First report of Phytopythium vexans causing gummosis and root rot of Khasi Mandarin (Citrus reticulata Blanco) in north eastern states of India.

Khasi mandarin (Citrus reticulata Blanco) is the most economically important crop among the citrus growing region in the north-eastern India (Singh et al. 2016). An extensive survey was conducted to identify the causal agent of citrus root rot and gummosis in north eastern states (Meghalaya, Tripura, Manipur, Arunachal Pradesh, Sikkim, Nagaland and Assam) of India during October 2021-23. The gummosis disease incidence ranged from 5 to 95 % in 10 to 25 years old Khasi mandarin plants showing relatively more chronic symptoms on mature trees. Yellowing and dropping of leaves, twigs die back, gum oozing from infected bark and loss of feeder roots were the typical symptoms of the disease. Infected bark tissue and young lemon leaf baits in rhizosphere soil were plated on corn meal agar medium supplemented with pimaricin (10 µg/ml), ampicillin (250 µg/ml), rifamycin (10 µg/ml) and 300µg/ml carbendazim and incubated at 26℃. Fifty isolates were purified and maintained on Carrot agar medium. These isolates showed similar cultural and morphological characteristics. Two representative isolates from Arunachal Pradesh (AP21 and AP26) were selected for further experiments and deposited to Indian Type culture collection (ITCC), New Delhi with accession no. 9156 and 9157 respectively. The colonies were fast growing, showing rosette pattern along with whitish blooming mycelium appearance with no visual sporulation at the surface. The hyphae were coenocytic with initially right-angled branching. Sporangia were globose or sub globose and papillated. Oogonia were smooth and globose (16.29-21.09 µm) in diameter. Antheridia were irregular, cylindrical and broadly attached to oogonia. Empty sporangia were also observed. Multilocus phylogenetic analysis using internal transcribed spacer region (Das et al. 2011), ß tubulin (Blair et al. 2008) and Cytochrome oxidase II gene (Noireung et al. 2020) showed that these isolates formed a stable clade with Phytopythium vexans (CBS119.80) sequence retrieved from NCBI database. BLAST analysis showed that ITS sequence of AP21 (OQ372986) and AP26 (OQ381083) had >99 % identity with P. vexans isolate NS-3 (ON533631). Further, BLAST analysis of ß tubulin (AP21 OQ446053, AP26 OR405377) and Cox II gene (AP21 OQ473414, AP26 OR552422) sequences showed that our Indian isolates showed >99 % similarity with P. vexans voucher strain CBS119.80. To fulfil Koch's postulates, Khasi mandarin (Citrus reticulata) seedlings were inoculated by adding 100 ml zoospore suspension of P. vexans (1x105 spores/ml) in sterilized soil (Thao et al. 2020). The experiment was carried out in triplicate. Yellowing of leaves and leaf drop were observed 7 days post inoculation while 30 days post inoculation, treated plants started showing symptoms of root rot, including mild root decay. No symptoms were observed in control treatment. The pathogen was reisolated from symptomatic roots and confirmed through colony and sporangium morphology. Recently, it was reported that P. vexans is associated with apple and pear decline in the Saiss plain of Morocco (Jabiri et al. 2021), root rot on mandarin in Thailand (Noireung et al. 2020) and on Durian in Vietnam (Thao et al. 2020). As per our knowledge, this is the first report of P. vexans causing root rot and gummosis in Khasi mandarin from north eastern states of India. This finding is significantly important for the development of a successful disease management strategy in India.

Evaluating the potential of ethyl cellulose/eudragit-based griseofulvin loaded nanosponge matrix for topical antifungal drug delivery in a sustained release pattern.

Fungal infections are very alarming nowadays and are common throughout the world. Severe fungal infections may lead to a significant risk of mortality and morbidity worldwide. Sustained delivery of antifungal agents is needed to mitigate this problem. In the current study, an attempt has been made to formulate griseofulvin-loaded nanosponges using the quasi-emulsion solvent diffusion technique. For characterization, griseofulvin loaded nanosponges were tested by different instrumental techniques such as optical microscopy, scanning electron microscopy (SEM), powder X-ray diffractometer (PXRD), Fourier transform infrared spectroscopy (FT-IR), differential scanning calorimetry (DSC), and transmission electron microscopy (TEM). The antifungal activity of the nanosponges was assessed against Candida albican strain using the agar well-diffusion method. Finally, the drug-loaded nanosponges' in vitro sustained release activity was evaluated. FTIR spectra showed no chemical interference between the drug and polymers. Some of the peaks of the drug are not visible in the FTIR spectrum, which suggests drug entrapment. PXRD data showed that the drug lost its high crystallinity when entrapped in the nanosponge matrix. From the morphological studies via SEM and TEM, a brief idea of the surface morphology of the nanosponges was obtained. The small pores throughout the structure proved its high porosity. The antifungal sensitivity assay was successful, and a zone of inhibition was observed in all the formulations. The in-vitro drug release study showed sustained behaviour. The sustaining effect was due to the polymer and cross-linker used, which gave rise to a porous scaffold matrix. From the results, it can be concluded that griseofulvin-loaded nanosponges can be used for antifungal drug delivery against various topical skin infections.

Characterizing genetic diversity of Sclerotium rolfsii isolates by biomapping of mycelial compatibility groupings and multilocus sequence analysis.

Genetic diversity in Sclerotium rolfsii is useful for understanding its population structure, identifying different mycelial compatibility groups (MCGs), and developing targeted strategies for disease management in affected crops. In our study, a comprehensive genetic analysis was conducted on 50 isolates of S. rolfsii, collected from various geographic regions and host plants. Two specific genes, TEF1α and RPB2, were utilized to assess the genetic diversity and relationships among these isolates. Notably, out of 1225 pairings examined, only 154 exhibited a compatible reaction, while the majority displayed antagonistic reactions, resulting in the formation of a barrier zone. The isolates were grouped into 10 distinct MCGs. These MCGs were further characterized using genetic sequencing. TEF1α sequences distinguished the isolates into 17 distinct clusters, and RPB2 sequences classified them into 20 clusters. Some MCGs shared identical gene sequences within each gene, while others exhibited unique sequences. Intriguingly, when both TEF1α and RPB2 sequences were combined, all 10 MCGs were effectively differentiated, even those that appeared identical with single-gene analysis. This combined approach provided a comprehensive understanding of the genetic diversity and relationships among the S. rolfsii isolates, allowing for precise discrimination between different MCGs. The results shed light on the population structure and genetic variability within this plant pathogenic fungus, providing valuable insights for disease management and control strategies. This study highlights the significance of comprehending the varied virulence characteristics within S. rolfsii isolates, categorizing them into specific virulence groups based on disease severity index (DSI) values. The association with MCGs provides additional insights into the genetic underpinnings of virulence in this pathogen. Furthermore, the identification of geographical patterns in virulence implies the influence of region-specific factors, with potential implications for disease control and crop protection strategies.Please confirm if the author names are presented accurately and in the correct sequence (given name, middle name/initial, family name). Author 1 Given name: [G. M. Sandeep] Last name [Kumar]. Author 2 Given name: [Praveen Kumar] Last name [Singh]. Also, kindly confirm the details in the metadata are correct.I confirm that the given names are accurate and presented in the correct sequence.

Autonomous Image-Based Corrosion Detection in Steel Structures Using Deep Learning.

Steel structures are susceptible to corrosion due to their exposure to the environment. Currently used non-destructive techniques require inspector involvement. Inaccessibility of the defective part may lead to unnoticed corrosion, allowing the corrosion to propagate and cause catastrophic structural failure over time. Autonomous corrosion detection is essential for mitigating these problems. This study investigated the effect of the type of encoder-decoder neural network and the training strategy that works the best to automate the segmentation of corroded pixels in visual images. Models using pre-trained DesnseNet121 and EfficientNetB7 backbones yielded 96.78% and 98.5% average pixel-level accuracy, respectively. Deeper EffiecientNetB7 performed the worst, with only 33% true-positive values, which was 58% less than ResNet34 and the original UNet. ResNet 34 successfully classified the corroded pixels, with 2.98% false positives, whereas the original UNet predicted 8.24% of the non-corroded pixels as corroded when tested on a specific set of images exclusive to the investigated training dataset. Deep networks were found to be better for transfer learning than full training, and a smaller dataset could be one of the reasons for performance degradation. Both fully trained conventional UNet and ResNet34 models were tested on some external images of different steel structures with different colors and types of corrosion, with the ResNet 34 backbone outperforming conventional UNet.

Renal Thrombotic Microangiopathy: A Complication of Paediatric Systemic Lupus Erythematosus That Requires Attention.

Systemic lupus erythematosus (SLE) is a multi-system disorder with a variety of clinical presentations. A wide range of renal vascular lesions (VL) is described predominantly in adult patients. The exact prevalence of renal VL in the pediatric SLE (pSLE) population is yet to be determined. A 10-year-old female patient with lupus nephritis (LN) presented with deteriorating kidney function. An exhaustive array of clinical-biochemical and pathological evaluations resulted in a diagnosis of class IV LN with thrombotic microangiopathy (TMA) associated with malignant hypertension and hypocomplementemia. Renal VL is overlooked or underreported in SLE patients, as it is neither accorded much importance in the International Society of Nephrology/Renal Pathology Society (ISN/RPS) classification nor in the activity and chronicity scoring. The TMA lesions in LN patients can be managed following the recently devised PLASMIC score; hence, reporting such VL has therapeutic implications.

Metabolomic profiling and its association with the bio-efficacy of Aspergillus niger strain against Fusarium wilt of guava.

Bio-control agents are the best alternative to chemicals for the successful management of plant diseases. The fungus Aspergillus niger is known to produce diverse metabolites with antifungal activity, attracting researchers to exploit it as a bio-control agent for plant disease control. In the present study, 11 A. niger strains were isolated and screened for their antagonism against the guava wilt pathogen under in vitro and in planta conditions. Strains were identified morphologically and molecularly by sequencing the internal transcribed spacer (ITS), ß-tubulin, and calmodulin genes. The strains were evaluated through dual culture, volatile, and non-volatile methods under an in vitro study. AN-11, AN-6, and AN-2 inhibited the test pathogen Fusarium oxysporum f. sp. psidii (FOP) at 67.16%, 64.01%, and 60.48%, respectively. An in planta study was conducted under greenhouse conditions with 6 months old air-layered guava plants (var. Allahabad Safeda) by pre- and post-inoculation of FOP. The AN-11 strain was found to be effective under both pre- and post-inoculation trials. Furthermore, gas chromatography-mass spectrometry (GC-MS) analysis was carried out to characterize the volatile compounds of the most potential strain, A. niger. The hexane soluble fraction showed the appearance of characteristic peaks of hexadecenoic acid methyl ester (4.41%), 10-octadecanoic acid methyl ester (3.79%), dodecane (3.21%), undecane (3.19%), gibepyrone A (0.15%), 3-methylundecane (0.36%), and citroflex A (0.38%). The ethyl acetate fraction of the bio-control fungi revealed the occurrence of major antifungal compounds, such as acetic acid ethyl ester (17.32%), benzopyron-4-ol (12.17%), 1,2,6-hexanetriol (7.16%), 2-propenoic acid ethanediyl ester (2.95%), 1-(3-ethyloxiranyl)-ethenone (0.98%), 6-acetyl-8-methoxy dimethyl chromene (0.96%), 4-hexyl-2,5-dihydro dioxo furan acetic acid (0.19%), and octadecanoic acid (1.11%). Furthermore, bio-control abilities could be due to hyper-parasitism, the production of secondary metabolites, and competition for sites and nutrients. Indeed, the results will enrich the existing knowledge of metabolomic information and support perspectives on the bio-control mechanism of A. niger.

Transcription Factors YAP/TAZ and SRF Cooperate To Specify Renal Myofibroblasts in the Developing Mouse Kidney.

BACKGROUND: The embryonic renal stroma consists of multiple molecularly distinct cell subpopulations, the functional significance of which is largely unknown. Previous work has demonstrated that the transcription factors YAP and TAZ play roles in the development and morphogenesis of the nephrons, collecting ducts, and nephron progenitor cells. METHODS: In embryonic mouse kidneys, we identified a subpopulation of stromal cells with enriched activity in YAP and TAZ. To evaluate the function of these cell types, we genetically ablated both Yap and Taz from the stromal progenitor population and examined how gene activity and development of YAP/TAZ mutant kidneys are affected over a developmental time course. RESULTS: We found that YAP and TAZ are active in a subset of renal interstitium and that stromal-specific coablation of YAP/TAZ disrupts cortical fibroblast, pericyte, and myofibroblast development, with secondary effects on peritubular capillary differentiation. We also demonstrated that the transcription factor SRF cooperates with YAP/TAZ to drive expression of at least a subset of renal myofibroblast target genes and to specify myofibroblasts but not cortical fibroblasts or pericytes. CONCLUSIONS: These findings reveal a critical role for YAP/TAZ in specific embryonic stromal cells and suggest that interaction with cofactors, such as SRF, influence the expression of cell type-specific target genes, thus driving stromal heterogeneity. Further, this work reveals functional roles for renal stroma heterogeneity in creating unique microenvironments that influence the differentiation and maintenance of the renal parenchyma.

Significance of Cardiac Magnetic Resonance Feature Tracking of the Right Ventricle in Predicting Subclinical Dysfunction in Patients with Thalassemia Major.

In patients with thalassemia major (TM), cardiac magnetic resonance feature-tracking (CMR-FT) has been shown to be an effective method for diagnosing subclinical left ventricular (LV) dysfunction. This study aimed to determine whether CMR-FT could detect abnormal RV dysfunction in patients with a normal right ventricular ejection fraction (RVEF). We performed a retrospective analysis of TM patients admitted to Dubai's Rashid Hospital between July 2019 and March 2021. The inclusion criteria were TM patients with SSFP cine with T2* (T2*-weighted imaging), while exclusion criteria included any other cardiovascular disease. When there was no myocardial iron overload (MIO) (T2* ≥ 20 ms) and when there was significant MIO (T2* < 20 ms), the CMR-FT was used to correlate with EF. Among the 89 participants, there were 46 men (51.7%) and 43 women (48.3%), with a mean age of 26.14 ± 7.4 years (range from 10 to 48 years). Forty-six patients (51.69%) did not have MIO, while 43 individuals did (48.31%). Thirty-nine patients (32.6%) were diagnosed with severe MIO, while seventeen (19.1%) were diagnosed with mild to moderate MIO. A significant correlation existed between RVEF and T2* values (r = 0.274, p = 0.014) and between left ventricular ejection fraction (LVEF) and T2* values (r = 0.256, p = 0.022). Using a multiple logistic regression model with predictors such as right ventricular longitudinal strain (RVGLS), LV ejection fraction (LV EF), and hemoglobin, abnormal myocardial iron overload can be predicted. This model demonstrates an AUC of 78.3%, a sensitivity of 72%, and a specificity of 76%. In the group with preserved RVEF > 53%, the left ventricular radial strain (LVGRS) (p = 0.001), right ventricular radial strain (RVGRS) (p = 0.000), and right ventricular basal circumferential strain (RVGCS-basal) (p = 0.000) CMR-FT strain values are significantly lower than those of the control group (p > 0.05). There was no significant correlation between the LVGLS and T2*. RVGLS was ranked among the most accurate predictors of abnormal myocardial iron overload. The LVGRS, RVGRS, and RVGCS-basal CMR-FT strain values were the best predictors of subclinical RV dysfunction in the group with preserved RVEF. The most accurate way to diagnose MIO is still T2*, but FT-strain can help us figure out how MIO affects the myocardium from a pathophysiological point of view.

Aluminium complexes: next-generation catalysts for selective hydroboration.

Organoboranes obtained from hydroboration reactions are one of the important classes of compounds that could be used to provide valuable synthons for follow-up transformations such as various functional group incorporation or C-C bond forming reactions. For decades, various transition metals were utilised as catalysts in such transformations. Recently Earth-abundant and less toxic main group metals have revived their importance in hydroboration chemistry, among which the suitable candidates are aluminium complexes as catalysts. In this regard, the development of aluminium complexes to achieve more robust catalytic systems with greater efficiency is appreciable.

Rh(I)-catalysed imine-directed C-H functionalization via the oxidative [3 + 2] cycloaddition of benzylamine derivatives with maleimides.

The Rh(I)-catalysed imine-directed oxidative [3 + 2] cycloaddition of benzylamines with maleimides is reported. A wide range of both benzylamines and maleimides is applicable to the reaction. A one-pot three component strategy using benzylamines, 2-pyridinecarboxaldehyde, and maleimides is successfully achieved. Mechanistic studies including deuterium labelling experiments suggest that a zwitterionic intermediate is formed and is a key intermediate through the Rh-catalysed activation of a benzylic C(sp3)-H bond of the imine.

HO-3867 Induces ROS-Dependent Stress Response and Apoptotic Cell Death in Leishmania donovani.

Lack of vaccine and increasing chemotherapeutic toxicities currently necessitate the development of effective and safe drugs against various forms of leishmaniases. We characterized the cellular stress induced by a novel curcumin analogue, HO-3867, encapsulated within the phosphatidylcholine-stearylamine (PC-SA) liposome for the first time against Leishmania. The liposomal formulation of HO-3867 (i.e., PC-SA/HO-3867) initiated oxidative stress-induced apoptosis in L. donovani, revealed by altered cell morphology, phosphatidylserine externalization, mitochondrial depolarization, intracellular lipid accumulation, and cell cycle arrest in promastigotes. Liposomal HO-3867 was observed to be a strong apoptosis inducer in L. donovani and L. major in a dose-dependent manner, yet completely safe for normal murine macrophages. Moreover, PC-SA/HO-3867 treatment induced L. donovani metacaspase and PARP1 activation along with downregulation of the Sir2 gene. PC-SA/HO-3867 arrested intracellular L. donovani amastigote burden in vitro, with reactive oxygen species (ROS) and nitric oxide (NO)-mediated parasite killing. These data suggest that liposomal HO-3867 represents a highly promising and non-toxic nanoparticle-based therapeutic platform against leishmaniasis inspiring further preclinical developments.

Transovarial Transmission of Dolichos Yellow Mosaic Virus by Its Vector, Bemisia tabaci Asia II 1.

The cultivation of dolichos bean [Lablab purpureus (L.) Sweet] has been severely affected by dolichos yellow mosaic virus (DoYMV, Begomovirus) transmitted by whitefly, Bemisia tabaci (Hemiptera: Aleyrodidae). We tested the transovarial transmission of DoYMV in next-generation B. tabaci by PCR, real-time PCR, Southern blot hybridization, and biological transmission. The eggs, laid by DoYMV-exposed B. tabaci, carry the virus in a unique pattern. Only the eggs laid in between 3 and 6 days post virus acquisition by a parent B. tabaci were DoYMV positive. When tested individually in real-time PCR, around 31-53% of the eggs carried the virus. The presence of DoYMV in ovaries and F1 eggs was further substantiated by the hybridization of a Cy3-conjugated nucleic acid probe complementary to the viral strand of DoYMV. Viral DNA was also detected in F1 adults and F2 eggs. B. tabaci progenies carried not only the DoYMV DNA but were also infective. The F1 adults transmitted DoYMV to all tested plants and produced strong yellow mosaic symptoms. An increase in viral copies from egg to nymphal stage indicated propagation of DoYMV in B. tabaci. However, the increase was for a short period and decreased thereafter. The present study provides the first evidence of transovarial transmission and propagation of a bipartite begomovirus in its vector, B. tabaci Asia II 1. The transovarial transmission and replication of DoYMV in B. tabaci have great epidemiological relevance as B. tabaci can serve as a major host of the virus to bridge the gap between the cropping seasons.

Nanovaccine: an emerging strategy.

INTRODUCTION: Vaccination is so far the most effective way of eradicating infections. Rapidly emerging drug resistance against infectious diseases and chemotherapy-related toxicities in cancer warrant immediate vaccine development to save mankind. Subunit vaccines alone, however, fail to elicit sufficiently strong and long-lasting protective immunity against deadly pathogens. Nanoparticle (NP)-based delivery vehicles like microemulsions, liposomes, virosomes, nanogels, micelles and dendrimers offer promising strategies to overcome limitations of traditional vaccine adjuvants. Nanovaccines can improve targeted delivery, antigen presentation, stimulation of body's innate immunity, strong T cell response combined with safety to combat infectious diseases and cancers. Further, nanovaccines can be highly beneficial to generate effective immutherapeutic formulations against cancer. AREAS COVERED: This review summarizes the emerging nanoparticle strategies highlighting their success and challenges in preclinical and clinical trials in infectious diseases and cancer. It provides a concise overview of current nanoparticle-based vaccines, their adjuvant potential and their cellular delivery mechanisms. EXPERT OPINION: The nanovaccines (50-250 nm in size) are most efficient in terms of tissue targeting, prolonged circulation and preferential uptake by the professional APCs chiefly due to their small size. More rational designing, improved antigen loading, extensive functionalization and targeted delivery are some of the future goals of ideal nanovaccines.

COVID-19 associated rhino-orbital-cerebral mucormycosis: An observational study from Eastern India, with special emphasis on neurological spectrum.

AIMS: 1: Describe the epidemiology and determine risk factors for COVID-19 associated mucormycosis. 2: Elaborate the clinical spectrum of Rhino-Orbital-Cerebral Mucormycosis (ROCM), pattern of neuroaxis involvement and it's radiological correlates. METHODS: Observational study. Consecutive, confirmed cases of mucormycosis (N = 55) were included. A case of mucormycosis was defined as one who had clinical and radiological features consistent with mucormycosis along with demonstration of the fungus in tissue via KOH mount/culture/histopathological examination (HPE). Data pertaining to epidemiology, risk factors, clinico-radiological features were analysed using percentage of total cases. RESULTS: Middle aged, diabetic males with recent COVID-19 infection were most affected. New onset upper jaw toothache was a striking observation in several cases. Among neurological manifestations headache, proptosis, vision loss, extraocular movement restriction; cavernous sinus, meningeal and parenchymal involvement were common. Stroke in ROCM followed a definitive pattern with watershed infarction. CONCLUSIONS: New onset upper jaw toothache and loosening of teeth should prompt an immediate search for mucormycosis in backdrop of diabetic patients with recent COVID-19 disease, aiding earlier diagnosis and treatment initiation. Neuroaxis involvement was characterized by a multitude of features pertaining to involvement of optic nerve, extraocular muscles, meninges, brain parenchyma and internal carotid artery.

Pyrimidine-directed metal-free C-H borylation of 2-pyrimidylanilines: a useful process for tetra-coordinated triarylborane synthesis.

Convenient, easily handled, laboratory friendly, robust approaches to afford synthetically important organoboron compounds are currently of great interest to researchers. Among the various available strategies, a metal-free approach would be overwhelmingly accepted, since the target boron compounds can be prepared in a metal-free state. We herein present a detailed study of the metal-free directed ortho-C-H borylation of 2-pyrimidylaniline derivatives. The approach allowed us to synthesize various boronates, which are synthetically important compounds and various four-coordinated triarylborane derivatives, which could be useful in materials science as well as Lewis-acid catalysts. This metal-free directed C-H borylation reaction proceeds smoothly without any interference by external impurities, such as inorganic salts, reactive functionalities, heterocycles and even transition metal precursors, which further enhance its importance.

Rh(i)- and Rh(ii)-catalyzed C-H alkylation of benzylamines with alkenes and its application in flow chemistry.

The Rh-catalyzed C-H alkylation of benzylamines with alkenes using a picolinamide derivative as a directing group is reported. Both Rh(i) and Rh(ii) complexes can be used as active catalysts for this transformation. In addition, a flow set up was designed to successfully mimic this process under flow conditions. Several examples are presented under flow conditions and it was confirmed that a flow process is advantageous over a batch process. Deuterium labelling experiments were performed to elucidate the mechanism of the reaction, and the results indicated a possible carbene mechanism for this C-H alkylation process.

Rh(II)-Catalyzed C-H Alkylation of Benzylamines with Unactivated Alkenes: The Influence of Acid on Linear and Branch Selectivity.

The Rh-catalyzed C-H alkylation of benzylamine derivatives with unactivated 1-alkenes that proceeds via a picolinamide directing group is reported. The crucial role of an acid additive in this transformation is confirmed. Aromatic acids showed high linear selectivity, and aliphatic acids provided branched alkylation products as the major product. The reaction has a broad scope for benzylamines and alkenes. Deuterium labeling experiments suggest that a Rh-carbene intermediate is involved in the case of linear product formation. A different reaction pathway, however, appears to be involved in the case of branched alkylation products, and this pathway also appeared to be a minor pathway in linear-selective reactions.

Chemo-profiling of bioactive metabolites from Chaetomium globosum for biocontrol of Sclerotinia rot and plant growth promotion.

Chaetomium globosum Kunze ex. Fries has been known to produce diverse bioactive metabolites, attracting researchers to exploit the biocontrol agent for plant disease management. However, distinct research gaps are visible regarding detail characterization of bioactive metabolites. Thus the current study has been planned to characterize volatile and nonvolatile compounds of most potential strain of C. globosum 5157. GC-MS analysis of hexane fraction revealed twenty-six volatile organic compounds, representing 65.5% of total components in which 3-octanone (21.4%) was found to be most abundant. UPLC-QTOF-MS/MS analysis of ethyl acetate and methanolic fractions resulted tentative characterization of fifteen and eleven metabolites, respectively. Among these, nine metabolites were isolated, purified and characterized using 1H NMR and High resolution mass spectrometric analysis to delineate mass fragmentation pattern for the first time. Antifungal potential of hexane fraction exhibited high inhibitory action against Sclerotium rolfsii (139.2 µg mL-1) whereas ethyl acetate fraction was highly effective against Sclerotinia sclerotiorum (112.1 µg mL-1). Comparative assessment of C. globosum 5157 vis a vis Trichoderma harzianum A28 revealed promising effect of C. globosum 5157 with respect to antifungal properties and plant growth promotion of Brassica seedlings.

An Automated High-Throughput Fluorescence In Situ Hybridization (FISH) Assay Platform for Use in the Identification and Optimization of siRNA-Based Therapeutics.

Since the revolutionary discovery of RNA interference (RNAi) more than 20 years ago, synthetic small interfering RNAs (siRNAs) have held great promise as therapeutic agents for treating human diseases by the specific knockdown of disease-causing gene products. To facilitate the development of siRNA therapeutics, a robust, high-throughput in vitro assay for measuring gene silencing is imperative during the initial siRNA lead sequence identification and, later, during the lead optimization with chemically modified siRNAs. There are several potential assays for measuring gene expression. Quantitative reverse transcription PCR (qRT-PCR) has been widely used to quantitate messenger RNA (mRNA). This method has a few disadvantages, however, such as the requirement for RNA isolation, complementary DNA (cDNA) generation, and PCR reaction, which are labor-intensive, limit the assay throughput, and introduce variability. We chose a high-content imaging assay, bDNA FISH, that combines the branched DNA (bDNA) technology with fluorescence in situ hybridization (FISH) to measure gene silencing by siRNAs because it is sensitive and robust with a short reagent procurement and assay development time. We also built a fully automated liquid-handling platform for executing bDNA FISH assays to increase throughput, and the system has a capacity of generating 192 concentration-response curves in a single run. We have successfully developed and executed the bDNA FISH assays for multiple targets using this automated platform to identify and optimize siRNA candidate molecules. Examples of the bDNA FISH assay for selected targets are presented.

Identification and characterization of cellular heterogeneity within the developing renal interstitium.

Kidney formation requires the coordinated growth of multiple cell types including the collecting ducts, nephrons, vasculature and interstitium. There is a long-held belief that interactions between progenitors of the collecting ducts and nephrons are primarily responsible for kidney development. However, over the last several years, it has become increasingly clear that multiple aspects of kidney development require signaling from the interstitium. How the interstitium orchestrates these various roles is poorly understood. Here, we show that during development the interstitium is a highly heterogeneous patterned population of cells that occupies distinct positions correlated to the adjacent parenchyma. Our analysis indicates that the heterogeneity is not a mere reflection of different stages in a linear developmental trajectory but instead represents several novel differentiated cell states. Further, we find that ß-catenin has a cell autonomous role in the development of a medullary subset of the interstitium and that this non-autonomously affects the development of the adjacent epithelia. These findings suggest the intriguing possibility that the different interstitial subtypes may create microenvironments that play unique roles in development of the adjacent epithelia and endothelia.