Non-encapsulated, encapsulated, and lyophilized probiotic Limosilactobacillus reuteri SW23 influenced the growth and gut health in calves.

The present study was conducted to assess the impact of non-encapsulated, air-dried microencapsulated, and lyophilized microencapsulated probiotics in indigenous cattle calves (Bos indicus). Twenty-four (5-7 days old) indigenous cattle calves were selected and assigned into four groups, with six calves in each as follows: control (CON), fed milk and basal diet alone, and treatment groups supplemented with non-encapsulated (NEC), air-dried microencapsulated (AEC) and lyophilized microencapsulated (LEC) probiotic L. reuteri SW23 at 108 CFU/head/day in skim milk as a carrier provided for 60 days. The animals were divided into four groups, adopting a complete randomized design, and the effects were considered significant at p ≤ 0.05. Probiotics supplementation increased (p < 0.05) body weight gain (kg), average daily gain, and structural growth measurements in calves of all treatment groups. Dry matter intake (g/d), feed conversion efficiency, and fecal counts of Lactobacilli and Bifidobacteria were also increased in the treatment groups compared to CON. The fecal consistency index was highest in CON (0.70 ± 0.03), followed by NEC (0.68 ± 0.01), AEC (0.66 ± 0.02), and LEC (0.65 ± 0.02). Fecal pH and ammonia levels were reduced (p < 0.05) in the probiotic-fed groups compared to CON, with a concomitant increase in fecal lactate, acetate, and propionate levels. In addition, cell-mediated and humoral immunity were significantly increased in supplemented groups as compared to CON. Thus, it can be concluded that supplementation of the probiotics in microencapsulated/non-encapsulated forms to neonatal calves had a variety of positive effects on their health, including better performance, improved gut health, and a lower fecal consistency index. Moreover, among all supplemented groups, the lyophilized microencapsulated group outperformed air-dried microencapsulated and non-microencapsulated groups in terms of ADG, DMI, and gut health.

Multiscale Process Modelling in Translational Systems Biology of Leishmania major: A Holistic view.

Present work aims to utilize systems biology and molecular modelling approach to understand the inhibition kinetics of Leishmania major GLO I and identifying potential hit followed by their validation through in vitro and animal studies. Simulation of GLO I inhibition has shown to affect reaction fluxes of almost all reactions in the model that led to increased production of various AGEs and free radicals. Further, in vitro testing of C1 and C2, selected through molecular modelling revealed remarkable morphological alterations like size reduction, membrane blebbing and loss in motility of the parasite, however, only C1 showed better antileishmanial activity. Additionally, C1 showed apoptosis mediated leishmanicidal activity (apoptosis-like cell death) along with cell-cycle arrest at sub-G0/G1 phase and exhibited potent anti-leishmanial effect against intracellular amastigotes. Furthermore, decrease in parasite load was also observed in C1 treated BALB/c female mice. Our results indicate that C1 has healing effect in infected mice and effectively reduced the parasitic burden. Hence, we suggest C1 as a lead molecule which on further modification, may be used to develop novel therapeutics against Leishmaniasis.

Integrative Computational Framework for Understanding Metabolic Modulation in Leishmania.

Background: The integration of computational and mathematical approaches is used to provide a key insight into the biological systems. Through systems biology approaches we seek to find detailed and more robust information on Leishmanial metabolic network. Forman/Forman-Ricci curvature measures were applied to identify important nodes in the network(s). This was followed by flux balance analysis (FBA) to decipher important drug targets. Results: Our results revealed several key high curvature nodes (metabolites) belonging to common yet crucial metabolic networks, thus, maintaining the integrity of the network which signifies its robustness. Further analysis revealed the presence of some of these metabolites, MGO, in redox metabolism of the parasite. Being a component in the glyoxalase pathway and highly cytotoxic, we further attempted to study the outcome of the deletion of the key enzyme (GLOI) mainly involved in the neutralization of MGO by utilizing FBA. The model and the objective function kept as simple as possible demonstrated an interesting emergent behavior. The non-functional GLOI in the model contributed to "zero" flux which signifies the key role of GLOI as a rate limiting enzyme. This has led to several fold increase production of MGO, thereby, causing an increased level of MGO•- generation. Conclusions: The integrated computational approaches have deciphered GLOI as a potential target both from curvature measures as well as FBA which could further be explored for kinetic modeling by implying various redox-dependent constraints on the model. Furthermore, a constraint-based FBA on a larger model could further be explored to get broader picture to understand the exact underlying mechanisms. Designing various in vitro experimental perspectives could churn the therapeutic importance of GLOI.

In silico pharmacophore modeling and simulation studies for searching potent antileishmanials targeted against Leishmania donovani nicotinamidase.

Nicotinamidase is a key enzyme for the salvage pathway catalyzing the first step for the conversion of nicotinamide (NAm) to nicotinic acid (NA) required for the synthesis of Nicotinamide Adenine Dinucleotide (NAD+) in the subsequent steps. Leishmania protozoan parasites are NAD+ auxotrophs and need precursors (nicotinamide, nicotinic acid, nicotinamide riboside) from their host environment to synthesize NAD+ for their survival. Interestingly, absence of this enzyme in higher eukaryotes and its absolute requirement in the developmental cycle of Leishmania has led nicotinamidase an attractive drug target towards leishmaniasis. Hence, we report some potential inhibitors for nicotinamidase screened based on 3-D pharmacophore model consisting of "ML", "Hyd|Aro", "Acc" and "Excl vol" features. Subsequently, dynamics simulation studies validate the proposed pharmacophore model suggesting its reliability for future studies. Furthermore, these essential site-specific features will help in enhancing the inhibition of nicotinamidase activity. Results of our study suggest that blocking of active site of nicotinamidase by the identified lead inhibitor will have major impact on the infectious processes and the survival of the parasite. Furthermore, due to the structural homology in the enzyme among L. donovani, L. infantum, L. major, we anticipate that our study would help to design more potent drug candidates against leshmaniasis for these three species.

Understanding the Cross-Talk of Redox Metabolism and Fe-S Cluster Biogenesis in Leishmania Through Systems Biology Approach.

Leishmania parasites possess an exceptional oxidant and chemical defense mechanism, involving a very unique small molecular weight thiol, trypanothione (T[SH]2), that helps the parasite to manage its survival inside the host macrophage. The reduced state of T[SH]2 is maintained by NADPH-dependent trypanothione reductase (TryR) by recycling trypanothione disulfide (TS2). Along with its most important role as central reductant, T[SH]2 have also been assumed to regulate the activation of iron-sulfur cluster proteins (Fe/S). Fe/S clusters are versatile cofactors of various proteins and execute a much broader range of essential biological processes viz., TCA cycle, redox homeostasis, etc. Although, several Fe/S cluster proteins and their roles have been identified in Leishmania, some of the components of how T[SH]2 is involved in the regulation of Fe/S proteins remains to be explored. In pursuit of this aim, a systems biology approach was undertaken to get an insight into the overall picture to unravel how T[SH]2 synthesis and reduction is linked with the regulation of Fe/S cluster proteins and controls the redox homeostasis at a larger scale. In the current study, we constructed an in silico kinetic model of T[SH]2 metabolism. T[SH]2 reduction reaction was introduced with a perturbation in the form of its inhibition to predict the overall behavior of the model. The main control of reaction fluxes were exerted by TryR reaction rate that affected almost all the important reactions in the model. It was observed that the model was more sensitive to the perturbation introduced in TryR reaction, 5 to 6-fold. Furthermore, due to inhibition, the T[SH]2 synthesis rate was observed to be gradually decreased by 8 to 14-fold. This has also caused an elevated level of free radicals which apparently affected the activation of Fe/S cluster proteins. The present kinetic model has demonstrated the importance of T[SH]2 in leishmanial cellular redox metabolism. Hence, we suggest that, by designing highly potent and specific inhibitors of TryR enzyme, inhibition of T[SH]2 reduction and overall inhibition of most of the downstream pathways including Fe/S protein activation reactions, can be accomplished.

Efflux pumps and antimicrobial resistance: Paradoxical components in systems genomics.

Efflux pumps play a major role in the increasing antimicrobial resistance rendering a large number of drugs of no use. Large numbers of pathogens are becoming multidrug resistant due to inadequate dosage and use of the existing antimicrobials. This leads to the need for identifying new efflux pump inhibitors. Design of novel targeted therapies using inherent complexity involved in the biological network modeling has gained increasing importance in recent times. The predictive approaches should be used to determine antimicrobial activities with high pathogen specificity and microbicidal potency. Antimicrobial peptides, which are part of our innate immune system, have the ability to respond to infections and have gained much attention in making resistant strain sensitive to existing drugs. In this review paper, we outline evidences linking host-directed therapy with the efflux pump activity to infectious disease.

Comparative analysis of different DNA-binding drugs for leishmaniasis cure: a pharmacoinformatics approach.

Several experiments have been performed to test DNA-binding drugs to cure Leishmania infection. However, there are no details of pharmacoinformatics study. Herein, we have selected a good number of compounds from experimentally verified studies and performed a comparative analysis based on pharmacoinformatics techniques. In silico docking study was performed to observe the molecular level interactions of these known ligands with the DNA receptor by automated computational docking using Glide. A comparison between the calculated interaction energies and in silico ADME/T study was made. In agreement with drug likeness rules, our study suggests that seco-hydroxy-aza-CBI-TMI (compound 4b; GScore, -12.058) is a potential molecule for targeting the DNA to cure leishmaniasis.

Comparative multivariate analysis of codon and amino acid usage in three Leishmania genomes.

Multivariate analysis of codon and amino acid usage was performed for three Leishmania species, including L. donovani, L. infantum and L. major. It was revealed that all three species are under mutational bias and translational selection. Lower GC12 and higher GC3S in all three parasites suggests that the ancestral highly expressed genes (HEGs), compared to lowly expressed genes (LEGs), might have been rich in AT-content. This also suggests that there must have been a faster rate of evolution under GC-bias in LEGs. It was observed from the estimation of synonymous/non-synonymous substitutions in HEGs that the HEG dataset of L. donovani is much closer to L. major evolutionarily. This is also supported by the higher dN value as compared to dS between L. donovani and L. major, suggesting the conservation of synonymous codon positions between these two species and the role of translational selection in shaping the composition of protein-coding genes.