Prognostic Impact of Para-Aortic Lymph Node Metastasis in Resected Non-Pancreatic Periampullary Cancers.

BACKGROUND: Surgery remains debatable in para-aortic lymph node (PALN, station 16b1) metastasis in non-pancreatic periampullary cancer (NPPAC). This study examined the impact of PALN metastasis on outcomes following pancreaticoduodenectomy (PD) in NPPAC. METHODS: A retrospective analysis of patients with NPPAC who were explored for PD with PALN dissection was performed. Based on the extent of nodal involvement on final histopathology, they were stratified as node-negative (N0), regional node involved (N+) and metastatic PALN (N16+) and their outcomes were compared. RESULTS: Between 2011 and 2022, 153/887 PD patients underwent a PALN dissection, revealing N16+ in 42 patients (27.4%), of whom 32 patients underwent resection. The 3-years overall survival (OS) for patients with N16+ was 28% (95% confidence interval [CI] 13-60%), notably lower than the 67% (95% CI 53-83.5%; p = 0.007) for those without PALN metastasis. Stratified by nodal involvement, the median OS for N+ and N16+ patients was similar (28.4 months and 26.2 months, respectively). The N0 subgroup had a significantly longer 3-years OS of 87.5% (95% CI 79-96.7%; p = 0.0051). Interestingly, 10 patients not offered resection following N16+ identified on frozen section had a median survival of only 9 months. The perioperative morbidity and mortality in patients undergoing PD with PALN dissection were similar to standard resections. CONCLUSION: In a select group of patients with NPPAC, PD in isolated PALN metastasis was associated with improved OS. The survival in this group of patients was comparable with regional node-positive patients and significantly better than palliative treatment alone.

Intussusception after Rotavirus Vaccine Introduction in India.

BACKGROUND: A three-dose, oral rotavirus vaccine (Rotavac) was introduced in the universal immunization program in India in 2016. A prelicensure trial involving 6799 infants was not large enough to detect a small increased risk of intussusception. Postmarketing surveillance data would be useful in assessing whether the risk of intussusception would be similar to the risk seen with different rotavirus vaccines used in other countries. METHODS: We conducted a multicenter, hospital-based, active surveillance study at 27 hospitals in India. Infants meeting the Brighton level 1 criteria of radiologic or surgical confirmation of intussusception were enrolled, and rotavirus vaccination was ascertained by means of vaccination records. The relative incidence (incidence during the risk window vs. all other times) of intussusception among infants 28 to 365 days of age within risk windows of 1 to 7 days, 8 to 21 days, and 1 to 21 days after vaccination was evaluated by means of a self-controlled case-series analysis. For a subgroup of patients, a matched case-control analysis was performed, with matching for age, sex, and location. RESULTS: From April 2016 through June 2019, a total of 970 infants with intussusception were enrolled, and 589 infants who were 28 to 365 days of age were included in the self-controlled case-series analysis. The relative incidence of intussusception after the first dose was 0.83 (95% confidence interval [CI], 0.00 to 3.00) in the 1-to-7-day risk window and 0.35 (95% CI, 0.00 to 1.09) in the 8-to-21-day risk window. Similar results were observed after the second dose (relative incidence, 0.86 [95% CI, 0.20 to 2.15] and 1.23 [95% CI, 0.60 to 2.10] in the respective risk windows) and after the third dose (relative incidence, 1.65 [95% CI, 0.82 to 2.64] and 1.08 [95% CI, 0.69 to 1.73], respectively). No increase in intussusception risk was found in the case-control analysis. CONCLUSIONS: The rotavirus vaccine produced in India that we evaluated was not associated with intussusception in Indian infants. (Funded by the Bill and Melinda Gates Foundation and others.).

Deciphering insights into rhizospheric microbial community and soil parameters under the influence of herbicides in zero-tillage tropical rice-agroecosystem.

Extensive use of chemicals like herbicides in rice and other fields to manage weeds is expected to have a lasting influence on the soil environment. Considering this rationale, we aimed to decipher the effects of herbicides, Pendimethalin and Pretilachlor, applied at 0.5 and 0.6 kg ha-1, respectively on the rhizosphere microbial community and soil characteristics in the tropical rice field, managed under zero tillage cultivation. The quantity of herbicide residues declined gradually since application up to 60 days thereafter it reached the non-detectable level. Most of the soil variables viz., microbial biomass, soil enzymes etc., exhibited slight reduction in the treated soils compared to the control. A gradual decline was observed in Mineral-N, MBC, MBN and enzyme activities. Quantitative polymerase chain reaction results showed maximal microbial abundance of bacteria, fungi and archaea at mid-flowering stage of rice crop. The 16 rRNA and ITS region targeted amplicons high throughput sequencing microbial metagenomic approach revealed total of 94, 1353, and 510 species for archaea, bacteria and fungi, respectively. The metabarcoding of core microbiota revealed that the archaea comprised of Nitrososphaera, Nitrosocosmicus, and Methanosarcina. In the bacterial core microbiome, Neobacillus, Nitrospira, Thaurea, and Microvigra were found as the predominant taxa. Fusarium, Clonostachys, Nigrospora, Mortierella, Chaetomium, etc., were found in core fungal microbiome. Overall, the study exhibited that the recommended dose of herbicides found to be detrimental to the microbial dynamics, though a negative relation between residues and soil variables was observed that might alter the microbial diversity. The outcomes offer a comprehensive understanding of how herbicides affect the microbial community in zero tillage rice soil, thus has a critical imputation for eco-friendly and sustainable rice agriculture. Further, the long-term studies will be helpful in elucidating the role of identified microbial groups in sustaining the soil fertility and crop productivity.

De novo genome assembly and comparative annotation reveals metabolic versatility in cellulolytic bacteria from cropland and forest soils.

Cellulose, the most abundant polysaccharide in nature, is a rich source of renewable energy and sustains soil nutrients. Among the microorganisms known to degrade cellulose, bacteria are less studied compared to fungi. In the present work, we have investigated the culturable bacteria actively involved in cellulose degradation in forest and crop field soils. Based on clear zone formation and enzyme activity assay, we identified 7 bacterial strains positive for cellulose degradation. Of these, two most efficient strains (Bacillus cereus strains BHU1 and BHU2) were selected for whole genome sequencing, annotation, and information regarding GC content, number of genes, total subsystems, starch, and cellulose degradation pathways. Average nucleotide identity (ANI) showed more than 90% similarity between both the strains (BHU1 and BHU2) and with B. cereus ATCC 14579. Both the strains have genes and enzyme families like endoglucanase and ß-glucosidase as evident from whole genome sequence. Cellulase containing gene families (GH5, GH8, GH1), and many other carbohydrate-degrading enzymes, were present in both the bacterial strains. Taken together, the results suggest that the strains were efficient in cellulose degradation, and can be used for energy generation and production of value-added product.

Effect of roughage on rumen microbiota composition in the efficient feed converter and sturdy Indian Jaffrabadi buffalo (Bubalus bubalis).

BACKGROUND: The rumen microbiota functions as an effective system for conversion of dietary feed to microbial proteins and volatile fatty acids. In the present study, metagenomic approach was applied to elucidate the buffalo rumen microbiome of Jaffrabadi buffalo adapted to varied dietary treatments with the hypothesis that the microbial diversity and subsequent in the functional capacity will alter with diet change and enhance our knowledge of effect of microbe on host physiology. Eight adult animals were gradually adapted to an increasing roughage diet (4 animals each with green and dry roughage) containing 50:50 (J1), 75:25 (J2) and 100:0 (J3) roughage to concentrate proportion for 6 weeks. Metagenomic sequences of solid (fiber adherent microbiota) and liquid (fiber free microbiota) fractions obtained using Ion Torrent PGM platform were analyzed using MG-RAST server and CAZymes approach. RESULTS: Taxonomic analysis revealed that Bacteroidetes was the most abundant phylum followed by Firmicutes, Fibrobacter and Proteobacteria. Functional analysis revealed protein (25-30 %) and carbohydrate (15-20 %) metabolism as the dominant categories. Principal component analysis demonstrated that roughage proportion, fraction of rumen and type of forage affected rumen microbiome at taxonomic as well as functional level. Rumen metabolite study revealed that rumen fluid nitrogen content reduced in high roughage diet fed animals and pathway analysis showed reduction in the genes coding enzymes involved in methanogenesis pathway. CAZyme annotation revealed the abundance of genes encoding glycoside hydrolases (GH), with the GH3 family most abundant followed by GH2 and GH13 in all samples. CONCLUSIONS: Results reveals that high roughage diet feed improved microbial protein synthesis and reduces methane emission. CAZyme analysis indicated the importance of microbiome in feed component digestion for fulfilling energy requirements of the host. The findings help determine the role of rumen microbes in plant polysaccharide breakdown and in developing strategies to maximize productivity in ruminants.

Identification of putative SNPs in progressive retinal atrophy affected Canis lupus familiaris using exome sequencing.

Progressive retinal atrophy (PRA) is one of the major causes of retinal photoreceptor cell degeneration in canines. The inheritance pattern of PRA is autosomal recessive and genetically heterogeneous. Here, using targeted sequencing technology, we have performed exome sequencing of 10 PRA-affected (Spitz=7, Cocker Spaniel=1, Lhasa Aphso=1 and Spitz-Labrador cross breed=1) and 6 normal (Spitz=5, Cocker Spaniel=1) dogs. The high-throughput sequencing using 454-Roche Titanium sequencer generated about 2.16 Giga bases of raw data. Initially, we have successfully identified 25,619 single nucleotide polymorphisms (SNPs) that passed the stringent SNP calling parameters. Further, we performed association study on the cohort, and the highly significant (0.001) associations were short-listed and investigated in-depth. Out of the 171 significant SNPs, 113 were previously unreported. Interestingly, six among them were non-synonymous coding (NSC) SNPs, which includes CPPED1 A>G (p.M307V), PITRM1 T>G (p.S715A), APP G>A (p.T266M), RNF213 A>G (p.V1482A), C>A (p.V1456L), and SLC46A3 G>A (p.R168Q). On the other hand, 35 out of 113 unreported SNPs were falling in regulatory regions such as 3'-UTR, 5'-UTR, etc. In-depth bioinformatics analysis revealed that majority of NSC SNPs have damaging effect and alter protein stability. This study highlighted the genetic markers associated with PRA, which will help to develop genetic assay-based screening in effective breeding.

Microbial profiles of liquid and solid fraction associated biomaterial in buffalo rumen fed green and dry roughage diets by tagged 16S rRNA gene pyrosequencing.

The microbiome of buffalo rumen plays an important role in animal health and productivity. The rumen bacterial composition of both liquid and solid fraction was surveyed using pyrosequencing of the 16S rRNA gene. Sequences were analyzed using taxonomy-dependent clustering methods and revealed that the dominant ruminal bacteria shared by samples belonged to phyla Bacteroidetes, Firmicutes, Fibrobacteres and Proteobacteria. The core rumen microbiome of the rumen consisted of 10 phyla, 19 classes, 22 orders and 25 families. However, the relative abundance of these bacterial groups was markedly affected by diet composition as well as in type of biomaterial. In animals fed with a green and dry roughage diet, the cellulolytic bacteria, Ruminococcaceae, and Fibrobacteraceae was found in highest abundance in all biomaterials which reflected the need for enhanced fiber-digesting capacity in buffalo. The polysaccharide-degrading Prevotellaceae bacteria were most abundant in buffalo rumen. In taxonomic comparison of rumen bacteria, about 26 genera were differentially abundant among liquid and solid fraction of ruminal fluid. These results highlight the buffalo ruminal microbiome's ability to adapt to feed with different composition.

Insights into resistome and stress responses genes in Bubalus bubalis rumen through metagenomic analysis.

Buffalo rumen microbiota experience variety of diets and represents a huge reservoir of mobilome, resistome and stress responses. However, knowledge of metagenomic responses to such conditions is still rudimentary. We analyzed the metagenomes of buffalo rumen in the liquid and solid phase of the rumen biomaterial from river buffalo adapted to varying proportion of concentrate to green or dry roughages, using high-throughput sequencing to know the occurrence of antibiotics resistance genes, genetic exchange between bacterial population and environmental reservoirs. A total of 3914.94 MB data were generated from all three treatments group. The data were analysed with Metagenome rapid annotation system tools. At phyla level, Bacteroidetes were dominant in all the treatments followed by Firmicutes. Genes coding for functional responses to stress (oxidative stress and heat shock proteins) and resistome genes (resistance to antibiotics and toxic compounds, phages, transposable elements and pathogenicity islands) were prevalent in similar proportion in liquid and solid fraction of rumen metagenomes. The fluoroquinolone resistance, MDR efflux pumps and Methicillin resistance genes were broadly distributed across 11, 9, and 14 bacterial classes, respectively. Bacteria responsible for phages replication and prophages and phage packaging and rlt-like streptococcal phage genes were mostly assigned to phyla Bacteroides, Firmicutes and proteaobacteria. Also, more reads matching the sigma B genes were identified in the buffalo rumen. This study underscores the presence of diverse mechanisms of adaptation to different diet, antibiotics and other stresses in buffalo rumen, reflecting the proportional representation of major bacterial groups.

Characterization of the rumen microbiome of Indian Kankrej cattle (Bos indicus) adapted to different forage diet.

Present study described rumen microbiome of Indian cattle (Kankrej breed) to better understand the microbial diversity and largely unknown functional capacity of the rumen microbiome under different dietary treatments. Kankrej cattle were gradually adapted to a high-forage diet (four animals with dry forage and four with green forage) containing 50 % (K1), 75 % (K2) to 100 % (K3) forage, and remaining concentrate diet, each for 6 weeks followed by analysis of rumen fiber adherent and fiber-free metagenomic community by shotgun sequencing using ion torrent PGM platform and EBI-metagenomics annotation pipeline. Taxonomic analysis indicated that rumen microbiome was dominated by Bacteroidetes followed by Firmicutes, Fibrobacter, Proteobacteria, and Tenericutes. Functional analysis based on gene ontology classified all reads in total 157 categories based on their functional role in biological, molecular, and cellular component with abundance of genes associated with hydrolase activity, membrane, transport, transferase, and different metabolism (such as carbohydrate and protein). Statistical analysis using STAMP revealed significant differences (P < 0.05) between solid and liquid fraction of rumen (in 65 categories), between all three treatments (in 56 categories), and between green and dry roughage (17 categories). Diet treatment also exerted significant difference in environmental gene tags (EGTs) involved in metabolic pathways for production of volatile fatty acids. EGTs for butyrate production were abundant in K2, whereas EGTs for propionate production was abundant during K1. Principal component analysis also demonstrated that diet proportion, fraction of rumen, and type of forage affected rumen microbiome at taxonomic as well as functional level.

Deciphering the Genomic Landscape and Virulence Mechanisms of the Wheat Powdery Mildew Pathogen Blumeria graminis f. sp. tritici Wtn1: Insights from Integrated Genome Assembly and Conidial Transcriptomics.

A high-quality genome sequence from an Indian isolate of Blumeria graminis f. sp. tritici Wtn1, a persistent threat in wheat farming, was obtained using a hybrid method. The assembly of over 9.24 million DNA-sequence reads resulted in 93 contigs, totaling a 140.61 Mb genome size, potentially encoding 8480 genes. Notably, more than 73.80% of the genome, spanning approximately 102.14 Mb, comprises retro-elements, LTR elements, and P elements, influencing evolution and adaptation significantly. The phylogenomic analysis placed B. graminis f. sp. tritici Wtn1 in a distinct monocot-infecting clade. A total of 583 tRNA anticodon sequences were identified from the whole genome of the native virulent strain B. graminis f. sp. tritici, which comprises distinct genome features with high counts of tRNA anticodons for leucine (70), cysteine (61), alanine (58), and arginine (45), with only two stop codons (Opal and Ochre) present and the absence of the Amber stop codon. Comparative InterProScan analysis unveiled "shared and unique" proteins in B. graminis f. sp. tritici Wtn1. Identified were 7707 protein-encoding genes, annotated to different categories such as 805 effectors, 156 CAZymes, 6102 orthologous proteins, and 3180 distinct protein families (PFAMs). Among the effectors, genes like Avra10, Avrk1, Bcg-7, BEC1005, CSEP0105, CSEP0162, BEC1016, BEC1040, and HopI1 closely linked to pathogenesis and virulence were recognized. Transcriptome analysis highlighted abundant proteins associated with RNA processing and modification, post-translational modification, protein turnover, chaperones, and signal transduction. Examining the Environmental Information Processing Pathways in B. graminis f. sp. tritici Wtn1 revealed 393 genes across 33 signal transduction pathways. The key pathways included yeast MAPK signaling (53 genes), mTOR signaling (38 genes), PI3K-Akt signaling (23 genes), and AMPK signaling (21 genes). Additionally, pathways like FoxO, Phosphatidylinositol, the two-component system, and Ras signaling showed significant gene representation, each with 15-16 genes, key SNPs, and Indels in specific chromosomes highlighting their relevance to environmental responses and pathotype evolution. The SNP and InDel analysis resulted in about 3.56 million variants, including 3.45 million SNPs, 5050 insertions, and 5651 deletions within the whole genome of B. graminis f. sp. tritici Wtn1. These comprehensive genome and transcriptome datasets serve as crucial resources for understanding the pathogenicity, virulence effectors, retro-elements, and evolutionary origins of B. graminis f. sp. tritici Wtn1, aiding in developing robust strategies for the effective management of wheat powdery mildew.

Organic Farming Favors phoD-Harboring Rhizospheric Bacterial Community and Alkaline Phosphatase Activity in Tropical Agroecosystem.

The bacteria harboring phoD encodes alkaline phosphatase (ALP), a secretory enzyme that hydrolyzes organic phosphorous (P) to a usable form in the soil. The impact of farming practices and crop types on phoD bacterial abundance and diversity in tropical agroecosystems is largely unknown. In this research, the aim was to study the effect of farming practices (organic vs. conventional) and crop types on the phoD-harboring bacterial community. A high-throughput amplicon (phoD gene) sequencing method was employed for the assessment of bacterial diversity and qPCR for phoD gene abundance. Outcomes revealed that soils treated for organic farming have high observed OTUs, ALP activity, and phoD population than soils managed under conventional farming with the trend of maize > chickpea > mustard > soybean vegetated soils. The relative abundance of Rhizobiales exhibited dominance. Ensifer, Bradyrhizobium, Streptomyces, and Pseudomonas were observed as dominant genera in both farming practices. Overall, the study demonstrated that organic farming practice favors the ALP activity, phoD abundance, and OTU richness which varied across crop types with maize crops showing the highest OTUs followed by chickpea, mustard, and least in soybean cropping.

Unraveling the transcriptional network regulated by miRNAs in blast-resistant and blast-susceptible rice genotypes during Magnaporthe oryzae interaction.

The plant pathogen Magnaporthe oryzae poses a significant threat to global food security, and its management through the cultivation of resistant varieties and crop husbandry practices, including fungicidal sprays, has proven to be inadequate. To address this issue, we conducted small-RNA sequencing to identify the roles of miRNAs and their target genes in both resistant (PB1637) and susceptible (PB1) rice genotypes. We confirmed the expression of differentially expressed miRNAs using stem-loop qRT-PCR analysis and correlated them with rice patho-phenotypic and physio-biochemical responses. Our findings revealed several noteworthy differences between the resistant and susceptible genotypes. The resistant genotype exhibited reduced levels of total chlorophyll and carotenoids compared to the susceptible genotype. However, it showed increased levels of total protein, callose, H2O2, antioxidants, flavonoids, and total polyphenols. Additionally, among the defense-associated enzymes, guaiacol peroxidase and polyphenol oxidase responses were higher in the susceptible genotypes. In our comparative analysis, we identified 27 up-regulated and 43 down-regulated miRNAs in the resistant genotype, while the susceptible genotype exhibited 44 up-regulated and 62 down-regulated miRNAs. Furthermore, we discovered eight up-regulated and five down-regulated miRNAs shared between the resistant and susceptible genotypes. Notably, we also identified six novel miRNAs in the resistant genotype and eight novel miRNAs in the susceptible genotype. These novel miRNAs, namely Chr8_26996, Chr12_40110, and Chr12_41899, were found to negatively correlate with the expression of predicted target genes, including Cyt-P450 monooxygenase, serine carboxypeptidase, and zinc finger A20 domain-containing stress-associated protein, respectively. The results of our study on miRNA and transcriptional responses provide valuable insights for the development of future rice lines that are resistant to blast disease. By understanding the roles of specific miRNAs and their target genes in conferring resistance, we can enhance breeding strategies and improve crop management practices to ensure global food security.

Fine-scale mapping of the microbiome on phylloplane and spermoplane of aromatic and non-aromatic rice genotypes.

Adaxial, abaxial phylloplane (leaf), and spermoplane (seed) are proximal yet contrasting habitats for a microbiota that needs to be adequately explored. Here, we proposed novel methods to decipher the adaxial/abaxial-phylloplane and spermoplane-microbiomes. Comparison of 22 meta barcoded-NGS datasets (size of total data set-1980.48 Mb) enabled us to fine-map the microbiome of the rice foliar niche, which encompasses the lower, middle, top leaf as well panicle. Here, the total- and the cultivable-microbiome profiling revealed 157 genera representing ten phyla and 87 genera from 4 bacterial phyla, respectively, with a predominance of Proteobacteria and Actinobacteria. Interestingly, more bacterial communities (124-genera) preferred the abaxial than the adaxial phylloplane (104-genera) and spermoplane (67-genera) for colonization. The microbiome profiles were nearly identical on the aromatic (125-genera) and non-aromatic rice (116-genera) with high representation of Pantoea, Methylobacterium, Curtobacterium, Sphingopyxis, and Microbacterium. The culturomics investigation confirmed the abundance of Pantoea, Chryseobacterium, Pseudomonas, Acinetobacter, Sphingobacterium, and Exiguobacterium. One hundred bacterial isolates characterized and identified by polyphasic-taxonomic tools revealed the dominance of Acinetobacter, Chryseobacterium, Enterobacter, Massilia, Pantoea, Pseudomonas, and Stenotrophomonas on adaxial/abaxial-phylloplane and spermoplane. The study culminated in identifying hitherto unexplored bacterial communities on the adaxial/abaxial phylloplane and spermoplane of rice that can be harnessed for microbiome-assisted rice cultivation in the future.

Structured Framework and Genome Analysis of Magnaporthe grisea Inciting Pearl Millet Blast Disease Reveals Versatile Metabolic Pathways, Protein Families, and Virulence Factors.

Magnaporthe grisea (T.T. Herbert) M.E. Barr is a major fungal phytopathogen that causes blast disease in cereals, resulting in economic losses worldwide. An in-depth understanding of the basis of virulence and ecological adaptation of M. grisea is vital for devising effective disease management strategies. Here, we aimed to determine the genomic basis of the pathogenicity and underlying biochemical pathways in Magnaporthe using the genome sequence of a pearl millet-infecting M. grisea PMg_Dl generated by dual NGS techniques, Illumina NextSeq 500 and PacBio RS II. The short and long nucleotide reads could be draft assembled in 341 contigs and showed a genome size of 47.89 Mb with the N50 value of 765.4 Kb. Magnaporthe grisea PMg_Dl showed an average nucleotide identity (ANI) of 86% and 98% with M. oryzae and Pyricularia pennisetigena, respectively. The gene-calling method revealed a total of 10,218 genes and 10,184 protein-coding sequences in the genome of PMg_Dl. InterProScan of predicted protein showed a distinct 3637 protein families and 695 superfamilies in the PMg_Dl genome. In silico virulence analysis revealed the presence of 51VFs and 539 CAZymes in the genome. The genomic regions for the biosynthesis of cellulolytic endo-glucanase and beta-glucosidase, as well as pectinolytic endo-polygalacturonase, pectin-esterase, and pectate-lyases (pectinolytic) were detected. Signaling pathways modulated by MAPK, PI3K-Akt, AMPK, and mTOR were also deciphered. Multicopy sequences suggestive of transposable elements such as Type LTR, LTR/Copia, LTR/Gypsy, DNA/TcMar-Fot1, and Type LINE were recorded. The genomic resource presented here will be of use in the development of molecular marker and diagnosis, population genetics, disease management, and molecular taxonomy, and also provide a genomic reference for ascomycetous genome investigations in the future.

Deciphering core phyllomicrobiome assemblage on rice genotypes grown in contrasting agroclimatic zones: implications for phyllomicrobiome engineering against blast disease.

BACKGROUND: With its adapted microbial diversity, the phyllosphere contributes microbial metagenome to the plant holobiont and modulates a host of ecological functions. Phyllosphere microbiome (hereafter termed phyllomicrobiome) structure and the consequent ecological functions are vulnerable to a host of biotic (Genotypes) and abiotic factors (Environment) which is further compounded by agronomic transactions. However, the ecological forces driving the phyllomicrobiome assemblage and functions are among the most understudied aspects of plant biology. Despite the reports on the occurrence of diverse prokaryotic phyla such as Proteobacteria, Firmicutes, Bacteroides, and Actinobacteria in phyllosphere habitat, the functional characterization leading to their utilization for agricultural sustainability is not yet explored. Currently, the metabarcoding by Next-Generation-Sequencing (mNGS) technique is a widely practised strategy for microbiome investigations. However, the validation of mNGS annotations by culturomics methods is not integrated with the microbiome exploration program. In the present study, we combined the mNGS with culturomics to decipher the core functional phyllomicrobiome of rice genotypes varying for blast disease resistance planted in two agroclimatic zones in India. There is a growing consensus among the various stakeholder of rice farming for an ecofriendly method of disease management. Here, we proposed phyllomicrobiome assisted rice blast management as a novel strategy for rice farming in the future. RESULTS: The tropical "Island Zone" displayed marginally more bacterial diversity than that of the temperate 'Mountain Zone' on the phyllosphere. Principal coordinate analysis indicated converging phyllomicrobiome profiles on rice genotypes sharing the same agroclimatic zone. Interestingly, the rice genotype grown in the contrasting zones displayed divergent phyllomicrobiomes suggestive of the role of environment on phyllomicrobiome assembly. The predominance of phyla such as Proteobacteria, Actinobacteria, and Firmicutes was observed in the phyllosphere irrespective of the genotypes and climatic zones. The core-microbiome analysis revealed an association of Acidovorax, Arthrobacter, Bacillus, Clavibacter, Clostridium, Cronobacter, Curtobacterium, Deinococcus, Erwinia, Exiguobacterium, Hymenobacter, Kineococcus, Klebsiella, Methylobacterium, Methylocella, Microbacterium, Nocardioides, Pantoea, Pedobacter, Pseudomonas, Salmonella, Serratia, Sphingomonas and Streptomyces on phyllosphere. The linear discriminant analysis (LDA) effect size (LEfSe) method revealed distinct bacterial genera in blast-resistant and susceptible genotypes, as well as mountain and island climate zones. SparCC based network analysis of phyllomicrobiome showed complex intra-microbial cooperative or competitive interactions on the rice genotypes. The culturomic validation of mNGS data confirmed the occurrence of Acinetobacter, Aureimonas, Curtobacterium, Enterobacter, Exiguobacterium, Microbacterium, Pantoea, Pseudomonas, and Sphingomonas in the phyllosphere. Strikingly, the contrasting agroclimatic zones showed genetically identical bacterial isolates suggestive of vertical microbiome transmission. The core-phyllobacterial communities showed secreted and volatile compound mediated antifungal activity on M. oryzae. Upon phyllobacterization (a term coined for spraying bacterial cells on the phyllosphere), Acinetobacter, Aureimonas, Pantoea, and Pseudomonas conferred immunocompetence against blast disease. Transcriptional analysis revealed activation of defense genes such as OsPR1.1, OsNPR1, OsPDF2.2, OsFMO, OsPAD4, OsCEBiP, and OsCERK1 in phyllobacterized rice seedlings. CONCLUSIONS: PCoA indicated the key role of agro-climatic zones to drive phyllomicrobiome assembly on the rice genotypes. The mNGS and culturomic methods showed Acinetobacter, Aureimonas, Curtobacterium, Enterobacter, Exiguobacterium, Microbacterium, Pantoea, Pseudomonas, and Sphingomonas as core phyllomicrobiome of rice. Genetically identical Pantoea intercepted on the phyllosphere from the well-separated agroclimatic zones is suggestive of vertical transmission of phyllomicrobiome. The phyllobacterization showed potential for blast disease suppression by direct antibiosis and defense elicitation. Identification of functional core-bacterial communities on the phyllosphere and their co-occurrence dynamics presents an opportunity to devise novel strategies for rice blast management through phyllomicrobiome reengineering in the future.

The costs of HIV prevention for different target populations in Mumbai, Thane and Bangalore.

BACKGROUND: Avahan, the India AIDS Initiative, delivers HIV prevention services to high-risk populations at scale. Although the broad costs of such HIV interventions are known, to-date there has been little data available on the comparative costs of reaching different target groups, including female sex workers (FSWs), replace with 'high risk men who have sex with men (HR-MSM) and trans-genders. METHODS: Costs are estimated for the first three years of Avahan scale up differentiated by typology of female sex workers (brothel, street, home, lodge based, bar based), HR-MSM and transgenders in urban districts in India: Mumbai and Thane in Maharashtra and Bangalore in Karnataka. Financial and economic costs were collected prospectively from a provider perspective. Outputs were measured using data collected by the Avahan programme. Costs are presented in US$2008. RESULTS: Costs were found to vary substantially by target group. Non-governmental organisations (NGOs) working with transgender populations had a higher mean cost (US $116) per person reached compared to those dealing primarily with FSWs (US $75-96) and MSWs (US $90) by the end of year three of the programme in Mumbai. The mean cost of delivering the intervention to HR-MSMs (US $42) was higher than delivering it to FSWs (US $37) in Bangalore. The package of services delivered to each target group was similar, and our results suggest that cost variation is related to the target population size, the intensity of the programme (in terms of number of contacts made per year) and a number of specific issues related to each target group. CONCLUSIONS: Based on our data policy makers and program managers need to consider the ease of accessing high risk population when planning and budgeting for HIV prevention services for these populations and avoid funding programmes on the basis of target population size alone.

Exploring the allochthonous pollution influence on bacterial community and co-occurrence dynamics of River Ganga water through 16S rRNA-tagged amplicon metagenome.

River Ganga is one of the largest and most sacred rivers of India. This river is largely affected by anthropogenic activities causing significant increase in water pollution. The impact of drains discharging polluted water on the bacterial community dynamics in the river remains unexplored. To elucidate this, the targeted 16S rRNA V3-V4 variable region amplicon sequencing and bioinformatic analysis were performed using water from upstream, drain, and downstream of river Ganga. Analysis revealed significant difference in relative abundances of bacterial communities. The increase in bacterial abundance and alpha diversity was detected in the downstream compared to the upstream. Environmental factors were found significantly different between upstream and downstream water. At the phyla level, highly abundant taxa such as Proteobacteria, Actinobacteria, Planctomycetes, Bacteroidetes, and Verrucomicrobia were observed. Bacterial genera like Prevotella, Bacteroides, Blautia, and Faecalibacterium (fecal indicator) had higher abundance in the downstream site. Network co-occurrence revealed that bacterial communities have a modular profile with reduced interaction in drain and downstream water. The network of co-occurring bacterial communities consists of 283 nodes with edge connectivity of 6900, 7074, and 5294 in upstream, drain, and downstream samples, respectively. Upstream communities exhibited the highest positive interaction followed by the drain and the downstream sites. Additionally, highly abundant pathogenic species such as Acinetobacter baumannii and Prevotella copri were also detected in all samples. This study suggests the drain to be allochthonous pollution vector that significantly contributes to bacterial community enrichment. From the results of this study, it is apparent that the lotic water may be used as the ecological reference to understand and monitor the variations in the bacterial communities and their co-occurrence dynamics in the fresh water ecosystems.

Hybrid de novo genome-reassembly reveals new insights on pathways and pathogenicity determinants in rice blast pathogen Magnaporthe oryzae RMg_Dl.

Blast disease incited by Magnaporthe oryzae is a major threat to sustain rice production in all rice growing nations. The pathogen is widely distributed in all rice paddies and displays rapid aerial transmissions, and seed-borne latent infection. In order to understand the genetic variability, host specificity, and molecular basis of the pathogenicity-associated traits, the whole genome of rice infecting Magnaporthe oryzae (Strain RMg_Dl) was sequenced using the Illumina and PacBio (RSII compatible) platforms. The high-throughput hybrid assembly of short and long reads resulted in a total of 375 scaffolds with a genome size of 42.43 Mb. Furthermore, comparative genome analysis revealed 99% average nucleotide identity (ANI) with other oryzae genomes and 83% against M. grisea, and 73% against M. poe genomes. The gene calling identified 10,553 genes with 10,539 protein-coding sequences. Among the detected transposable elements, the LTR/Gypsy and Type LINE showed high occurrence. The InterProScan of predicted protein sequences revealed that 97% protein family (PFAM), 98% superfamily, and 95% CDD were shared among RMg_Dl and reference 70-15 genome, respectively. Additionally, 550 CAZymes with high GH family content/distribution and cell wall degrading enzymes (CWDE) such endoglucanase, beta-glucosidase, and pectate lyase were also deciphered in RMg_Dl. The prevalence of virulence factors determination revealed that 51 different VFs were found in the genome. The biochemical pathway such as starch and sucrose metabolism, mTOR signaling, cAMP signaling, MAPK signaling pathways related genes were identified in the genome. The 49,065 SNPs, 3267 insertions and 3611 deletions were detected, and majority of these varinats were located on downstream and upstream region. Taken together, the generated information will be useful to develop a specific marker for diagnosis, pathogen surveillance and tracking, molecular taxonomy, and species delineation which ultimately leads to device improved management strategies for blast disease.

Potential Environmental and Human Health Risks Caused by Antibiotic-Resistant Bacteria (ARB), Antibiotic Resistance Genes (ARGs) and Emerging Contaminants (ECs) from Municipal Solid Waste (MSW) Landfill.

The disposal of municipal solid waste (MSW) directly at landfills or open dump areas, without segregation and treatment, is a significant concern due to its hazardous contents of antibiotic-resistant bacteria (ARB), antibiotic resistance genes (ARGs), and metal resistance genes (MGEs). The released leachate from landfills greatly effects the soil physicochemical, biological, and groundwater properties associated with agricultural activity and human health. The abundance of ARB, ARGs, and MGEs have been reported worldwide, including MSW landfill sites, animal husbandry, wastewater, groundwater, soil, and aerosol. This review elucidates the occurrence and abundance of ARB, ARGs, and MRGs, which are regarded as emerging contaminants (ECs). Recently, ECs have received global attention because of their prevalence in leachate as a substantial threat to environmental and public health, including an economic burden for developing nations. The present review exclusively discusses the demands to develop a novel eco-friendly management strategy to combat these global issues. This review also gives an intrinsic discussion about the insights of different aspects of environmental and public health concerns caused due to massive leachate generation, the abundance of antibiotics resistance (AR), and the effects of released leachate on the various environmental reservoirs and human health. Furthermore, the current review throws light on the source and fate of different ECs of landfill leachate and their possible impact on the nearby environments (groundwater, surface water, and soil) affecting human health. The present review strongly suggests the demand for future research focuses on the advancement of the removal efficiency of contaminants with the improvement of relevant landfill management to reduce the potential effects of disposable waste. We propose the necessity of the identification and monitoring of potential environmental and human health risks associated with landfill leachate contaminants.

Integrated Metabarcoding and Culturomic-Based Microbiome Profiling of Rice Phyllosphere Reveal Diverse and Functional Bacterial Communities for Blast Disease Suppression.

Phyllosphere-the harsh foliar plant part exposed to vagaries of environmental and climatic variables is a unique habitat for microbial communities. In the present work, we profiled the phyllosphere microbiome of the rice plants using 16S rRNA gene amplicon sequencing (hereafter termed metabarcoding) and the conventional microbiological methods (culturomics) to decipher the microbiome assemblage, composition, and their functions such as antibiosis and defense induction against rice blast disease. The blast susceptible rice genotype (PRR78) harbored far more diverse bacterial species (294 species) than the resistant genotype (Pusa1602) that showed 193 species. Our metabarcoding of bacterial communities in phyllomicrobiome revealed the predominance of the phylum, Proteobacteria, and its members Pantoea, Enterobacter, Pseudomonas, and Erwinia on the phyllosphere of both rice genotypes. The microbiological culturomic validation of metabarcoding-taxonomic annotation further confirmed the prevalence of 31 bacterial isolates representing 11 genera and 16 species with the maximum abundance of Pantoea. The phyllomicrobiome-associated bacterial members displayed antifungal activity on rice blast fungus, Magnaporthe oryzae, by volatile and non-volatile metabolites. Upon phyllobacterization of rice cultivar PB1, the bacterial species such as Enterobacter sacchari, Microbacterium testaceum, Pantoea ananatis, Pantoea dispersa, Pantoea vagans, Pseudomonas oryzihabitans, Rhizobium sp., and Sphingomonas sp. elicited a defense response and contributed to the suppression of blast disease. qRT-PCR-based gene expression analysis indicated over expression of defense-associated genes such as OsCEBiP, OsCERK1, and phytohormone-associated genes such as OsPAD4, OsEDS1, OsPR1.1, OsNPR1, OsPDF2.2, and OsFMO in phyllobacterized rice seedlings. The phyllosphere bacterial species showing blast suppressive activity on rice were found non-plant pathogenic in tobacco infiltration assay. Our comparative microbiome interrogation of the rice phyllosphere culminated in the isolation and identification of agriculturally significant bacterial communities for blast disease management in rice farming through phyllomicrobiome engineering in the future.