PNEUMOCOCCAL BIOFILMS AND THEIR INTERVENTION STRATEGIES

Pneumonia is a fatal infection with hard time breathing, cough, and fever. The children are at high risk worldwide due to pneumonia. This is responsible for childhood mortality and morbidity worldwide. It is mainly caused by bacteria. Pneumonia-causing bacteria are resistant to most of the antibiotics and therapeutic agents due to the formation of biofilms. Laboratories around the world are trying to develop strategies to combat pneumococcal biofilms. This review deals with the formation of pneumococcal biofilms and their different intervention strategies.Â

An overview of traditional rice beer of North-east India: ethnic preparation, challenges and prospects

744-757Fermented drinks are always found to be connected with the culture and existence of the ethnic tribes throughout the world. Traditional rice beer of North-east India is an indispensible part of tribal life style attached culturally and religiously with them. There are more than 150 major tribes in Northeast India and almost all are having a traditional way of preparing rice beer from time immemorial. The present paper is a comparative review on the methods of preparing rice beer by some popular tribes of Northeastern India. The methodology they use is found to be unique in many senses from rest of the world. The uniqueness not only lies in the starter culture preparation, which is the prime source of yeast but also in the other ingredients like indigenous plant species and rice varieties used as substrate resulting variation in taste and aroma. Almost all the traditional rice beer has unique soothing taste, aroma, colour and nutritional as well as therapeutic values having significant role in their socio cultural lives. In the present scenario the tradition is fading and facing lots of challenges. But, still the traditional rice beer has got lots of scope for its scientific up-gradation and commercialization

In silico studies reveal antiviral effects of traditional Indian spices on COVID-19

BACKGROUND: The global health emergency due to SARS-CoV-2 causing the COVID-19 pandemic emphasized the scientific community to intensify their research work for its therapeutic solution. In this study, Indian traditional spices owing to various medicinal properties were tested in silico for their inhibitory activity against SARS-CoV-2 proteins. SARS-CoV-2 spike proteins (SP) and main proteases (Mpro) play a significant role in infection development were considered as potential drug targets. METHODS: A total of 75 phytochemicals present in traditional Indian spices retrieved from the published literature and Dr. Duke\u27s Phytochemical and Ethnobotanical Database, were docked with Mpro (PDB IDs: 6YNQ), and the SP (PDB IDs: 6LXT and 6YOR). RESULTS: Through the screening process, 75 retrieved phytochemicals were docked with spike protein (PDB IDs: 6LXT and 6YOR) and main protease (PDB ID: 6YNQ) of SARS-CoV-2. Among them, myricetin, a flavonoid (rank score: 6LXT: -11.72383; 6YOR: -9.87943; 6YNQ: -11.68164) from Allium sativumL and Isovitexin, an example of flavone (rank score:6LXT: -12.14922; 6YOR: -10.19443; 6YNQ: - 12.60603) from Pimpinella anisumL were the most potent ligands against SP and Mproof SARS-CoV-2. Whereas, Astragalin from Crocus sativusL.; Rutin from Illicium verum, Oxyguttiferone from Garcinia cambogia; Scopolin from Apium graveolens L, Luteolin from Salvia officinalis, Emodin, Aloe-emodin from Cinnamomum zeylanicium and Apigenin from Allium sativumL showed better inhibition against Mpro than SP of SARS-CoV-2. The amino acid residues like SER, LYS, ASP and TYR were found playing important role in protein-ligand interactions via hydrogen bonding and Vander Waals forces. CONCLUSION: Optimal use of traditional spices in our daily meals may help fight against COVID-19. This study also paves the path for herbal drug formulation against SARS-CoV-2 after wet lab validation

Silybin B and Cianidanol Inhibit M pro and Spike Protein of SARS-CoV-2: Evidence from in Silico Molecular Docking Studies

BACKGROUND: The main proteases (Mpro) and Spike Proteins (SP) of Severe Acute Respiratory Syndrome Coronavirus 2 (SARSCoV-2) play a major role in viral infection development by producing several non-structural proteins (nsPs) and penetrating the host cells respectively. In this study, the potential of in silico molecular docking-based drug repositioning approach was exploited for identifying the inhibitors of Mpro and SP of SARS-CoV-2. METHODS: A total of 196 compounds including various US-FDA-approved drugs, vitamins and their analogs were docked with Mpro (PDB IDs: 6YB7 and 6Y84), and the top six ligands were further tested for ADME properties followed by docking with SP (PDB IDs: 6LXT and 6W41). RESULTS: Out of 196 compounds, binding energy (DE) of Silybin B (6YB7: DE: -11.20 kcal/mol; 6Y84: DE: -10.18 kcal/mol; 6LXT:DE: -10.47 kcal/mol; 6W41:DE: -10.96 kcal/mol) and Cianidanol (6YB7:DE: -8.85 kcal/mol; 6Y84:DE:-10.02 kcal/mol; 6LXT:DE:-9.36 kcal/mol; 6W41:DE: -9.52 kcal/mol) demonstrated better binding and ADME properties compared with the currently endeavored drugs like Hydroxychloroquine and Lopinavir. Additionally, Elliptinone, Diospyirin, SCHEMBL94263 and Fiboflavin have shown encouraging results. Fiboflavin, an immunity booster, was found to inhibit both the Mpro and spike protein of SARS-CoV-2. It was observed that amino acid residues MET6, ALA7, PHE8, PRO9, ASP295, GLY302, VAL303 and THR304 play significant roles in protein-ligand interactions through hydrogen bonds and Vander Waals forces. CONCLUSION: Silybin B and Cianidanol showed excellent binding and ADME properties compared with the currently endeavored drugs and can be exploited as therapeutic options against SARS-CoV-2 infection after experimental validation and clinical trials

Identification of a Protein Mediating Respiratory Supercomplex Stability

SummaryThe complexes of the electron transport chain associate into large macromolecular assemblies, which are believed to facilitate efficient electron flow. We have identified a conserved mitochondrial protein, named respiratory supercomplex factor 1 (Rcf1—Yml030w), that is required for the normal assembly of respiratory supercomplexes. We demonstrate that Rcf1 stably and independently associates with both Complex III and Complex IV of the electron transport chain. Deletion of the RCF1 gene caused impaired respiration, probably as a result of destabilization of respiratory supercomplexes. Consistent with the hypothetical function of these respiratory assemblies, loss of RCF1 caused elevated mitochondrial oxidative stress and damage. Finally, we show that knockdown of HIG2A, a mammalian homolog of RCF1, causes impaired supercomplex formation. We suggest that Rcf1 is a member of an evolutionarily conserved protein family that acts to promote respiratory supercomplex assembly and activity

Preparation and evaluation of the ZnO NP-Ampicillin/Sulbactam nanoantibiotic: Optimization of formulation variables using RSM coupled GA method and antibacterial activities

Nanoparticles (NPs) possessing antibacterial activity represent an effective way of overcoming bacterial resistance. In the present work, we report a novel formulation of a nanoantibiotic formed using Ampicillin/sulbactam (Ams) and a zinc oxide nanoparticle (ZnO NP). ‘ZnO NP–Ams’ nanoantibiotic formulation is optimized using response surface methodology coupled genetic algorithm approach. The optimized formulation of nanoantibiotic (ZnO NP: 49.9 μg/mL; Ams: 33.6 μg/mL; incubation time: 27 h) demonstrated 15% enhanced activity compared to the unoptimized formulation against K. pneumoniae. The reactive oxygen species (ROS) generation was directly proportional to the interaction time of nanoantibiotic and K. pneumoniae after the initial lag phase of ~18 h as evident from 2'-7'-Dichlorodihydrofluorescein diacetate assay. A low minimum inhibitory concentration (6.25 μg/mL) of nanoantibiotic formulation reveals that even a low concentration of nanoantibiotic can prove to be effective against K. pneumoniae. The importance of nanoantibiotic formulation is also evident by the fact that the 100 μg/mL of Ams and 25 μg of ZnO NP was required individually to inhibit the growth of K. pneumonia, whereas only 6.25 μg/mL of optimized nanoantibiotic formulation (ZnO NP and Ams in the ratio of 49.9: 33.6 in μg/mL and conjugation time of 27 h) was needed for the same

BONCAT-FACS-Seq reveals the active fraction of a biocrust community undergoing a wet-up event

Determining which microorganisms are active within soil communities remains a major technical endeavor in microbial ecology research. One promising method to accomplish this is coupling bioorthogonal non-canonical amino acid tagging (BONCAT) with fluorescence activated cell sorting (FACS) which sorts cells based on whether or not they are producing new proteins. Combined with shotgun metagenomic sequencing (Seq), we apply this method to profile the diversity and potential functional capabilities of both active and inactive microorganisms in a biocrust community after being resuscitated by a simulated rain event. We find that BONCAT-FACS-Seq is capable of discerning the pools of active and inactive microorganisms, especially within hours of applying the BONCAT probe. The active and inactive components of the biocrust community differed in species richness and composition at both 4 and 21 h after the wetting event. The active fraction of the biocrust community is marked by taxa commonly observed in other biocrust communities, many of which play important roles in species interactions and nutrient transformations. Among these, 11 families within the Firmicutes are enriched in the active fraction, supporting previous reports indicating that the Firmicutes are key early responders to biocrust wetting. We highlight the apparent inactivity of many Actinobacteria and Proteobacteria through 21 h after wetting, and note that members of the Chitinophagaceae, enriched in the active fraction, may play important ecological roles following wetting. Based on the enrichment of COGs in the active fraction, predation by phage and other bacterial members, as well as scavenging and recycling of labile nutrients, appear to be important ecological processes soon after wetting. To our knowledge, this is the first time BONCAT-FACS-Seq has been applied to biocrust samples, and therefore we discuss the potential advantages and shortcomings of coupling metagenomics to BONCAT to intact soil communities such as biocrust. In all, by pairing BONCAT-FACS and metagenomics, we are capable of highlighting the taxa and potential functions that typifies the microbes actively responding to a rain event

Generation of Long Insert Pairs Using a Cre-LoxP Inverse PCR Approach

Large insert mate pair reads have a major impact on the overall success of de novo assembly and the discovery of inherited and acquired structural variants. The positional information of mate pair reads generally improves genome assembly by resolving repeat elements and/or ordering contigs. Currently available methods for building such libraries have one or more of limitations, such as relatively small insert size; unable to distinguish the junction of two ends; and/or low throughput. We developed a new approach, Cre-LoxP Inverse PCR Paired-End (CLIP-PE), which exploits the advantages of (1) Cre-LoxP recombination system to efficiently circularize large DNA fragments, (2) inverse PCR to enrich for the desired products that contain both ends of the large DNA fragments, and (3) the use of restriction enzymes to introduce a recognizable junction site between ligated fragment ends and to improve the self-ligation efficiency. We have successfully created CLIP-PE libraries up to 22 kb that are rich in informative read pairs and low in small fragment background. These libraries have demonstrated the ability to improve genome assemblies. The CLIP-PE methodology can be implemented with existing and future next-generation sequencing platforms

Optimizing de novo genome assembly from PCR-amplified metagenomes

Background Metagenomics has transformed our understanding of microbial diversity across ecosystems, with recent advances enabling de novo assembly of genomes from metagenomes. These metagenome-assembled genomes are critical to provide ecological, evolutionary, and metabolic context for all the microbes and viruses yet to be cultivated. Metagenomes can now be generated from nanogram to subnanogram amounts of DNA. However, these libraries require several rounds of PCR amplification before sequencing, and recent data suggest these typically yield smaller and more fragmented assemblies than regular metagenomes. Methods Here we evaluate de novo assembly methods of 169 PCR-amplified metagenomes, including 25 for which an unamplified counterpart is available, to optimize specific assembly approaches for PCR-amplified libraries. We first evaluated coverage bias by mapping reads from PCR-amplified metagenomes onto reference contigs obtained from unamplified metagenomes of the same samples. Then, we compared different assembly pipelines in terms of assembly size (number of bp in contigs ≥ 10 kb) and error rates to evaluate which are the best suited for PCR-amplified metagenomes. Results Read mapping analyses revealed that the depth of coverage within individual genomes is significantly more uneven in PCR-amplified datasets versus unamplified metagenomes, with regions of high depth of coverage enriched in short inserts. This enrichment scales with the number of PCR cycles performed, and is presumably due to preferential amplification of short inserts. Standard assembly pipelines are confounded by this type of coverage unevenness, so we evaluated other assembly options to mitigate these issues. We found that a pipeline combining read deduplication and an assembly algorithm originally designed to recover genomes from libraries generated after whole genome amplification (single-cell SPAdes) frequently improved assembly of contigs ≥10 kb by 10 to 100-fold for low input metagenomes. Conclusions PCR-amplified metagenomes have enabled scientists to explore communities traditionally challenging to describe, including some with extremely low biomass or from which DNA is particularly difficult to extract. Here we show that a modified assembly pipeline can lead to an improved de novo genome assembly from PCR-amplified datasets, and enables a better genome recovery from low input metagenomes

31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC 2016) : part two

Background The immunological escape of tumors represents one of the main ob- stacles to the treatment of malignancies. The blockade of PD-1 or CTLA-4 receptors represented a milestone in the history of immunotherapy. However, immune checkpoint inhibitors seem to be effective in specific cohorts of patients. It has been proposed that their efficacy relies on the presence of an immunological response. Thus, we hypothesized that disruption of the PD-L1/PD-1 axis would synergize with our oncolytic vaccine platform PeptiCRAd. Methods We used murine B16OVA in vivo tumor models and flow cytometry analysis to investigate the immunological background. Results First, we found that high-burden B16OVA tumors were refractory to combination immunotherapy. However, with a more aggressive schedule, tumors with a lower burden were more susceptible to the combination of PeptiCRAd and PD-L1 blockade. The therapy signifi- cantly increased the median survival of mice (Fig. 7). Interestingly, the reduced growth of contralaterally injected B16F10 cells sug- gested the presence of a long lasting immunological memory also against non-targeted antigens. Concerning the functional state of tumor infiltrating lymphocytes (TILs), we found that all the immune therapies would enhance the percentage of activated (PD-1pos TIM- 3neg) T lymphocytes and reduce the amount of exhausted (PD-1pos TIM-3pos) cells compared to placebo. As expected, we found that PeptiCRAd monotherapy could increase the number of antigen spe- cific CD8+ T cells compared to other treatments. However, only the combination with PD-L1 blockade could significantly increase the ra- tio between activated and exhausted pentamer positive cells (p= 0.0058), suggesting that by disrupting the PD-1/PD-L1 axis we could decrease the amount of dysfunctional antigen specific T cells. We ob- served that the anatomical location deeply influenced the state of CD4+ and CD8+ T lymphocytes. In fact, TIM-3 expression was in- creased by 2 fold on TILs compared to splenic and lymphoid T cells. In the CD8+ compartment, the expression of PD-1 on the surface seemed to be restricted to the tumor micro-environment, while CD4 + T cells had a high expression of PD-1 also in lymphoid organs. Interestingly, we found that the levels of PD-1 were significantly higher on CD8+ T cells than on CD4+ T cells into the tumor micro- environment (p < 0.0001). Conclusions In conclusion, we demonstrated that the efficacy of immune check- point inhibitors might be strongly enhanced by their combination with cancer vaccines. PeptiCRAd was able to increase the number of antigen-specific T cells and PD-L1 blockade prevented their exhaus- tion, resulting in long-lasting immunological memory and increased median survival