Application of flow cytometry for rapid, high-throughput, multiparametric analysis of environmental microbiomes.

Quantification of the abundance and understanding of the dynamics of the microbial communities is essential to establish a basis for microbiome characterization. The conventional techniques used for the quantification of microbes are complicated and time-consuming. With scientific advancement, many techniques evolved and came into account. Among them, flow cytometry is a robust, high-throughput technique through which microbial dynamics, morphology, microbial distribution, physiological characteristics, and many more attributes can be studied in a high-throughput manner with comparatively less time and resources. Flow cytometry, when combined with other omics-based methods, offers a rapid and efficient platform to analyze and understand the composition of microbiome at the cellular level. The microbial diversity observed through flow cytometry will not be equivalent to that obtained by sequencing methods, but this integrated approach holds great potential for high throughput characterization of microbiomes. Flow cytometry is regarded as an established characterization tool in haematology, oncology, immunology, and medical microbiology research; however, its application in environmental microbiology is yet to be explored. This comprehensive review aims to delve into the diverse environmental applications of flow cytometry across various domains, including but not limited to bioremediation, landfills, anaerobic digestion, industrial bioprocesses, water quality regulation, and soil quality regulation. By conducting an in-depth analysis, this article seeks to shed light on the potential benefits and challenges associated with the utilization of flow cytometry in addressing environmental concerns.

Modifications of native lotus (Nelumbo nucifera G.) rhizome starch and its overall characterization: A review.

Lotus (Nelumbo nucifera G.) rhizomes are an under-utilized and sustainable starch source that constitutes up to 20 % starch. The review mainly focused on the extraction methods of starch, the chemical composition of LRS, and techno-functional characteristics such as swelling power, solubility, in vitro digestibility, pasting property, and gelatinization is highlighted in LRS review. Lotus rhizome starch (LRS) is also used as a water retention agent, thickening, gelling, stabilizing, and filling in food and non-food applications. Native starch has limited functional characteristics in food applications so by modifying the starch, functional characteristics are enhanced. Single and dual treatment processes are available to enhance microstructural properties, resistant starch, techno-functional, morphological, and, film-forming properties. Compared with other starch sources, there is a lack of systematic information on the LRS. Many industries are interested in developing food products based on starch such as nanoparticles, hydrogels, edible films, and many others. Additionally, there are several recommendations to improve the applications in the food industry. Finally, we provide an outlook on the future possibility of LRS.

Effect of fungal pretreatment by Pycnoporus sanguineus and Trichoderma longibrachiatum on the anaerobic digestion of rice straw.

Rice straw is composed of complex lignocellulosic biomass, representing a major obstacle in its conversion to bioenergy. The objective of this study was to evaluate the usefulness of less explored fungal strains Trichoderma longibrachiatum (TL) and Pycnoporus sanguineus (PS) in improving hydrolysis and bioavailability of rice straw in anaerobic digestion (AD). The fungal treatment of rice straw for 10 days by PS and TL increased biogas production by 20.79% and 17.85% and reduced soluble chemical oxygen demand (sCOD) by 71.43% and 64.70%, respectively. The AD samples containing fungal-treated rice straw showed higher lignocellulolytic enzyme activities contributing to better process performance. The taxonomic profile of microbial communities in treated samples showed increased diversity that could sustain consistent system performance and exhibit enhanced resilience against pH fluctuations. Metagenomic analysis revealed 60.82% increase in Proteobacteria in PS and 11.58% increase in Bacteroidetes in TL-treated rice straw samples resulting in improved hydrolysis.

Scientific characterization methods for better utilization of cattle dung and urine: a concise review.

Cattle are usually raised for food, manure, leather, therapeutic, and draught purposes. Biowastes from cattle, such as dung and urine, harbor a diverse group of crucial compounds, metabolites/chemicals, and microorganisms that may benefit humans for agriculture, nutrition, therapeutics, industrial, and other utility products. Several bioactive compounds have been identified in cattle dung and urine, which possess unique properties and may vary based on agro-climatic zones and feeding practices. Therefore, cattle dung and urine have great significance, and a balanced nutritional diet may be a key to improved quality of these products/by-products. This review primarily focuses on the scientific aspects of biochemical and microbial characterization of cattle biowastes. Various methods including genomics for analyzing cattle dung and gas chromatography-mass spectroscopy for cattle urine have been reviewed. The presented information might open doors for the further characterization of cattle resources for heterogeneous applications in the production of utility items and addressing research gaps. Methods for cattle's dung and urine characterization.

Genetics of diabetes.

Diabetes mellitus is a complicated disease characterized by a complex interplay of genetic, epigenetic, and environmental variables. It is one of the world's fastest-growing diseases, with 783 million adults expected to be affected by 2045. Devastating macrovascular consequences (cerebrovascular disease, cardiovascular disease, and peripheral vascular disease) and microvascular complications (like retinopathy, nephropathy, and neuropathy) increase mortality, blindness, kidney failure, and overall quality of life in individuals with diabetes. Clinical risk factors and glycemic management alone cannot predict the development of vascular problems; multiple genetic investigations have revealed a clear hereditary component to both diabetes and its related complications. In the twenty-first century, technological advancements (genome-wide association studies, next-generation sequencing, and exome-sequencing) have led to the identification of genetic variants associated with diabetes, however, these variants can only explain a small proportion of the total heritability of the condition. In this review, we address some of the likely explanations for this "missing heritability", for diabetes such as the significance of uncommon variants, gene-environment interactions, and epigenetics. Current discoveries clinical value, management of diabetes, and future research directions are also discussed.

Quantum Circuit Components for Cognitive Decision-Making.

This paper demonstrates that some non-classical models of human decision-making can be run successfully as circuits on quantum computers. Since the 1960s, many observed cognitive behaviors have been shown to violate rules based on classical probability and set theory. For example, the order in which questions are posed in a survey affects whether participants answer 'yes' or 'no', so the population that answers 'yes' to both questions cannot be modeled as the intersection of two fixed sets. It can, however, be modeled as a sequence of projections carried out in different orders. This and other examples have been described successfully using quantum probability, which relies on comparing angles between subspaces rather than volumes between subsets. Now in the early 2020s, quantum computers have reached the point where some of these quantum cognitive models can be implemented and investigated on quantum hardware, by representing the mental states in qubit registers, and the cognitive operations and decisions using different gates and measurements. This paper develops such quantum circuit representations for quantum cognitive models, focusing particularly on modeling order effects and decision-making under uncertainty. The claim is not that the human brain uses qubits and quantum circuits explicitly (just like the use of Boolean set theory does not require the brain to be using classical bits), but that the mathematics shared between quantum cognition and quantum computing motivates the exploration of quantum computers for cognition modeling. Key quantum properties include superposition, entanglement, and collapse, as these mathematical elements provide a common language between cognitive models, quantum hardware, and circuit implementations.

Biosynthesis of Silver Nanoparticles Utilizing Leaf Extract of Trigonella foenum-graecum L. for Catalytic Dyes Degradation and Colorimetric Sensing of Fe3+/Hg2.

The aqueous Trigonella foenum-graecum L. leaf extract belonging to variety HM 444 was used as reducing agent for silver nanoparticles (AgNPs) synthesis. UV-Visible spectroscopy, Particle size analyser (PSA), Field emission scanning electron microscopy coupled to energy dispersive X-ray spectroscopy (FESEM-EDX) and High-resolution transmission electron microscopy (HRTEM) were used to characterize AgNPs. Selected area electron diffraction (SAED) confirmed the formation of metallic Ag. Fourier Transform Infrared Spectroscopy (FTIR) was done to find out the possible phytochemicals responsible for stabilization and capping of the AgNPs. The produced AgNPs had an average particle size of 21 nm, were spherical in shape, and monodispersed. It showed catalytic degradation of Methylene blue (96.57%, 0.1665 ± 0.03 min-1), Methyl orange (71.45%, 0.1054 ± 0.002 min-1), and Rhodamine B (92.72%, 0.2004 ± 0.01 min-1). The produced AgNPs were excellent solid bio-based sensors because they were very sensitive to Hg2+ and Fe3+ metal ions with a detection limit of 11.17 µM and 195.24 µM, respectively. From the results obtained, it was suggested that aqueous leaf extract demonstrated a versatile and cost-effective method and should be utilized in future as green technology for the fabrication of nanoparticles.

Phytofabrication of Silver Nanoparticles Using Trigonella foenum-graceum L. Leaf and Evaluation of Its Antimicrobial and Antioxidant Activities.

Silver nanoparticles (AgNPs) were fabricated using Trigonella foenum-graceum L. leaf extract, belonging to the variety HM 425, as leaf extracts are a rich source of phytochemicals such as polyphenols, flavonoids, and sugars, which function as reducing, stabilizing, and capping agents in the reduction of silver ions to AgNPs. These phytochemicals were quantitatively determined in leaf extracts, and then, their ability to mediate AgNP biosynthesis was assessed. The optical, structural, and morphological properties of as-synthesized AgNPs were characterized using UV-visible spectroscopy, a particle size analyzer (PSA), FESEM (field emission scanning electron microscopy), HRTEM (high-resolution transmission electron microscopy), and FTIR (Fourier transform infrared spectroscopy). HRTEM analysis demonstrated the formation of spherically shaped AgNPs with a diameter of 4-22 nm. By using the well diffusion method, the antimicrobial potency of AgNPs and leaf extract was evaluated against microbial strains of Staphylococcus aureus, Xanthomonas spp., Macrophomina phaseolina, and Fusarium oxysporum. AgNPs showed significant antioxidant efficacy with IC50 = 426.25 µg/mL in comparison to leaf extract with IC50 = 432.50 µg/mL against 2,2-diphenyl-1-picrylhydrazyl (DPPH). The AgNPs (64.36 mg AAE/g) demonstrated greater total antioxidant capacity using the phosphomolybdneum assay compared to the aqueous leaf extract (55.61 mg AAE/g) at a concentration of 1100 µg/mL. Based on these findings, AgNPs may indeed be useful for biomedical applications and drug delivery systems in the future.

Identification of perturbed pathways rendering susceptibility to tuberculosis in type 2 diabetes mellitus patients using BioNSi simulation of integrated networks of implicated human genes.

In type 2 diabetes mellitus (T2DM) patients, chronic hyperglycemia and inflammation underlie susceptibility to tuberculosis (TB) and result in poor TB control. Here, an integrative pathway-based approach is used to investigate perturbed pathways in T2DM patients that render susceptibility to TB. We obtained 36 genes implicated in type 2 diabetes-associated tuberculosis (T2DMTB) from the literature. Gene expression analysis on T2DM patient data (GSE26168) showed that DEFA1 is differentially expressed at Padj <0.05. The human host TB susceptibility genes TNFRSF10A, MSRA, GPR148, SLC37A3, PXK, PROK2, REV3L, PGM1, HIST3H2A, PLAC4, LETM2, and EMP2 and hsa-miR-146a microRNA were also differentially expressed at Padj <0.05. We included all these genes and added the remaining 28 genes from the T2DMTB set and the remaining differentially expressed genes at Padj <0.05 in STRING and obtained a well-connected network with high confidence score (≥0.7). Further, we extracted the KEGG pathways at FDR <0.05 and retained only the diabetes and TB pathways. The network was simulated with BioNSi using gene expression data. It is evident from BioNSi analysis that the NF-kappa B and Toll-like receptor pathways are commonly perturbed with high ranking in multiple gene expression datasets of type 2 diabetes versus healthy controls. The other pathways, necroptosis pathway and FoxO signalling pathway, appear perturbed with high ranking in different gene expression datasets. These pathways likely underlie susceptibility to TB in T2DM patients.

Salivary AsHPX12 influences pre-blood meal associated behavioral properties in Anopheles stephensi.

BACKGROUND & OBJECTIVES: A successful blood meal acquisition process by an adult female mosquito is accomplished through salivary glands, which releases a cocktail of proteins to counteract the vertebrate host's immune homeostasis. Here, we characterize a salivary-specific Heme peroxidase family member HPX12, originally identified from Plasmodium vivax infected salivary RNAseq data of the mosquito Anopheles stephensi. METHODS: To demonstrate we utilized a comprehensive in silico and functional genomics approach. RESULTS: Our dsRNA-mediated silencing experiments demonstrate that salivary AsHPX12 may regulate pre-blood meal-associated behavioral properties such as probing time, probing propensity, and host attraction. Altered expression of the salivary secretory and antennal proteins expression may have accounted for salivary homeostasis disruption resulting in the unusual fast release of salivary cocktail proteins and delayed acquisition of blood meal in the AsHPX12 knockdown mosquitoes. We also observed a significant parallel transcriptional modulation in response to blood feeding and P. vivax infection. INTERPRETATION & CONCLUSION: With this work, we establish a possible functional correlation of AsHPX12 role in the maintenance of salivary physiological-homeostasis, and Plasmodium sporozoites survival/transmission, though the mechanism is yet to unravel.

Antibiotics in anaerobic digestion: Investigative studies on digester performance and microbial diversity.

The ever-increasing consumption of antibiotics in both humans and animals has increased their load in municipal and pharmaceutical industry waste and may cause serious damage to the environment. Impact of antibiotics on the performance of commercially used anaerobic digesters in terms of bioenergy output, antibiotics' removal and COD removal have been compared critically with a few studies indicating >90% removal of antibiotics. AnMBR performed the best in terms of antibiotic removal, COD removal and methane yield. Most of the antibiotics investigated have adverse effects on microbiome associated with different stages and methane generation pathways of AD which has been assessed using high throughput technologies like metatranscriptomics, metaproteomics and flow cytometry. Perspectives have been given for understanding the fate and elimination of antibiotics from AD. The challenge of optimization and process improvement needs to be addressed to increase efficiency of the anaerobic digesters.

Electrochemical, Photophysical, Morphological and DFT Study of Polymorphic Sn(IV)-Porphyrins Containing Fluorinated Axial Ligand.

In this study, we report the polymorphism of six coordinated Sn(IV)- tetrabromophenyl porphyrins axially armed with fluorine-substituted phenolate ligands (structural formula [Sn(TBrPP)2+ (A- )2 ], where A is the axial ligand=3,5-difluoro phenol, compound 1). One form stabilizes in triclinic system (namely, 1α), and the other stabilizes in monoclinic system (namely, 1ß). The two 1α and 1ß polymorphs display distinct photophysical and morphological properties in the solid state. X-ray diffraction study reveals that these polymorphs 1α and 1ß significantly differ in their supramolecular architecture, different axial phenolate conformations, and noncovalent interactions, which are responsible for their distinct solid-state properties. The crystal packing of these polymorphs dominates by intermolecular C-H⋅⋅⋅F, C-H⋅⋅⋅π and C-Br⋅⋅⋅F interhalogen interactions. Furthermore, the solid-state emission spectra of 1α showed red-shifted emission bands with respect to 1ß, in addition the redox behavior of 1α is slightly different in comparison to 1ß. Complementary theoretical studies with Hirshfeld surface analysis show the definite role of Br⋅⋅⋅F interhalogen interactions in the overall stability. Mapping the electrostatic potential isosurfaces with the aid of density functional theory in compound 1 clearly shows the presence of σ-hole, a requisite feature to show halogen interactions in the crystalline state. In addition, lattice energy and single point energy calculation shows that 1α was found to be energetically more favorable and thermodynamically more stable compare to 1ß.

Bidirectional Microbiome-Gut-Brain-Axis Communication Influences Metabolic Switch-Associated Responses in the Mosquito Anopheles culicifacies.

The periodic ingestion of a protein-rich blood meal by adult female mosquitoes causes a drastic metabolic change in their innate physiological status, which is referred to as a 'metabolic switch'. While understanding the neural circuits for host-seeking is modestly attended, how the gut 'metabolic switch' modulates brain functions, and resilience to physiological homeostasis, remains unexplored. Here, through a comparative brain RNA-Seq study, we demonstrate that the protein-rich diet induces the expression of brain transcripts related to mitochondrial function and energy metabolism, possibly causing a shift in the brain's engagement to manage organismal homeostasis. A dynamic mRNA expression pattern of neuro-signaling and neuro-modulatory genes in both the gut and brain likely establishes an active gut-brain communication. The disruption of this communication through decapitation does not affect the modulation of the neuro-modulator receptor genes in the gut. In parallel, an unusual and paramount shift in the level of neurotransmitters (NTs), from the brain to the gut after blood feeding, further supports the idea of the gut's ability to serve as a 'second brain'. After blood-feeding, a moderate enrichment of the gut microbial population, and altered immunity in the gut of histamine receptor-silenced mosquitoes, provide initial evidence that the gut-microbiome plays a crucial role in gut-brain-axis communication. Finally, a comparative metagenomics evaluation of the gut microbiome highlighted that blood-feeding enriches the family members of the Morganellaceae and Pseudomonadaceae bacterial communities. The notable observation of a rapid proliferation of Pseudomonas bacterial sp. and tryptophan enrichment in the gut correlates with the suppression of appetite after blood-feeding. Additionally, altered NTs dynamics of naïve and aseptic mosquitoes provide further evidence that gut-endosymbionts are key modulators for the synthesis of major neuroactive molecules. Our data establish a new conceptual understanding of microbiome-gut-brain-axis communication in mosquitoes.

Functional disruption of transferrin expression alters reproductive physiology in Anopheles culicifacies.

BACKGROUND: Iron metabolism is crucial to maintain optimal physiological homeostasis of every organism and any alteration of the iron concentration (i.e. deficit or excess) can have adverse consequences. Transferrins are glycoproteins that play important role in iron transportation and have been widely characterized in vertebrates and insects, but poorly studied in blood-feeding mosquitoes. RESULTS: We characterized a 2102 bp long transcript AcTrf1a with complete CDS of 1872bp, and 226bp UTR region, encoding putative transferrin homolog protein from mosquito An. culicifacies. A detailed in silico analysis predicts AcTrf1a encodes 624 amino acid (aa) long polypeptide that carries transferrin domain. AcTrf1a also showed a putative N-linked glycosylation site, a characteristic feature of most of the mammalian transferrins and certain non-blood feeding insects. Structure modelling prediction confirms the presence of an iron-binding site at the N-terminal lobe of the transferrin. Our spatial and temporal expression analysis under altered pathophysiological conditions showed that AcTrf1a is abundantly expressed in the fat-body, ovary, and its response is significantly altered (enhanced) after blood meal uptake, and exogenous bacterial challenge. Additionally, non-heme iron supplementation of FeCl3 at 1 mM concentration not only augmented the AcTrf1a transcript expression in fat-body but also enhanced the reproductive fecundity of gravid adult female mosquitoes. RNAi-mediated knockdown of AcTrf1a causes a significant reduction in fecundity, confirming the important role of transferrin in oocyte maturation. CONCLUSION: All together our results advocate that detailed characterization of newly identified AcTrf1a transcript may help to select it as a unique target to impair the mosquito reproductive outcome.

A testis-expressing heme peroxidase HPX12 regulates male fertility in the mosquito Anopheles stephensi.

In vertebrates dysregulation of the antioxidant defense system has a detrimental impact on male fertility and reproductive physiology. However, in insects, especially mosquitoes the importance of sperm quality has been poorly studied. Since long-term storage of healthy and viable sperm earmarks male reproductive competency, we tested whether the heme peroxidase, a member of antioxidant enzyme family proteins, and abundantly expressed in the testis, also influence male fertility in the mosquito An. stephensi. Here, we show that a heme peroxidase 12 (HPX12), is an important cellular factor to protect the sperms from oxidative stress, and maintains semen quality in the male mosquito reproductive organ. We demonstrate that knockdown of the HPX12 not only impairs the sperm parameters such as motility, viability but also causes a significant down-regulation of MAG expressing transcripts such as ASTEI02706, ASTEI00744, ASTEI10266, likely encoding putative Accessory gland proteins. Mating with HPX12 knockdown male mosquitoes, resulted in ~ 50% reduction in egg-laying, coupled with diminished larval hatchability of a gravid female mosquito. Our data further outlines that increased ROS in the HPX12 mRNA depleted mosquitoes is the ultimate cause of sperm disabilities both qualitatively as well as quantitatively. Our data provide evidence that testis expressing AsHPX12 is crucial for maintaining optimal homeostasis for storing and protecting healthy sperms in the male mosquito's reproductive organs. Since, high reproductive capacity directly influences the mosquito population, manipulating male mosquito reproductive physiology could be an attractive tool to combat vector-borne diseases.

A lexical review on Vishaghna Dravyas of Kaideva Nighantu.

Introduction: Agad Tantra being the unique branch that deals with toxicology and its management enlists several antitoxic drugs used for various poisonings. Ancient texts comprise the list of a wide range of traditional medicines, but they are not explored due to lack of incomprehension. Aims: The review has been written with the aim to dig out the hidden knowledge of Vishaghna Dravyas (alexeterics) mentioned in Kaiydeva Nighantu. Materials and methods: This Nighantu was written in the 15th century A. D. by Kaiydeva with the name "Pathya-Apathya-Vibodhaka" it comprising of total 514 Dravyas out of which 175 drugs possess Vishagna (anti-poisonous) property. Results: These Vishaghna Dravyas (alexeterics) have been distributed based on various Vargas (classes) which is described as 121 belonging to Ausghada Varga (drug class) as single drugs and four as groups, total of 16 from Dhatu Varga (metal class) as single drugs and one as compound or as a group, 9 from Dhanya Varga (cereal class) as a single drug, 15 in Dravya Varga as single drugs, and 5 as compound or as a group. Two each in Kritana Varga and Vihara Varga and one in Mansa Varga as compound or as a group. Out of all the 175 Vishagna Dravayas (alexeterics), 18 Dravyas (substances) are specific indications in combating particular types of envenomation or poisoning conditions. Conclusion: All the abovementioned drugs are screened for the purpose of revalidation to bring out their therapeutic utility.

In silico prediction of natural compounds as potential multi-target inhibitors of structural proteins of SARS-CoV-2.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a colossal loss to human health and lives and has deeply impacted socio-economic growth. Remarkable efforts have been made by the scientific community in containing the virus by successful development of vaccines and diagnostic kits. Initiatives towards drug repurposing and discovery have also been undertaken. In this study, we compiled the known natural anti-viral compounds using text mining of the literature and examined them against four major structural proteins of SARS-CoV-2, namely, spike (S) protein, nucleocapsid (N) protein, membrane (M) protein and envelope (E) protein. Following computational approaches, we identified fangchinoline and versicolactone C as the compounds to exhibit strong binding to the target proteins and causing structural deformation of three structural proteins (N, S and M). We recommend the inhibitory effects of these compounds from our study should be experimentally validated against SARS-CoV-2.Communicated by Ramaswamy H. Sarma.

Exosomes: A Forthcoming Era of Breast Cancer Therapeutics.

Despite the recent advancements in therapeutics and personalized medicine, breast cancer remains one of the most lethal cancers among women. The prognostic and diagnostic aids mainly include assessment of tumor tissues with conventional methods towards better therapeutic strategies. However, current era of gene-based research may influence the treatment outcome particularly as an adjunct to diagnostics by exploring the role of non-invasive liquid biopsies or circulating markers. The characterization of tumor milieu for physiological fluids has been central to identifying the role of exosomes or small extracellular vesicles (sEVs). These exosomes provide necessary communication between tumor cells in the tumor microenvironment (TME). The manipulation of exosomes in TME may provide promising diagnostic/therapeutic strategies, particularly in triple-negative breast cancer patients. This review has described and highlighted the role of exosomes in breast carcinogenesis and how they could be used or targeted by recent immunotherapeutics to achieve promising intervention strategies.

Hemocyte RNA-Seq analysis of Indian malarial vectors Anopheles stephensi and Anopheles culicifacies: From similarities to differences.

Anopheles stephensi and Anopheles culicifacies are dominant malarial vectors in urban and rural India, respectively. Both species carry significant biological differences in their behavioral adaptation and immunity, but the genetic basis of these variations are still poorly understood. Here, we uncovered the genetic differences of immune blood cells, that influence several immune-physiological responses. We generated, analyzed and compared the hemocyte RNA-Seq database of both mosquitoes. A total of 5,837,223,769 assembled bases collapsed into 7,595 and 3,791 transcripts, originating from hemocytes of laboratory-reared 3-4 days old naïve (sugar-fed) mosquitoes, Anopheles stephensi and Anopheles culicifacies respectively. Comparative GO annotation analysis revealed that both mosquito hemocytes encode similar proteins. Furthermore, while An. stephensi hemocytes showed a higher percentage of immune transcripts encoding APHAG (Autophagy), IMD (Immune deficiency pathway), PRDX (Peroxiredoxin), SCR (Scavenger receptor), IAP (Inhibitor of apoptosis), GALE (galactoside binding lectins), BGBPs (1,3 beta D glucan binding proteins), CASPs (caspases) and SRRP (Small RNA regulatory pathway), An. culicifacies hemocytes yielded a relatively higher percentage of transcripts encoding CLIP (Clip domain serine protease), FREP (Fibrinogen related proteins), PPO (Prophenol oxidase), SRPN (Serpines), ML (Myeloid differentiation 2-related lipid recognition protein), Toll path and TEP (Thioester protein), family proteins. However, a detailed comparative Interproscan analysis showed An. stephensi mosquito hemocytes encode proteins with increased repeat numbers as compared to An. culicifacies. Notably, we observed an abundance of transcripts showing significant variability of encoded proteins with repeats such as LRR (Leucine rich repeat), WD40 (W-D dipeptide), Ankyrin, Annexin, Tetratricopeptide and Mitochondrial substrate carrier repeat-containing family proteins, which may have a direct influence on species-specific immune-physiological responses. Summarily, our deep sequencing analysis unraveled that An. stephensi evolved with an expansion of repeat sequences in hemocyte proteins as compared to An. culicifacies, possibly providing an advantage for better adaptation to diverse environments.

Genetic changes of Plasmodium vivax tempers host tissue-specific responses in Anopheles stephensi.

Recently, we showed how an early restriction of gut flora proliferation by Plasmodium vivax favors immune-suppression and Plasmodium survival in the gut lumen (Sharma et al., 2020). Here, we asked post gut invasion how P. vivax interacts with individual tissues such as the midgut, hemocyte, and salivary glands, and manages its survival in the mosquito host. Our data from tissue-specific comparative RNA-Seq analysis and extensive temporal/spatial expression profiling of selected mosquito transcripts in the uninfected and P. vivax infected mosquito's tissues indicated that (i) a transient suppression of gut metabolic machinery by early oocysts; (ii) enriched expression of nutritional responsive proteins and immune proteins against late oocysts, together may ensure optimal parasite development and gut homeostasis restoration; (iii) pre-immune activation of hemocyte by early gut-oocysts infection via REL induction (p â€‹< â€‹0.003); and altered expression of hemocyte-encoded immune proteins may cause rapid removal of free circulating sporozoites from hemolymph; (iv) while a strong suppression of salivary metabolic activities, and elevated expression of salivary specific secretory, as well as immune proteins together, may favor the long-term storage and survival of invaded sporozoites. Finally, our RNA-Seq-based discovery of 4449 transcripts of Plasmodium vivax origin, and their developmental stage-specific expression modulation in the corresponding infected mosquito tissues, predicts a possible mechanism of mosquito responses evasion by P. vivax. Conclusively, our system-wide RNA-Seq analysis provides the first genetic evidence of direct mosquito-Plasmodium interaction and establishes a functional correlation.