Modification of Rifamycin Polyketide Backbone Leads to Improved Drug Activity Against Rifampicin-Resistant Mycobacterium tuberculosis

Rifamycin B, a product of Amycolatopsis mediterranei S699, is the precursor of clinically used antibiotics that are effective against tuberculosis, leprosy and AIDS related mycobacterial infections. However, prolonged usage of these antibiotics has resulted in the emergence of rifamycin resistant strains of Mycobacterium tuberculosis. As part of our effort to generate better analogs of rifamycin, we substituted the acyltransferase (AT) domain of module 6 of rifamycin polyketide synthase (rifPKS) with that of module 2 of rapamycin PKS. The resulting mutants (rifAT6::rapAT2) of A. mediterranei S699 produced new rifamycin analogs, 24-desmethylrifamycin B and 24-desmethylrifamycin SV, which contained modification in the polyketide backbone. 24-desmethylrifamycin B was then converted to 24-desmethylrifamycin S, whose structure was confirmed by MS, NMR, and X-ray crystallography. Subsequently, 24-desmethylrifamycin S was converted to 24-desmethylrifampicin, which showed excellent antibacterial activity against several rifampicin-resistant M. tuberculosis strains.This research was originally published in the Journal of Biological Chemistry. Nigam, A., Almabruk, K. H., Saxena, A., Yang, J., Mukherjee, U., Kaur, H., ... & Lal, R. Modification of Rifamycin Polyketide Backbone Leads to Improved Drug Activity against Rifampicin-resistant Mycobacterium tuberculosis. Journal of Biological Chemistry. 2014. 289:21142-21152. © the American Society for Biochemistry and Molecular Biology. This is an author's peer-reviewed final manuscript, as accepted by the publisher. The published article is copyrighted by The American Society for Biochemistry and Molecular Biology, Inc., and can be found at: http://www.jbc.org/Keywords: Domain Swapping, Polyketide Synthase, Rifamycin analogs, Multiple Drug Resistant, 24-desmethylrifamyci

Persistencia de malezas gramíneas en cultivos de trigo del sudeste bonaerense

En la presente tesis se estudió la persistencia de especies poáceas en cultivos de trigo del sudeste de Buenos Aires. En dicha región, Avena fatua L. y Lolium multiflorum Lam. son las malezas poáceas más importantes, tanto por la dificultad de control como por sus efectos competitivos sobre el cultivo. A los efectos de cuantificar la persistencia de dichas especies, se estudió la composición de la comunidad de malezas en dos momentos del ciclo: preaplicación de herbicidas y precosecha. Individuos de ambas malezas fueron registrados en ambos momentos como consecuencia de “escapes” al control realizado con herbicidas, siendo A. fatua más constante que L. mutiflorum. Posteriormente, se estudiaron los procesos que definen la persistencia de ambas malezas. Los resultados obtenidos indican que el ajuste del momento de emergencia es jerárquicamente el factor más importante para explicar la persistencia de A. fatua. Se demostró que los modelos de germinación son diferentes según las semillas provengan de un lote agrícola o de una condición de no cultivo, siendo estas diferencias de naturaleza genética. Por otro lado, la variabilidad en la supervivencia a los herbicidas es el factor que mejor explica la persistencia de L. multiflorum, habiéndose documentado resistencia cruzada a los herbicidas inhibidores de la ALS, pyroxsulam, imazamox y flucarbazone, sin antecedentes previos en la región. Los índices de resistencia encontrados presentan variación con la temperatura ambiente en post-aplicación del herbicida, habiéndose registrado mayor resistencia con mayor temperatura. Además, se comprobó que los individuos resistentes presentan menor tiempo a floración que los susceptibles. Tal atributo puede significar una ventaja demográfica para dichas poblaciones. Queda así demostrada la persistencia de A. fatua y L. multiflorum durante el ciclo del cultivo más allá de las prácticas de control realizadas y la participación de dos procesos demográficos distintos (establecimiento y supervivencia) en dicha persistencia

MahmudTaifoPharmacyModificationRifamycinPolyketide.pdf

Rifamycin B, a product of Amycolatopsis mediterranei S699, is the precursor of clinically used antibiotics that are effective against tuberculosis, leprosy and AIDS related mycobacterial infections. However, prolonged usage of these antibiotics has resulted in the emergence of rifamycin resistant strains of Mycobacterium tuberculosis. As part of our effort to generate better analogs of rifamycin, we substituted the acyltransferase (AT) domain of module 6 of rifamycin polyketide synthase (rifPKS) with that of module 2 of rapamycin PKS. The resulting mutants (rifAT6::rapAT2) of A. mediterranei S699 produced new rifamycin analogs, 24-desmethylrifamycin B and 24-desmethylrifamycin SV, which contained modification in the polyketide backbone. 24-desmethylrifamycin B was then converted to 24-desmethylrifamycin S, whose structure was confirmed by MS, NMR, and X-ray crystallography. Subsequently, 24-desmethylrifamycin S was converted to 24-desmethylrifampicin, which showed excellent antibacterial activity against several rifampicin-resistant M. tuberculosis strains.Keywords: Multiple Drug Resistant, Domain Swapping, Polyketide Synthase, Rifamycin analogs, 24-desmethylrifamycinKeywords: Multiple Drug Resistant, Domain Swapping, Polyketide Synthase, Rifamycin analogs, 24-desmethylrifamycinKeywords: Multiple Drug Resistant, Domain Swapping, Polyketide Synthase, Rifamycin analogs, 24-desmethylrifamycinKeywords: Multiple Drug Resistant, Domain Swapping, Polyketide Synthase, Rifamycin analogs, 24-desmethylrifamyci

Comparative Metagenomic Analysis of Soil Microbial Communities across Three Hexachlorocyclohexane Contamination Levels

This paper presents the characterization of the microbial community responsible for the in-situ bioremediation of hexachlorocyclohexane (HCH). Microbial community structure and function was analyzed using 16S rRNA amplicon and shotgun metagenomic sequencing methods for three sets of soil samples. The three samples were collected from a HCH-dumpsite (450 mg HCH/g soil) and comprised of a HCH/soil ratio of 0.45, 0.0007, and 0.00003, respectively. Certain bacterial; (Chromohalobacter, Marinimicrobium, Idiomarina, Salinosphaera, Halomonas, Sphingopyxis, Novosphingobium, Sphingomonas and Pseudomonas), archaeal; (Halobacterium, Haloarcula and Halorhabdus) and fungal (Fusarium) genera were found to be more abundant in the soil sample from the HCH-dumpsite. Consistent with the phylogenetic shift, the dumpsite also exhibited a relatively higher abundance of genes coding for chemotaxis/motility, chloroaromatic and HCH degradation (lin genes). Reassembly of a draft pangenome of Chromohalobacter salaxigenes sp. (~8X coverage) and 3 plasmids (pISP3, pISP4 and pLB1; 13X coverage) containing lin genes/clusters also provides an evidence for the horizontal transfer of HCH catabolism genes

Phylogenetic analysis of the microbiomes.

<p>(A) Dual dendrogram of top 50 bacterial genera across three metagenomes obtained after TEFAP analysis using four bacterial primer sets. Genera and sample categories were clustered using Manhattan distance metric, top 50 genera with standard deviation >0.4 and having at least 0.8% of the total abundance were selected. Colour scale is representing the relative abundance of sequence reads (normalized by sample-mean). (B) Phylogenetic correlation of microbial communities across increasing HCH contamination, a subset of 1000 randomly selected OTUs from each metagenome was used to construct an elucidan distance matrix. Matrices were pair-wise compared using Mantel-test (1000 permutation, 0.05 as standard <i>P</i> -value) and Pearson correlation values were calculated. Asterisks indicate the statistical significance <i>P</i><0.001(mean±sm). (C) Relative percentage of reads assigned to different archeal (I) and fungal (II) genera in TEFAP analysis.</p

Enrichment of lindane degradation (aerobic) pathway.

<p>Schematic representation for the enrichment of aerobic degradation pathway of lindane. Numerical values (on color gradient) at each enzyme represent the diversity (genera) of the corresponding gene present at each metagenome estimated using Transpipe analysis.</p

Chemical properties and sequencing data of soil gradients with HCH gradient.

<p>∑HCH: represents the sum of α and β HCH isomers concentration.</p><p>Salinity levels are representing the EC and cation concentration.</p