Young mycelia were white and almost hyaline, but inconspicuous zonations were recorded in other cultures. The mycelium was mostly flat and submerged into the substratum. Hyphae were thin-walled and of different diameters ranging from 0.7 to 3.5 mm. The hyphae were highly interwoven, often adhered together and gave the appearance of simple intertwined cords. The hyphae often showed anastomoses and were irregularly septated. They often intertwined and overlapped each other. In older cultures and on the root surface, hyphae were often irregularly inflated, showing a nodose to coralloid Plant Surface Microbiology A. Varma, L. Abbott, D. Werner, R. Hampp (Eds.)

Stem cell has immense potential in regenerative cellular therapy. Mesenchymal stem cells (MSCs) can become a potential attractive candidate for therapy due to its remarkable ability of self-renewal and differentiation into three lineages, i.e., ectoderm, mesoderm, and endoderm. Stem cell holds tremendous promises in the field of tissue regeneration and transplantation for disease treatments. Globally, medicinal plants are being used for the treatment and prevention of a variety of diseases. Phytochemicals like naringin, icariin, genistein, and resveratrol obtained from plants have been extensively used in traditional medicine for centuries. Certain bioactive compounds from plants increase the rate of tissue regeneration, differentiation, and immunomodulation. Several studies show that bioactive compounds from plants have a specific role (bioactive mediator) in regulating the rate of cell division and differentiation through complex signal pathways like BMP2, Runx2, and Wnt. The use of plant bioactive phytochemicals may also become promising in treating diseases like osteoporosis, neurodegenerative disorders, and other tissue degenerative disorders. Thus, the present review article is aimed at highlighting the roles and consequences of plant extracts on MSCs proliferation and desired lineage differentiations.

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At present, few organisms are known to and capable of naturally producing laccases and white rot fungi are one such group. In the present study, three fungal species, namely, Ganoderma lucidum-CDBT1, Ganoderma japonicum, and Lentinula edodes, isolated from their native habitat in Nepal were screened for laccase production, and G. lucidum-CDBT1 was found to express highest levels of enzyme (day 10 culture media showed 0.92 IU/mg total protein or 92 IU/mL laccase activity with ABTS as substrate). Lignin extracted from rice straw was used in Olga medium for laccase production and isolation from G. lucidum-CDBT1. Presence of lignin (5 g/L) and copper sulfate (30 μM) in the media increased the extracellular laccase content by 111% and 114%, respectively. The laccase enzyme produced by G. lucidum-CDBT1 was fractionated by ammonium sulfate and purified by DEAE Sepharose anion exchange chromatography. The purified enzyme was found to have a molecular mass of 43 kDa and exhibits optimal activity at pH 5.0 and 30°C. The isolated laccase was thermally stable for up to 70°C for 1 h and exhibited broad pH stability. The kinetic constants, K m, V max, and K cat, determined using 2,2′-azinobis-(-3-ethylbenzothiazoline-6-sulfonic acid) as substrate were found to be 110 μM, 36 μmol/min/mg, and 246 min−1, respectively. The isolated thermostable laccase will be used in future experiments for delignification process.

Dengue virus is a major health problem in Nepal. The endogenous dengue appeared in 2006 in the country with reported outbreaks in 2010, 2013 and 2016. Eleven years vertical data show there were sporadic cases in all the years and mostly adults between 25 and 40 years of age were infected with dengue virus. Compared with primary infections, secondary infections were observed in relatively larger numbers during the period of 2008-2016. Most of the cases had symptoms of dengue fever; while 7 and 19 cases demonstrated dengue hemorrhagic fever/dengue shock syndrome in 2010 and 2013 respectively. The proportion of dengue hemorrhagic fever amongst all cases of dengue fever was 2.5:4.7% in 2010 and 2013. We found there is shift of serotype from dengue virus serotype-1 (DENV-1) in 2010, DENV-2 in 2013 and DENV-1 in 2016. We feel there is urgent need for better community, hospital and laboratory based surveillance system capable of monitoring the circulating dengue virus (DENV) serotypes in different districts of Nepal. With improvement in surveillance system and efficient management of cases, the case fatality rate due to severe dengue can be reduced.

Phage therapy is one of the most promising alternatives to antibiotics as we face global antibiotic resistance crisis. However, the pharmacokinetics (PK) and pharmacodynamics (PD) of phage therapy are largely unknown. In the present study, we aimed to evaluate the PK/PD of a locally isolated virulent novel øKp_Pokalde_002 (Podoviridae, C1 morphotype) that infects carbapenem-resistant Klebsiella pneumoniae (Kp56) using oral and intraperitoneal (IP) route in a mouse model. The result showed that the øKp_Pokalde_002 rapidly distributed into the systemic circulation within an hour via both oral and IP routes. A higher concentration of phage in plasma was found after 4 h (2.3 x 105 PFU/ml) and 8 h (7.3 x 104 PFU/ml) of administration through IP and oral route, respectively. The phage titer significantly decreased in the blood and other tissues, liver, kidneys, and spleen after 24 h and completely cleared after 72 h of administration. In the Kp56 infection model, the bacterial count significantly decreased in the blood and other organs by 4–7 log10 CFU/ml after 24 h of øKp_Pokalde_002 administration. Elimination half-life of øKp_Pokalde_002 was relatively shorter in the presence of host-bacteria Kp56 compared to phage only, suggesting rapid clearance of phage in the presence of susceptible host. Further, administration of the øKp_Pokalde_002 alone in healthy mice (via IP or oral) did not stimulate pro-inflammatory cytokines (TNF-α and IL-6). Also, treatment with øKp_Pokalde_002 resulted in a significant reduction of pro-inflammatory cytokines (TNF-α and IL-6) caused by bacterial infection, thereby reducing the tissue inflammation. In conclusion, the øKp_Pokalde_002 possess good PK/PD properties and can be considered as a potent therapeutic candidate for future phage therapy in carbapenem-resistant K. pneumoniae infections.

Background & objectives:Epidemiological interventions and mosquito control are the available measures for dengue control. The former approach uses serotype and genetic information on the circulating virus strains. Dengue has been frequently reported from Nepal, but this information is mostly lacking. The present study was done to generate a comprehensive clinical and virological picture of a dengue outbreak in Nepal during 2013.Methods:A hospital-based study involving patients from five districts of Nepal was carried out. Demographic information, clinical details and dengue serological status were obtained. Viral RNA was characterized at the molecular level by reverse-transcription polymerase chain reaction (RT-PCR), nucleotide sequencing and phylogenetic analysis.Results:From among the 2340 laboratory-confirmed dengue cases during the study period, 198 patients consented for the study. Clinically they had fever (100%), headache (59.1%), rashes (18.2%), retro-orbital pain (30.3%), vomiting (15.1%), joint pain (28.8%) and thrombocytopenia (74.3%). Fifteen (7.5%) of them had mucosal bleeding manifestations, and the rest were uncomplicated dengue fever. The patients were mostly adults with a mean age of 45.75 ± 38.61 yr. Of the 52 acute serum samples tested, 15 were positive in RT-PCR. The causative virus was identified as DENV serotype 2 belonging to the Cosmopolitan genotype.Interpretations & conclusions:We report here the involvement of DENV serotype 2 in an outbreak in Nepal in 2013. Earlier outbreaks in the region in 2010 were attributed to serotype 1 virus. As serotype shifts are frequently associated with secondary infections and severe disease, there is a need for enhancing surveillance especially in the monsoon and post-monsoon periods to prevent large-scale, severe dengue outbreaks in the region.

Lignocellulose is one of the most abundant renewable materials that could serve as a source for fermentable sugars. However, its commercial deployment is limited by the fact that lignocellulose is recalcitrant and requires heroic efforts to release sugars. Although very few, there are cellulose degrading organisms in nature that can efficiently degrade the cellulosic biomass and one among them is termite. Termites are responsible for mass turnover of the cellulosic biomass and they accomplish this task by harbouring a consortium of cellulolytic and lignolytic microorganisms. In the present study, we have isolated eight bacterial and five fungal cellulose degrading organisms from the gut of a termite native to Nepal via basal culture medium enrichment with filter paper as substrate. The isolated organisms were identified as Bacillus sp., Cellulomonas sp., Enterobacter sp., and Aspergillus sp. The Congo red screening assay for cellulase production showed the largest zone of hydrolysis (38 mm) for Aspergillus sp. The carboxymethyl cellulose assay revealed that the Bacillus sp (S3B8; 0.12± 0.01 IU/ml obtained within96 hours) and Aspergillus sp. (S3F3; 0.07 ±0.01 IU/ml obtained within 168 hours) were most efficient cellulase producers. The saccharification and fermentation of lignocellulosic biomass to ethanol was successfully achieved via culturing of selected bacterial [Bacillus (S3B8)] and fungal isolates [Aspergillus (S3F3)] and then fermentation with Saccharomyces cerevisiae (S2Y1). Further characterization of these organisms and optimization of fermentation of lignocellulosic biomass is ongoing.