Swati Jindal

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.

Acinetobacter indicus sp. nov., isolated from a hexachlorocyclohexane dump site.

The taxonomic position of a Gram-negative, non-motile, oxidase negative and catalase positive strain, A648(T), isolated from a hexachlorocyclohexane (HCH) dump site located in Lucknow, India, was ascertained by using a polyphasic approach. A comparative analysis of a partial sequence of the rpoB gene and the 16S rRNA gene sequence revealed that strain A648(T) belonged to the genus Acinetobacter. DNA-DNA relatedness values between strain A648(T) and other closely related members (16S rRNA gene sequence similarity greater than 97%), namely Acinetobacter radioresistens DSM 6976(T), A. venetianus ATCC 31012(T), A. baumannii LMG 1041(T), A. parvus LMG 21765(T) A. junii LMG 998(T) and A. soli JCM 15062(T), were found to be less than 8%. The major cellular fatty acids of strain A648(T) were 18:1ω9c (19.6%), summed feature 3 (15.9%), 16:0 (10.6%) and 12:0 (6.4%). The DNA G+C content was 40.4 mol%. The polar lipid profile of strain A648(T) indicated the presence of diphosphatidylglycerol, phosphatidylethanolamine, followed by phosphatidylglycerol and phosphatidylcholine. The predominant polyamine of strain A648(T) was 1,3-diaminopropane and moderate amounts of putrescine, spermidine and spermine were also detected. The respiratory quinone consisted of ubiquinone with nine isoprene units (Q-9). On the basis of DNA-DNA hybridization, phenotypic characteristics and chemotaxonomic and phylogenetic comparisons with other members of the genus Acinetobacter, strain A648(T) is found to be a novel species of the genus Acinetobacter, for which the name Acinetobacter indicus sp. nov. is proposed. The type strain is A648(T) ( = DSM 25388(T) = CCM 7832(T)).

Pseudomonas sp. to Sphingobium indicum: a journey of microbial degradation and bioremediation of Hexachlorocyclohexane

The unusual process of production of hexachlorocyclohexane (HCH) and extensive use of technical HCH and lindane has created a very serious problem of HCH contamination. While the use of technical HCH and lindane has been banned all over the world, India still continues producing lindane. Bacteria, especially Sphingomonads have been isolated that can degrade HCH isomers. Among all the bacterial strains isolated so far, Sphingobium indicum B90A that was isolated from HCH treated rhizosphere soil appears to have a better potential for HCH degradation. This conclusion is based on studies on the organization of lin genes and degradation ability of B90A. This strain perhaps can be used for HCH decontamination through bioaugmentation.

Sphingopyxis indica sp. nov., isolated from a high dose point hexachlorocyclohexane (HCH)-contaminated dumpsite

A Gram-stain-negative, aerobic, non-motile, non-spore-forming, rod-shaped and light-yellow-pigmented bacterium, designated DS15(T), was isolated from a soil sample collected from a hexachlorocyclohexane dumpsite in Lucknow, Uttar Pradesh, India. Strain DS15(T) showed highest 16S rRNA gene sequence similarity to Sphingopyxis panaciterrulae DCY34(T) (98.7%) and Sphingopyxis soli BL03(T) (98.0%). The 16S rRNA gene sequence similarity between strain DS15(T) and species of genus Sphingopyxis with validly published names ranged from 92.5% to 98.7%. The DNA G+C content of strain DS15(T) was 67.5 mol%. The chemotaxonomic markers in strain DS15(T) were consistent with its classification in the genus Sphingopyxis, i.e. Q-10 as the major ubiquinone and summed feature 8 (C18:1ω7c/C18:1ω9c), C17:1ω6c, summed feature 3 (C16:1ω7c/C16:1ω6c), C14:0 2-OH, C15:0 2-OH, C16:0 and C17:1ω8c as the predominant fatty acids. The major polar lipids of strain DS15(T) were phosphatidylethanolamine (PE), diphosphatidylglycerol (DPG), phosphatidylcholine (PC), phosphatidylglycerol (PG) and sphingoglycolipids (SGL) and spermidine was detected as the major polyamine. Phylogenetic analysis, DNA-DNA hybridization, and chemotaxonomic and phenotypic analysis support the conclusion that strain DS15(T) represents a novel species within the genus Sphingopyxis, for which the name Sphingopyxis indica is proposed. The type strain is DS15(T) (=MTCC 9455(T)=CCM 7542(T)=MCC 2023(T)).

Functional screening of enzymes and bacteria for the dechlorination of hexachlorocyclohexane by a high-throughput colorimetric assay

Two distinct microbial dehalogenases are involved in the first steps of degradation of hexachlorocyclohexane (HCH) isomers. The enzymes, LinA and LinB, catalyze dehydrochlorination and dechlorination reactions of HCH respectively, each with distinct isomer specificities. The two enzymes hold great promise for use in the bioremediation of HCH residues in contaminated soils, although their kinetics and isomer specificities are currently limiting. Here we report the functional screening of a library of 700 LinA and LinB clones generated from soil DNA for improved dechlorination activity by means of a high throughput colorimetric assay. The assay relies upon visual colour change of phenol red in an aqueous medium, due to the pH drop associated with the dechlorination reactions. The assay is performed in a microplate format using intact cells, making it quick and simple to perform and it has high sensitivity, dynamic range and reproducibility. The method has been validated with quantitative gas chromatographic analysis of promising clones, revealing some novel variants of both enzymes with superior HCH degrading activities. Some sphingomonad isolates with potentially superior activities were also identified.

Bacterial diversity and real‐time PCR based assessment of linA and linB gene distribution at hexachlorocyclohexane contaminated sites

The disposal of hexachlorocyclohexane (HCH) muck has created large number of HCH dumpsites all over the world from where the harmful HCH isomers are leaking into the environment. Bacteria have evolved at such contaminated sites that have the ability to degrade HCH. Degradation of various HCH isomers in bacterial strains is mediated primarily by two genes: linA and linB which encode dehydrochlorinase and haloalkane dehalogenase respectively. In this study we explored one such highly contaminated HCH dumpsite located in Lucknow, Uttar Pradesh, India. To assess the biostimulation potential of the contaminated site, microbial diversity study and real‐time PCR based quantification of lin genes was carried out. The soil samples from dumpsite and surrounding areas were found to be highly contaminated with HCH residue levels as high as 1.8 × 105 mg kg−1. The residues were detected in areas upto 13 km from the dumpsite. Sphingomonads, Chromohalobacter, and Marinobacter were the dominant genera present at the dump‐site. Role of Sphingomonads in HCH degradation has been well documented. The highest copy numbers of linA and linB genes as determined using real‐time PCR were 6.2 × 104 and 5.3 × 105, respectively, were found in sample from the dump site. The presence of Sphingomonads, linA, and linB genes from HCH contaminated soil indicates the presence of indigenous bacterial communities capable of HCH degradation.

Bioremediation of Hexachlorocyclohexane (HCH) Pollution at HCH Dump Sites

Globally, the period from early the 1950s to late 1980s has shown an increased use of primarily three pesticides namely DDT.

A new life in a bacterium through synthetic genome: a successful venture by craig venter

The fi rst synthetic genome, a stripped down version of a natural organism, is only the beginning. I now want to go further. My company Synthetic Genomics Inc., is already trying to develop cassettes-modules of genes-to turn an organism into a biofactory that could make clean hydrogen fuel from sunlight and water or soak up more carbon dioxide. From there I want to take us far from shore into unknown waters, to a new phase of evolution, to the day when one DNA-based species can sit down at a computer to design another. I plan to show that we understand the software of life by creating true artifi cial life. And in this way I want to discover whether a life decoded is truly a life understood [1]. – A Life Decoded By J. Craig Venter