. The hemolymph polypeptide profile (SDS-PAGE followed by autoradiography) of infected insects was significantly different from that of healthy larvae. Hemocytes of infected larvae did not surround the fungal hyphae, although they encapsulated small foreign bodies (phase contrast microscopy). Infection had a negative effect on hemocytes, causing oenocyte and spherulocyte deformation, granulocyte degranulation, plasmatocyte vacuolization, and hemocyte disintegration. GC-MS analysis revealed the presence of 21 compounds in the hemolymph of control insects. C. coronatus infection caused the appearance of 5 fatty acids absent in healthy larvae (heptanoic, decanoic, adipic, suberic, tridecanoic), the disappearance of 4 compounds (monopalmitoylglycerol, monooleoylglycerol, monostearin, and cholesterol), and changes in the concentrations of 8 compounds. It remains an open question whether substances appearing in the hemolymph of infected insects are a product of the fungus or if they are released from the insect tissues damaged by the growing hyphae.

Immuno-compromised COVID-19 patients being treated with glucocorticoids like Dexamethasone for prolonged periods had led to the surge of mucormycosis. Mucor spores proliferate and spread faster in COVID afflicted persons. In India, more than 51,775 cases and 4300 deaths due to COVID-associated mucormycosis were reported till date. The Government of J&K declared an epidemic of COVID-19 associated mucormycosis under Sect. 2 of the Epidemic Disease Act, 1897. In the backdrop of current COVID scenario, it becomes imperative to study the etiological factors, occurrence of causative agents and aerosolisation of the Mucor spores from the environmental samples leading to mucormycosis. The present study was undertaken to check the presence of Mucor spores in the environment so as to find the reinforcing sources of COVID associated mucormycosis. We isolated three strains of Mucor species: Mucor circinelloides and two Mucor hiemalis species, from environmental samples (air, soil and water), all having a positive relevance to mucormycosis. This is the first study on the presence of mucormycosis causing Mucor species in Kashmir environment. In this study, some pathogenicity marker tests were performed to detect the virulence of these fungal strains such as Anti-fungal susceptibility test (AST), biofilm formation and heat stress test. The in vitro antifungal susceptibility of these species showed the major zone of inhibition by Amphotericin B (Amph) and Nystatin (NS), whereas, Fluconazole (FLC) showed no activity. Biofilm was formed by both Mucor species which was confirmed by crystal violet assay (CV) with positive absorbance readings in microplate reader. Under heat stress, pathogenicity marker test was performed in which both species formed considerable aseptate hyphal growth and small scattered spores. During SEM (Scanning Electron Microscopy) analysis, both the species of Mucor were observed with intact sporangia producing several sporangiospores, whose size dimorphism is linked to its virulence. Thus, augmented knowledge of the epidemiology and clinical presentations of these Mucor species may prevent environmental exposure of immune-compromised patients post-COVID and improve early diagnosis and treatment.