LiLA: lipid lung-based ATLAS built through a comprehensive workflow designed for an accurate lipid annotation.

Accurate lipid annotation is crucial for understanding the role of lipids in health and disease and identifying therapeutic targets. However, annotating the wide variety of lipid species in biological samples remains challenging in untargeted lipidomic studies. In this work, we present a lipid annotation workflow based on LC-MS and MS/MS strategies, the combination of four bioinformatic tools, and a decision tree to support the accurate annotation and semi-quantification of the lipid species present in lung tissue from control mice. The proposed workflow allowed us to generate a lipid lung-based ATLAS (LiLA), which was then employed to unveil the lipidomic signatures of the Mycobacterium tuberculosis infection at two different time points for a deeper understanding of the disease progression. This workflow, combined with manual inspection strategies of MS/MS data, can enhance the annotation process for lipidomic studies and guide the generation of sample-specific lipidome maps. LiLA serves as a freely available data resource that can be employed in future studies to address lipidomic alterations in mice lung tissue.

Detection of Mycobacterium tuberculosis in human tissue via RNA in situ hybridization.

Rationale: Accurate TB diagnosis is hampered by the variable efficacy of the widely-used Ziehl-Neelsen (ZN) staining method to identify Mycobacterium tuberculosis ( Mtb ) acid-fast bacilli (AFB). Here, we sought to circumvent this current limitation through direct detection of Mtb mRNA. Objectives: To employ RNAscope to determine the spatial distribution of Mtb mRNA within tuberculous human tissue, to appraise ZN-negative tissue from confirmed TB patients, and to provide proof-of-concept of RNAscope as a platform to inform TB diagnosis and Mtb biology. Methods: We examined ante- and postmortem human TB tissue using RNAscope to detect Mtb mRNA and a dual ZN/immunohistochemistry staining approach to identify AFB and bacilli producing antigen 85B (Ag85B). Measurements and main results: We adapted RNAscope for Mtb and identified intact and disintegrated Mtb bacilli and intra- and extracellular Mtb mRNA. Mtb mRNA was distributed zonally within necrotic and non-necrotic granulomas. We also found Mtb mRNA within, and adjacent to, necrotic granulomas in ZN-negative lung tissue and in Ag85B-positive bronchial epithelium. Intriguingly, we observed accumulation of Mtb mRNA and Ag85B in the cytoplasm of host cells. Notably, many AFB were negative for Ag85B staining. Mtb mRNA was observed in ZN-negative antemortem lymph node biopsies. Conclusions: RNAscope has diagnostic potential and can guide therapeutic intervention as it detects Mtb mRNA and morphology in ZN-negative tissues from TB patients, and Mtb mRNA in ZN-negative antemortem biopsies, respectively. Lastly, our data provide evidence that at least two phenotypically distinct populations of Mtb bacilli exist in vivo .

NAD(H) homeostasis underlies host protection mediated by glycolytic myeloid cells in tuberculosis.

Mycobacterium tuberculosis (Mtb) disrupts glycolytic flux in infected myeloid cells through an unclear mechanism. Flux through the glycolytic pathway in myeloid cells is inextricably linked to the availability of NAD+, which is maintained by NAD+ salvage and lactate metabolism. Using lung tissue from tuberculosis (TB) patients and myeloid deficient LDHA (LdhaLysM-/-) mice, we demonstrate that glycolysis in myeloid cells is essential for protective immunity in TB. Glycolytic myeloid cells are essential for the early recruitment of multiple classes of immune cells and IFNγ-mediated protection. We identify NAD+ depletion as central to the glycolytic inhibition caused by Mtb. Lastly, we show that the NAD+ precursor nicotinamide exerts a host-dependent, antimycobacterial effect, and that nicotinamide prophylaxis and treatment reduce Mtb lung burden in mice. These findings provide insight into how Mtb alters host metabolism through perturbation of NAD(H) homeostasis and reprogramming of glycolysis, highlighting this pathway as a potential therapeutic target.

Plant growth-promoting bacterial consortia improved the physiology and growth of maize by regulating osmolytes and antioxidants balance under salt-affected field conditions.

This study was undertaken to see how microbial consortia influenced maize development and yield under salt-affected conditions. The efficacy of the pre-isolated bacterial strains Burkholderia phytofirmans, Bacillussubtilis, Enterobacter aerogenes, and Pseudomonas syringae and Pseudomonas fluorescens to decrease the detrimental effects of salt on maize was tested in four distinct combinations using Randomized Complete Block Design with three replicates. The results revealed that these strains were compatible and collaborated synergistically, with an 80% co-aggregation percentage under salt-affected conditions. Following that, these strains were tested for their ability to increase maize growth and yield under salt-affected field conditions. The photosynthetic rate (11-50%), relative water content (10-34%), and grain yield (13-21%) of maize were all increased by these various combinations. However, when Burkholderia phytofirmans, Enterobacter aerogenes and Pseudomonas fluorescens were combined, the greatest increase was seen above the un-inoculated control. Furthermore, as compared to the un-inoculated control, the same combination resulted in a 1.5-fold increase in catalase and a 2.0-fold increase in ascorbate concentration. These findings showed that a multi-strain consortium might boost maize's total yield response as a result of better growth under salt stress.

Converging alfalfa (Medicago sativa L.) and petroleum hydrocarbon acclimated ACC-deaminase containing bacteria for phytoremediation of petroleum hydrocarbon contaminated soil.

Plant assisted bioremediation of petroleum hydrocarbon contaminated soil is considered an effective green technology whereby accelerated degradation occurs due to converged effect of microorganisms and plants. However, survival and growth of microbes and plants under stress conditions is challenging task for success of the technology. In this study, plant growth promoting bacteria containing 1-aminocyclopropane-1-carboxylate (ACC)-deaminase activity and tolerance to petroleum hydrocarbon contamination were used in association with alfalfa for bioremediation of petroleum hydrocarbon contaminated soil. Eight pre-isolated bacterial isolates from soil having previous history of petroleum contamination were used in convergence with alfalfa on sand soil which was artificially contaminated (10 g crude oil per kg-1 of coarse textured soil). Combined effect of bacteria and plants on the degradation of petroleum hydrocarbons under controlled conditions of light and temperature was observed for a period of 60 days. The results of the study revealed that four bacterial isolates Bacillus subtilis strain PM32Y, Bacillus cereus strain WZ3S1, Bacillus sp. strain SM73 and Bacillus sp. strain WZ3S3 in association with alfalfa significantly degraded petroleum hydrocarbons. The most significant biodegradation (47%) of petroleum hydrocarbons was recorded in the experimental unit receiving PM32Y inoculation in association with alfalfa. Biodegradation of petroleum hydrocarbons was 33% with alone inoculation (without alfalfa) of PM32Y. The study revealed that combined use of bacteria and alfalfa plant is more efficient than alone application of either bacteria or plants for degradation of petroleum hydrocarbons.

This study provides the evidence for phytoremediation and significant degradation of petroleum hydrocarbons by using plant growth promoting bacteria (PGPB), containing 1-aminocyclopropane-1-carboxylate deaminase (ACC-deaminase) in association with alfalfa (Medicago sativa L.). The most significant biodegradation of petroleum hydrocarbons was recorded with a new combination of Bacillus subtilis strain PM32Y in association with alfalfa.

Targeting dendritic cells with TLR-2 ligand-coated nanoparticles loaded with Mycobacterium tuberculosis epitope induce antituberculosis immunity.

Novel vaccination strategies are crucial to efficiently control tuberculosis, as proposed by the World Health Organization under its flagship program "End TB Strategy." However, the emergence of drug-resistant strains of Mycobacterium tuberculosis (Mtb), particularly in those coinfected with HIV-AIDS, constitutes a major impediment to achieving this goal. We report here a novel vaccination strategy that involves synthesizing a formulation of an immunodominant peptide derived from the Acr1 protein of Mtb. This nanoformulation in addition displayed on the surface a toll-like receptor-2 ligand to offer to target dendritic cells (DCs). Our results showed an efficient uptake of such a concoction by DCs in a predominantly toll-like receptor-2-dependent pathway. These DCs produced elevated levels of nitric oxide, proinflammatory cytokines interleukin-6, interleukin-12, and tumor necrosis factor-α, and upregulated the surface expression of major histocompatibility complex class II molecules as well as costimulatory molecules such as CD80 and CD86. Animals injected with such a vaccine mounted a significantly higher response of effector and memory Th1 cells and Th17 cells. Furthermore, we noticed a reduction in the bacterial load in the lungs of animals challenged with aerosolized live Mtb. Therefore, our findings indicated that the described vaccine triggered protective anti-Mtb immunity to control the tuberculosis infection.

Potential of plant growth promoting bacterial consortium for improving the growth and yield of wheat under saline conditions.

Owing to inconsistent results of a single bacterial strain, co-inoculation of more than one strain under salinity stress could be a more effective strategy to induce salt tolerance. Co-inoculation of more than one bacterial strain could be more effective due to the presence of several growths promoting traits. This study was conducted to evaluate the effectiveness of multi-strains bacterial consortium to promote wheat growth under salinity stress. Several plant growth promoting rhizobacteria (PGPR) had been isolated and tested for their ability to grow in increasing concentrations of sodium chloride (NaCl). Those rhizobacterial strains having tolerance against salinity were screened to evaluate their ability to promote wheat growth in the presence of salinity by conducting jar trials under axenic conditions. The rhizobacteria with promising results were tested for their compatibility with each other before developing multi-strain inoculum of PGPR. The compatible PGPR strains were characterized, and multi-strain inoculum was then evaluated for promoting wheat growth under axenic conditions at different salinity levels, i.e., 2.1 (normal soil), 6, 12, and 18 dS m-1. The most promising combination was further evaluated by conducting a pot trial in the greenhouse. The results showed that compared to a single rhizobacterial strain, better growth-promoting effect was observed when rhizobacterial strains were co-inoculated. The multi-strain consortium of PGPR caused a significant positive impact on shoot length, root length, shoot fresh weight, and root fresh weight of wheat at the highest salinity level in the jar as well as in the pot trial. Results showed that the multi-strain consortium of PGPR caused significant positive effects on the biochemical traits of wheat by decreasing electrolyte leakage and increasing chlorophyll contents, relative water contents (RWC), and K/Na ratio. It can be concluded that a multi-strain consortium of PGPR (Ensifer adhaerens strain BK-30, Pseudomonas fluorescens strain SN5, and Bacillus megaterium strain SN15) could be more effective to combat the salinity stress owing to the presence of a variety of growth-promoting traits. However, further work is going on to evaluate the efficacy of multi-strain inoculum of PGPR under salt-affected field conditions.

Phycoremediation of textile effluents with enhanced efficacy of biodiesel production by algae and potential use of remediated effluent for improving growth of wheat.

The uncontrolled industrialization and unrestricted textile production combined with inappropriate effluent treatment services in developing countries like Pakistan have multiplied the number of harmful effluent discharge. These effluents are enriched with dyes, heavy metal ions, and other hazardous materials that are poisonous and carcinogenic to living organisms. For that reason, the utilization of economic and efficient control techniques against such pollutants is imperative to protect natural resources. The triple algal role for phycoremediation of textile effluent was utilized in this study to make it suitable for irrigation and higher biofuel production. Locally isolated two strains, CKW1 (Spirogyra sp.) and PKS33 (Cladophora sp.), were used to treat the effluent collected from the direct outlets of the textile industries. The treated effluent was then tested for its toxicity and applied to wheat at initial stage grown under axenic conditions to check its effect on wheat (Triticum aestivum L.) vegetative growth and development. Finally, the algal biomass obtained after treatment was subjected to trans-esterification for predicting the amount of biodiesel production. Study outcomes revealed that the algal strains were able to decolorize the effluent entirely within 96-120 h. Compared to un-treated textile effluent, the phycoremediated wastewater application to wheat plants enhanced the plant biomass by 80%. Lastly, the production of biodiesel from algal biomass attained after phycoremediation was 35% less to algal biomass obtained under normal growth conditions. It can be concluded that the algal use helps to treat the contaminated effluent and marks them re-usable for irrigating plants and producing biomass which could be utilized for biodiesel production.

Water-soluble tocopherol derivatives inhibit SARS-CoV-2 RNA-dependent RNA polymerase.

The recent emergence of a novel coronavirus, SARS-CoV-2, has led to the global pandemic of the severe disease COVID-19 in humans. While efforts to quickly identify effective antiviral therapies have focused largely on repurposing existing drugs 1-4 , the current standard of care, remdesivir, remains the only authorized antiviral intervention of COVID-19 and provides only modest clinical benefits 5 . Here we show that water-soluble derivatives of α-tocopherol have potent antiviral activity and synergize with remdesivir as inhibitors of the SARS-CoV-2 RNA-dependent RNA polymerase (RdRp). Through an artificial-intelligence-driven in silico screen and in vitro viral inhibition assay, we identified D-α-tocopherol polyethylene glycol succinate (TPGS) as an effective antiviral against SARS-CoV-2 and ß-coronaviruses more broadly that also displays strong synergy with remdesivir. We subsequently determined that TPGS and other water-soluble derivatives of α-tocopherol inhibit the transcriptional activity of purified SARS-CoV-2 RdRp and identified affinity binding sites for these compounds within a conserved, hydrophobic interface between SARS-CoV-2 nonstructural protein 7 and nonstructural protein 8 that is functionally implicated in the assembly of the SARS-CoV-2 RdRp 6 . In summary, we conclude that solubilizing modifications to α-tocopherol allow it to interact with the SARS-CoV-2 RdRp, making it an effective antiviral molecule alone and even more so in combination with remdesivir. These findings are significant given that many tocopherol derivatives, including TPGS, are considered safe for humans, orally bioavailable, and dramatically enhance the activity of the only approved antiviral for SARS-CoV-2 infection 7-9 .

Biochar Mediated-Alleviation of Chromium Stress and Growth Improvement of Different Maize Cultivars in Tannery Polluted Soils.

Soil pollution with heavy metal is a serious problem across the globe and is on the rise due to the current intensification of chemical industry. The leather industry is one of them, discharging chromium (Cr) in huge quantities during the process of leather tanning and polluting the nearby land and water resources, resulting in deterioration of plant growth. In this study, the effects of biochar application at the rate of 3% were studied on four maize cultivars, namely NK-8441, P-1543, NK-8711, and FH-985, grown in two different tannery polluted Kasur (K) and Sialkot (S) soils. Maize plants were harvested at vegetative growth and results showed that Cr toxicity adversely not only affected their growth, physiology, and biochemistry, but also accumulated in their tissues. However, the level of Cr toxicity, accumulation, and its influence on maize cultivars varied greatly in both soils. In this pot experiment, biochar application played a crucial role in lessening the Cr toxicity level, resulting in significant increase in plant height, biomass (fresh and dry), leaf area, chlorophyll pigments, photosynthesis, and relative water content (RWC) over treatment set as a control. However, applied biochar significantly decreased the electrolyte leakage (EL), antioxidant enzymes, lipid peroxidation, proline content, soluble sugars, and available fraction of Cr in soil as well as Cr (VI and III) concentration in root and shoot tissues of maize plant. In addition to this, maize cultivar differences were also found in relation to their tolerance to Cr toxicity and cultivar P-1543 performed better over other cultivars in both soils. In conclusion, biochar application in tannery polluted soils could be an efficient ecofriendly approach to reduce the Cr toxicity and to promote plant health and growth.

Appraising the potential of EPS-producing rhizobacteria with ACC-deaminase activity to improve growth and physiology of maize under drought stress.

Rhizobacteria containing 1-aminocyclopropane-1-carboxylic acid-deaminase (ACC-deaminase) and exopolysaccharides (EPS) activity are important to induce stress tolerance in plants. The present study was conducted to screen and characterize plant growth-promoting rhizobacteria (PGPR) with ACC-deaminase and EPS-producing activity for improving maize growth under drought stress. Eighty-five rhizobacterial strains were isolated from the rain-fed areas, among those 69 isolates were able to utilize ACC and 31 strains were found positive for EPS production. These strains containing ACC-deaminase and/or EPS-producing activity were subjected to drought tolerance assay by inducing water stress in media using polyethylene glycol 6000. Based on results of the drought tolerance bioassay, 12 most prominent strains were selected to evaluate their growth-promoting abilities in maize under water-stressed conditions by conducting jar trial. The impact of strains on maize growth parameters was variable. Strains with co-existence of ACC-deaminase and EPS-producing activity showed comparatively better results than those with either ACC-deaminase or EPS-producing activity only. These strains were also significantly better in improving the plant physiological parameters including photosynthesis rate, stomatal conductance, vapor pressure, water-use efficiency and transpiration rate. The strain D3 with co-existence of ACC-deaminase and EPS-producing activity was significantly better in colonizing maize roots, improving plant growth and physiological parameters. The strain was named as Bacillus velezensis strain D3 (accession number MT367633) as confirmed through results of 16S rRNA partial gene sequencing. It is concluded that the strains with co-existence of ACC-deaminase and EPS-producing activity could be better suited for improving crop growth and physiology under drought stress.

The Role of Host-Generated H2S in Microbial Pathogenesis: New Perspectives on Tuberculosis.

For centuries, hydrogen sulfide (H2S) was considered primarily as a poisonous gas and environmental hazard. However, with the discovery of prokaryotic and eukaryotic enzymes for H2S production, breakdown, and utilization, H2S has emerged as an important signaling molecule in a wide range of physiological and pathological processes. Hence, H2S is considered a gasotransmitter along with nitric oxide (•NO) and carbon monoxide (CO). Surprisingly, despite having overlapping functions with •NO and CO, the role of host H2S in microbial pathogenesis is understudied and represents a gap in our knowledge. Given the numerous reports that followed the discovery of •NO and CO and their respective roles in microbial pathogenesis, we anticipate a rapid increase in studies that further define the importance of H2S in microbial pathogenesis, which may lead to new virulence paradigms. Therefore, this review provides an overview of sulfide chemistry, enzymatic production of H2S, and the importance of H2S in metabolism and immunity in response to microbial pathogens. We then describe our current understanding of the role of host-derived H2S in tuberculosis (TB) disease, including its influences on host immunity and bioenergetics, and on Mycobacterium tuberculosis (Mtb) growth and survival. Finally, this review discusses the utility of H2S-donor compounds, inhibitors of H2S-producing enzymes, and their potential clinical significance.

A multiple T cell epitope comprising DNA vaccine boosts the protective efficacy of Bacillus Calmette-Guérin (BCG) against Mycobacterium tuberculosis.

BACKGROUND: Approximately 80% - 90% of individuals infected with latent Mycobacterium tuberculosis (Mtb) remain protected throughout their life-span. The release of unique, latent-phase antigens are known to have a protective role in the immune response against Mtb. Although the BCG vaccine has been administered for nine decades to provide immunity against Mtb, the number of TB cases continues to rise, thereby raising doubts on BCG vaccine efficacy. The shortcomings of BCG have been associated with inadequate processing and presentation of its antigens, an inability to optimally activate T cells against Mtb, and generation of regulatory T cells. Furthermore, BCG vaccination lacks the ability to eliminate latent Mtb infection. With these facts in mind, we selected six immunodominant CD4 and CD8 T cell epitopes of Mtb expressed during latent, acute, and chronic stages of infection and engineered a multi-epitope-based DNA vaccine (C6). RESULT: BALB/c mice vaccinated with the C6 construct along with a BCG vaccine exhibited an expansion of both CD4 and CD8 T cell memory populations and augmented IFN-γ and TNF-α cytokine release. Furthermore, enhancement of dendritic cell and macrophage activation was noted. Consequently, illustrating the elicitation of immunity that helps in the protection against Mtb infection; which was evident by a significant reduction in the Mtb burden in the lungs and spleen of C6 + BCG administered animals. CONCLUSION: Overall, the results suggest that a C6 + BCG vaccination approach may serve as an effective vaccination strategy in future attempts to control TB.

Gut Dysbiosis Thwarts the Efficacy of Vaccine Against Mycobacterium tuberculosis.

The generation of enduring protective immunity by vaccines is of utmost importance. Intriguingly, there is considerable variation in the efficacy of vaccines amongst individuals. Various studies have shown that normal flora of gastrointestinal tract plays a vital role in maintaining host homeostasis and immunity. Since gut microbiome is also extremely variable between individuals, we speculate that it might impact individual's response to vaccines. Consequently, we administered broad spectrum antibiotics cocktail to induce gut dysbiosis and monitored its impact on the generation of long-lasting memory T cells and thereby BCG vaccine efficacy. Interestingly, gut dysbiosis significantly decreased the activation of CD4+ T cells and CD8+ T cells. Further, there was decline in the frequency of memory CD4+ T cells and CD8+ T cells in lungs and secondary lymphoid organs of the vaccinated animals. Moreover, it dampened the IFN-γ and TNF-α secretion and proliferation of Mtb-specific T cells. Most importantly, dysbiosis hampered Mtb clearance in vaccinated animals, as evidenced by increase in the colony forming units (CFUs) in lungs and spleen. Our findings indicate that gut dysbiosis can be one of the major factors responsible for variable efficacy of TB vaccines across the world.

Potential Role of Gut Microbiota in Induction and Regulation of Innate Immune Memory.

The gut microbiota significantly regulates the development and function of the innate and adaptive immune system. The attribute of immunological memory has long been linked only with adaptive immunity. Recent evidence indicates that memory is also present in the innate immune cells such as monocytes/macrophages and natural killer cells. These cells exhibit pattern recognition receptors (PRRs) that recognize microbe- or pathogen-associated molecular patterns (MAMPs or PAMPs) expressed by the microbes. Interaction between PRRs and MAMPs is quite crucial since it triggers the sequence of signaling events and epigenetic rewiring that not only play a cardinal role in modulating the activation and function of the innate cells but also impart a sense of memory response. We discuss here how gut microbiota can influence the generation of innate memory and functional reprogramming of bone marrow progenitors that helps in protection against infections. This article will broaden our current perspective of association between the gut microbiome and innate memory. In the future, this knowledge may pave avenues for development and designing of novel immunotherapies and vaccination strategies.

A genomic analysis of Mycobacterium immunogenum strain CD11_6 and its potential role in the activation of T cells against Mycobacterium tuberculosis.

BACKGROUND: Mycobacterium tuberculosis (Mtb) is an etiological agent of tuberculosis (TB). Tuberculosis is a mounting problem worldwide. The only available vaccine BCG protects the childhood but not adulthood form of TB. Therefore, efforts are made continuously to improve the efficacy of BCG by supplementing it with other therapies. Consequently, we explored the possibility of employing Mycobacterium immunogenum (Mi) to improve BCG potential to protect against Mtb. RESULTS: We report here the genome mining, comparative genomics, immunological and protection studies employing strain CD11_6 of Mi. Mycobacterium immunogenum was isolated from duodenal mucosa of a celiac disease patient. The strain was whole genome sequenced and annotated for identification of virulent genes and other traits that may make it suitable as a potential vaccine candidate. Virulence profile of Mi was mapped and compared with two other reference genomes i.e. virulent Mtb strain H37Rv and vaccine strain Mycobacterium bovis (Mb) AFF2122/97. This comparative analysis revealed that Mi is less virulent, as compared to Mb and Mtb, and contains comparable number of genes encoding for the antigenic proteins that predict it as a probable vaccine candidate. Interestingly, the animals vaccinated with Mi showed significant augmentation in the generation of memory T cells and reduction in the Mtb burden. CONCLUSION: The study signifies that Mi has a potential to protect against Mtb and therefore can be a future vaccine candidate against TB.

A lipidated bi-epitope vaccine comprising of MHC-I and MHC-II binder peptides elicits protective CD4 T cell and CD8 T cell immunity against Mycobacterium tuberculosis.

BACKGROUND: The clinical trials conducted at Chingleput India suggest that BCG fails to protect against tuberculosis (TB) in TB-endemic population. Recent studies advocate that non-tuberculous mycobacteria and latent Mycobacterium tuberculosis (Mtb) infection interferes in the antigen processing and presentation of BCG in inducing protective immunity against Mtb. Thereby, indicating that any vaccine that require extensive antigen processing may not be efficacious in TB-endemic zones. Recently, we have demonstrated that the vaccine candidate L91, which is composed of lipidated promiscuous MHC-II binder epitope, derived from latency associated Acr1 antigen of Mtb is immunogenic in the murine and Guinea pig models of TB and conferred better protection than BCG against Mtb. METHODS: In this study, we have used a multi-stage based bi-epitope vaccine, namely L4.8, comprising of MHC-I and MHC-II binding peptides of active (TB10.4) and latent (Acr1) stages of Mtb antigens, respectively. These peptides were conjugated to the TLR-2 agonist Pam2Cys. RESULTS: L4.8 significantly elicited both CD8 T cells and CD4 T cells immunity, as evidenced by increase in the enduring polyfunctional CD8 T cells and CD4 T cells. L4.8 efficiently declined Mtb-burden and protected animals better than BCG and L91, even at the late stage of Mtb infection. CONCLUSIONS: The BCG-L4.8 prime boost strategy imparts a better protection against TB than the BCG alone. This study emphatically denotes that L4.8 can be a promising future vaccine candidate for controlling active and latent TB.

Preliminary study on phosphate solubilizing Bacillus subtilis strain Q3 and Paenibacillus sp. strain Q6 for improving cotton growth under alkaline conditions.

BACKGROUND: Low phosphorus availability limits crop production in alkaline calcareous soils in semi-arid regions including Pakistan. Phosphate solubilizing bacteria may improve crop growth on alkaline calcareous soils due to their ability to enhance P availability. METHODS: Twenty rhizobacterial isolates (Q1-Q20) were isolated from rhizosphere of cotton and characterized for their growth promoting attributes in vitro. The selected phosphate solubilizing isolates were further screened for their ability to improve cotton growth under axenic conditions (jar trial). The phosphorus solubilization capacities of selected strains were quantified and these strains were identified through 16S rDNA sequencing. RESULTS: Isolates Q2, Q3, Q6, Q7, Q8, Q13 and Q14 were able to solubilize phosphate from insoluble sources. Most of these isolates also possessed other traits including catalase activity and ammonia production. The growth promotion assay showed that Q3 was significantly better than most of the other isolates followed by Q6. Maximum root colonization (4.34 × 106 cfu g-1) was observed in case of isolate Q6 followed by Q3. The phosphorus solubilization capacities of these strains were quantified, showing a maximum phosphorus solubilization by Q3 (optical density 2.605 ± 0.06) followed by the Q6 strain. The strain Q3 was identified as Bacillus subtilis (accession # KX788864) and Q6 as Paenibacillus sp. (accession # KX788865) through 16S rDNA sequencing. DISCUSSION: The bacterial isolates varied in their abilities for different growth promoting traits. The selected PGPR Bacillus subtilis strain Q3 and Paenibacillus sp. strain Q6 have multifarious growth promoting traits including ability to grow at higher EC and pH levels, and phosphorus solubilizing ability. These strains can efficiently colonize cotton roots under salt affected soils and help plants in phosphorus nutrition. It is concluded that both strains are potential candidates for promoting cotton growth under alkaline conditions, however further investigation is required to determine their potential for field application.

Nickel phytoextraction through bacterial inoculation in Raphanus sativus.

A pot experiment was conducted to evaluate the potential of two plant growth promoting rhizobacteria (PGPR) viz. Bacillus sp. CIK-516 and Stenotrophomonas sp. CIK-517Y for improving the growth and Ni uptake of radish (Raphanus sativus) in the presence of four different levels of Ni contamination (0, 50, 100, 150 mg Ni kg-1 soil). Plant growth, dry biomass, chlorophyll and nitrogen contents were significantly reduced by the exogenous application of Ni, however, bacterial inoculation diluted the negative impacts of Ni stress on radish by improving these parameters. PGPR strain CIK-516 increased root length (9-27%), shoot length (8-27%), root dry biomass (2-32%), shoot dry biomass (9-51%), root girth (6-48%), total chlorophyll (4-38%) and shoot nitrogen contents (11-15%) in Ni contaminated and non-contaminated soils. Positive regulation of chlorophyll and nitrogen contents by the inoculated plants shows plant tolerance mechanism of Ni stress. Bacterial strain (CIK-516) exhibited indole acetic acid and 1-amino-cyclopropane-1-carboxylate deaminase potentials which would have helped radish plant to stabilize in Ni contaminated soil and thereby increased Ni uptake (24-257 in shoot and 58-609 in root mg kg-1 dry biomass) and facilitated accumulation in radish (bioaccumulation factor = 0.6-1.7) depending upon soil Ni contamination. Based on the findings of this study, it might be suggested that inoculation with bacterial strain CIK-516 could be an efficient tool for enhanced Ni phytoextraction in radish.

A lipidated peptide of Mycobacterium tuberculosis resuscitates the protective efficacy of BCG vaccine by evoking memory T cell immunity.

BACKGROUND: The current BCG vaccine induces only short-term protection against Mycobacterium tuberculosis (Mtb), suggesting its failure to generate long-lasting memory T cells. Previously, we have demonstrated that a self-adjuvanting peptide of Mtb (L91), successfully generated enduring memory Th1 cells. Consequently, we investigated if L91 was able to recuperate BCG potency in perpetuating the generation of memory T cells and protection against Mtb infected mice. METHODS: In the present study, we evaluated the potency of a self adjuvanting Mtb peptide vaccine L91 in invigorating BCG immune response against Mtb in mice. Female BALB/c mice were immunized with BCG. Later, they were boosted twice with L91 or an antigenically irrelevant lipidated influenza virus hemagglutinin peptide (LH). Further, PBMCs obtained from BCG vaccinated healthy subjects were cultured in vitro with L91. T cell responses were determined by surface markers and intracellular cytokine staining. Secretion of cytokines was estimated in the culture supernatants (SNs) by ELISA. RESULTS: Compared to the BCG-vaccinated controls, L91 booster significantly enhanced the percentage of memory Th1 cells and Th17 cells and reduced the mycobacterial burden in BCG primed and L91-boosted (BCG-L91) group, even after 229 days of BCG vaccination. Further, substantial augmentation in the central (CD44hiCD62LhiCD127hi) and effector memory (CD44hiCD62LloCD127lo) CD4 T cells was detected. Furthermore, greater frequency of polyfunctional Th1 cells (IFN-γ+TNF-α+) and Th17 cells (IFN-γ+IL-17A+) was observed. Importantly, BCG-L91 successfully prevented CD4 T cells from exhaustion by decreasing the expression of PD-1 and Tim-3. Additionally, augmentation in the frequency of Th1 cells, Th17 cells and memory CD4 T cells was observed in the PBMCs of the BCG-vaccinated healthy individuals following in vitro stimulation with L91. CONCLUSIONS: Our study demonstrated that L91 robustly reinvigorate BCG potency to invoke enduring protection against Mtb. This novel vaccination stratagem involving BCG-priming followed by L91-boosting can be a future prophylactic measure to control TB.