Research Highlights


1. Genetic manipulation of Amycolatopsis mediterrarei S699 for production of Rifamycin analogs.

This project aims in genetic manipulation of A. mediterrarei S699 to produce rifamycin analogs which will be effective against multi drug resistant tuberculosisstrain. This project aims in genetic manipulations of polyketide synthase (PKS) gene cluster of Rifamycin B producing A. mediterranei S699. The genetic manipulation involves domain deletion, addition or swapping that will result in various rifamycin analogs which will be effective against infecting strains. For this purpose, firstly we developed cloning vectors and standardized the transformation system. Our efforts of several years of research led to the development of plasmid pRL1 by manipulating an indigenous plasmid pA387 (isolated from Amycolatopsis benzoylytica) by cloning its origin of replication in E.coli. Subsequently, series of cloning vectors, developed was patented (Lal, R. US Patent 005985560A; Tuteja et al., 2000) and used for standardization of transformation protocol in A. mediterranei. Secondly, we also cloned and characterize the rifamycin polyketide synthase (rifPKS) gene cluster from A.mediterranei (Kaur et al., 2001) and picked up entire rifPKS cluster on cosmid clones. This was followed by manipulation of A. mediterranei by combinatorial approach to develop proof of concept for the production of rifamycin analogues effective against MDR strains. We were successful in swapping acyltransferase domain of module 6 of rifPKS (rifAT6), that adds propionate unit with that of acyltransferase domain of module 2 of rapamycin PKS (rapAT2) from Streptomyces hygroscopicus. This successful genetic manipulation of AT6 domain swapping led to the generation of novel analog 24-desmethylrifamycin B. The resulting analog 24-desmethylrifamycin B produced was further converted to 24-desmethylrifamycin S & 24-desmethylrifamycin SV that were found to have a better antibacterial activity than rifamycin B. Before this technology is commercialized, an Indian Patent followed by PCT is being filed. For details, please visit the Tab "Patent".

Important Papers :

Nigam, A., Almabruk, K. H., Saxena, A., Yang, J., Mukherjee, U., Kaur, H., Kohli, P., Kumari, R., Singh, P., Zakharov, L. N., Singh, Y., Mahmud, T., and Lal, R. 2014. Modification of Rifamycin Polyketide Backbone Leads to Improved Drug Activity Against Rifampicin-Resistant Mycobacterium tuberculosis . J Biochem . doi: 10.1074/jbc.M114.572636.

Lal, R., Lal, S., Gründ E and Eichenlaub, R. Construction of a hybrid plasmid capable of replication in Amycolatopsis mediterranei . Appl. Environ. Microbiol. 1991, 57, 665-671.

Lal, R., Khanna, R., Dhingra, N., Khanna, M and Lal, S. 1998. Development of an improved cloning vector and transformation system in Amycolatopsis mediterranei (Nocardia mediterranei). J. Antibiot. 51, 161- 169.

Tuteja, D., Dua, M., Khanna, R., Dhingra, N., Khanna, M., Kaur, H., Saxena, D. and Lal, R. 2000. The importance of homologous recombination in the generation of large deletions in hybrid plasmids in Amycolatopsis mediterranei. Plasmid . 43, 1- 11.

Verma, M., Kaur, J., Kumar, M., Kumari, K., Saxena, A., Anand, S., Nigam, A., Ravi, V., Raghuvanshi, S., Khurana, P., Tyagi, A.K., Khurana, J.P., and Lal, R. 2011. Whole Genome Sequence of Rifamycin B Producing Amycolatopsis mediterranei S699. J. Bacteriol. 193, 5562-5563


2. Genomics

2.1 Genome of Rifamycin B ProducingAmycolatopsis mediterranei  S699 (Accession No. CP002896)

Whole genome sequencing of strain S699 was performed by using Roche 454 system (GS20 version) and Sanger shotgun sequencing. The hybrid assembly of Sanger-pyrosequencing data using Phrap resulted in 386 contigs. Subsequently, sequence gaps were filled by primer walking, transposon mutagenesis and complete sequencing of linker clones using Roche 454 system (GS FLX). The final assembly that consisted of 1,993,024 reads was done with MIRA3 Assembler by mapping to the reference genome of  A. mediterranei  U32 (GenBank accession number CP002000). The complete genome sequence of   A. mediterranei  S699 contains a single circular 10,236,779 bp chromosome with a GC content of 71.3 %. There are 9,575 CDSs falling into 6,883 functional COG groups. For details please see:

Important Papers

Verma, M., Kaur, J., Kumar, M., Kumari, K., Saxena, A., Anand, S., Nigam, A., Ravi, V., Raghuvanshi, S., Khurana, P., Tyagi, A.K., Khurana, J.P., and   Lal, R . (2011)   Whole Genome Sequence of Rifamycin B Producing  Amycolatopsis mediterranei  S699. J. Bacteriol.  193: 5562-5563.

2.2 Genome of hexachlorocyclohexane (HCH) degrading bacterium  Sphingobium indicum  B90A (Accession No. AJXQ00000000)

Sphingobium indicum  B90A was isolated from sugarcane rhizosphere soil in 1990 and became the first strain to degrade one of the most recalcitrant man-made compounds, the β- hexachlorocyclohexane (HCH). Since then B90A has been the focus of many studies leading to novel discoveries such as the association of   lin genes with IS   6100   , enantioselective transformation of chiral α-HCH by   linA1   and   linA2   gene, the presence of efficient HCH dehydrochlorinase and haloalkane dehalogenase systems, evolution of isomeric specific (α-, β-, γ-, δ- and ε- HCH) degradation pathways and potential application of B90A in HCH bioremediation. The draft genome of   S. indicum   B90A was generated by using Illumina Genome Analyzer platform. The sequencing data (88.3% of the total raw reads) was assembled into 149 contigs.

The availability of the genome sequence of   S. indicum   B90A coupled with community genomics data from the HCH dumpsite (R. Lal-unpublished) will act as an invaluable supplement to the ongoing research efforts towards understanding several unanswered questions associated with the degradation of HCH isomers and would thus aid in the development of   in situ ; bioremediation technology in the future. For details please see

Anand, S., Sangwan, N., Lata, P., Kaur, J., Dua, A., Singh, A., Verma, M., Kaur J., Khurana, P., Khurana, J.P., Mathur, S., and   Lal, R.   2012.   Genome Sequence of   Sphingobium indicum B90A, a Hexachlorocyclohexane (HCH) Degrading Bacterium. J. Bacteriol . 194:4472

2.3 Genome of Thermus sp. RLT( Accession No. AIJQ00000000)


Thermus sp. RL genome featured on the cover page of J.Bacteriology 194,3534 (cover page can be viewed at

The hot water springs located atop the Himalayan ranges in Parvati Valley at an altitude of 1,760 m in Manikaran, India, are known for their scenic beauty and spiritual sanctity. These are the hottest springs (90°C to 98°C) in the country and contain low levels of helium. The microbial diversity from these hot water springs has not been explored previously. Thermus sp. strain RL was isolated from a hot water spring, and its genome was sequenced. Metagenomic analysis is being carried out to understand microbial diversity at this site and to understand microbial life and biochemical processes at these high temperatures

Thermus sp. RL represents genome size of 20,36,600 bp with an average G+C content of 68.77%. Genome  annotations predicted 1986 protein coding genes and 710 hypothetical proteins. Strain RL has two rRNA operons (5S-16S-23S and 5S-5S-16S-23S ) and 47 tRNA genes . A total of 111 tandem repeats, 2825 CpG islands and a single CRISPR element.Further determination of the taxonomic position of strain RL, comparative genomics of Thermus spp. and metagenomic analyses of the hotspring are under way. We are also investigating the Taq DNA polymerase activity of this stain.For details please see

Dwivedi, V., Sangwan, N., Nigam, A., Garg, N., Niharika, N., Khurana, P., Khurana, J.P., and   Lal, R. Draft Genome Sequence of Thermus sp. RL Isolated from Hot Water Spring Located atop the Himalayan Ranges at Manikaran, India   . J. Bacteriol. 194:3534

2.4 Genome of Acinetobacter sp. HA (Accession No. AJXD00000000)

Acinetobacter sp. HA was isolated from the gut of the insect Helicoverpa armigera . It has a G+C content of 41% and the estimated genome size is 3.12Mb that has been assembled into 102 contigs. The bacterium possesses genes for natural competence as well as for esterase activity. The genome is being proceeded with for gap filling at the finishing stage by sequencing of a cosmid library (1156 clones in Super cos vector) that will validate the overall assembly. For details please see

Malhotra, J., Dua, A., Saxena, A., Sangwan, N., Mukherjee, U., Pandey, N., Raman, R., Khurana, P., Khurana, J,P and Lal, R. Genome sequence of Acinetobacter sp. HA isolated from the gut of polyphagous insect pest Helicoverpa armigera. J. Bacteriol. 194: 5156

More Genome Announcements:

Dua, A., Sangwan, N., Kaur, J., Saxena, A., Kohli, P., Gupta, A.K. and Lal, R . 2013. Draft Genome Sequence of Agrobacterium sp. Strain UHFBA-218, Isolated from Rhizosphere Soil of Crown Gall-Infected Cherry Rootstock Colt. Genome Announc . doi:10.1128/genomeA.00302-13.

Niharika, N., Sangwan, N., Ahmad, S., Singh, P., Khurana, J.P. and Lal, R. 2013. Draft genome sequence of Sphingobium chinhatense strain IP26 T isolated from the hexachlorocyclohexane dumpsite. Genome Announc . doi:10.1128/genomeA.00680-13.

Saxena, A., Nayyar, N., Sangwan, N., Kumari, R., Khurana, J.P., and Lal, R. 2013. Genome Sequence of Novosphingobium lindaniclasticum LE124 T , isolated from a Hexachlorocyclohexane (HCH) dumpsite. Genome Announc . doi:10.1128/genomeA.00715-13.

Singh, A. K., Sangwan, N., Sharma, A., Gupta, V., Khurana, J.P., and Lal, R . 2013. Draft Genome Sequence of Sphingobium quisquiliarum P25 T , a Novel Hexachlorocylohexane (HCH) Degrading Bacterium Isolated from the HCH Dumpsite. Genome Announc doi:10.1128/genomeA.00717-13.

Mukherjee, U., Kumar, R., Mahato, N.K.., Khurana, J.P., and Lal, R. 2013. Draft Genome Sequence of Sphingobium sp. HDIPO4, an Avid Degrader ofHexachlorocyclohexane. Genome Announc. doi:10.1128/genomeA.00749-13

Kaur, J., Verma, H., Tripathi, C., Khurana, J.P., and Lal, R. 2013. Draft Genome Sequence of a Hexachlorocyclohexane-DegradingBacterium, Sphingobium baderi Strain LL03 T . Genome Announc. doi:10.1128/genomeA.00751-13.

Kumar, R., Dwivedi, V., Negi, V., Khurana, J.P., and Lal, R. 2013. Draft Genome Sequence of Sphingobium lactosutens DS20Hexachlorocyclohexane (HCH) Dumpsite. Genome Announc. doi:10.1128/genomeA.00753-13.

Mahato NK., Tripathi C., Verma H., Singh N., Lal R. 2014. Draft genome sequence of Deinococcus sp. strain RL isolated from sediments of hot Spring located at Manikaran, India. Genome Announc. doi:10.1128/genomeA.00703-14.

Mukherjee U., Saxena A., Kumari R., Singh P., Lal R. 2014. Draft genome sequence of Amycolatopsis mediterranei DSM 40773, a tangible antibiotic producer. Genome Announc. doi: 10.1128/genomeA.00752-14.

Saxena A., Kumari R., Mukherjee U., Singh P., Lal R . 2014. Draft Genome Sequence of Rifamycin producer Amycolatopsis rifamycinica 46095. Genome Announc. doi:10.1128/genomeA.00662-14.

Negi V., Lata Pushp, Sangwan N., Gupta S., Das S., Rao D.L.N., and Lal R . Draft genome sequence of Hexachlorocyclohexane (HCH)- degrading Sphingobium lucknowense Strain F2, isolated from the HCH Dumpsite. Genome Announc. doi:10.1128/genomeA.00788-14.

Singh P., Kumari R., Mukherjee U., Saxena A., Sood U., Lal R. Draft genome sequence of Rifamycin derivatives producing Amycolatopsis mediterranei strain DSM 46096/S955. Genome Announc. doi:10.1128/genomeA.00837-14

Sharma A, Hira P, Shakarad M, Lal R . 2014. Draft genome sequence of Cellulosimicrobium sp. MM, isolated from arsenic rich microbial mats of a Himalayan Hot Spring. Genome Announc . 5: e01020-14


3. HCH Bioremediation

Another major area his group is working on, is the molecular genetics of hexachlorocyclohexane (HCH) degradation by sphingomonads that could eventually lead to the development of a technology for bioremediation of HCH contaminated soils. He has extensively studied the degradation of HCH by Sphingobium indicum (formerly known as Sphingomonas paucimobilis B90A). His group has unravelled the physiological and genetic aspects of HCH degradation in B90A and has provided evidence for the stability horizontal transfer of ' lin ' genes (responsible for HCH degradation) among different bacteria. The study has also led to the cloning of genes responsible for the mineralization of α- and γ- isomers of HCH in B90A and dynamics of ' lin ' gene expression. The group is working on deciphering the degradation pathway (hitherto not known) of α- δ- β and ε- HCH. Alongside, his group is attempting bioremediation of HCH contaminated sites by 'bioaugmentation', 'biostimulation' and 'enzymatic bioremediation' approaches.

Important Papers

Pandey R., Lucent D., Kumari K., Sharma P., Lal R ., Oakeshott J. G., and Pandey G. 2014. Kinetic and sequence-structure-function analysis of LinB enzyme variants with β- and δ-hexachlorocyclohexane. PLoS One. DOI: 10.1371/journal.pone.0103632

Nayyar N., Sangwan N., Kohli P., Verma H., Kumar R., Negi V., Oldach P., Mahato NK., Gupta V., Lal R . 2014. Hexachlorocyclohexane: Persistence, Toxicity and Decontamination. Rev Environ Health . 29 : 49-52

Sharma , P., Jindal , S., Bala , K., Kumari, K., Niharika, N., Kaur ,J., Pandey , G., Pandey , R., Russell , R. J., Oakeshott , J. G. and Lal, R. 2013. Functional screening of enzymes and bacteria for the dechlorination of hexachlorocyclohexane by a high-throughput colorimetric assay. Biodegradation. 48 : 65–79.


4. Metagenomics of Extreme Environments supplemented with Comparative Genomics

Metagenomic analysis of hexachlorocyclohexane contaminated environments (soil, sediment and water) has been done. We have been able to resolve the taxonomical and functional dynamics of enriched, yet un-cultivated microbial communities using amplicon based and shotgun sequencing data (3.2 Gbp). Using paired end assembled and validated downstream analysis we have reconstructed various genotypes (near complete plasmids and draft genomes) corresponding to their pangenomic abundance. Our future projects (in the pipeline) are targeting the community genomics of hot spring at Manikaran, Himachal Pradesh, India. For details please see

Important Papers:

Sangwan, N., Lata, P., Dwivedi, V., Singh, A., Niharika, N., Kaur, J., Anand, S., Malhotra, J., Jindal, S., Nigam, A., Lal, D., Dua, A., Saxena, A., Garg, N., Verma, M., Kaur, J., Mukherjee, U., Gilbert, J.A., Dowd, S.E., Raman, R., Khurana, P., Khurana, J.P., and Lal, R. 2012.Comparative Metagenomic analysis of soil microbial communities across three Hexachlorocyclohexane contamination levels. PLos One : 7, e46219

Sangwan, N., Verma, H., Kumar, R., Negi, V., Lax, S., Khurana,P., Khurana, J.P., Gilbert, J. A. and Lal,R. 2013. Reconstructing an ancestral genotype of two hexachlorocyclohexane degrading Sphingobium species using metagenomic sequence data. ISME J. doi:10.1038/ismej.2013.153

Verma H., Kumar R., Oldach P., Sangwan N., Khurana J.P., Gilbert J.A., Lal R . 2014. Comparative genomic analysis of nine Sphingobium strains: Insights into their Evolution and Hexachlorocyclohexane (HCH) Degradation Pathway. BMC Genomics .15 :1014.

Sharma A., Sangwan N., Negi V., Kohli P., Rao D.L.N., Lal R . 2015. Pan-genome dynamics of Pseudomonas gene complements across hexachlorocyclohexane dumpsite. BMC Genomics . 16:313.

Sangwan N., Lambert C., Sharma A., Gupta V., Khurana P., Khurana J.P., Sockett R.E., Gilbert J.A., Lal R . Arsenic rich Himalayan hot spring metagenomics reveal genetically novel predator-prey genotypes. Environ. Microbiol. Reports DOI: 10.1111/1758-2229.12297.;jsessionid=A06D0DFD46F2434D496BFC3F85CFB92E.f01t01


5. Microbial Diversity

In our ongoing effort to identify the culturable from exteme environment especially the hexachlorocyclohexane (HCH) dump site, Lucknow, Uttar Pradesh in North India, we have characterized 30 novel species using polyphasic approach . The species belong to different generar like Sphingomonas, Sphingobium , Novosphingobium , Sphingopyxis , Rhizobium, Devosia , Pontibacter etc. and are described in International Journal of Systemic and Evolutionary Microbiology .

Important Papers

Malhotra, J., Anand, S., Jindal, S and Lal, R . 2012. Acinetobacter indicus sp. nov., isolated from Hexachlorocyclohexane (HCH) dumpsite. Int. J. Syst. Evol. Microbiol . 62 : 2883-2890.

Kaur, J., Kaur, J., Niharika, N and Lal, R . 2012. Sphingomonas laterariae LNB2 T sp. nov. isolated from hexachlorocyclohexane (HCH) contaminated dumpsite in Lucknow. Int. J. Syst. Evol. Microbiol. 62 :2891-2896 .

Niharika, N., Jindal, S., Kaur, J and Lal, R . 2012. Sphingominas indica sp. nov., isolated from hexachlorocyclohexane (HCH) contaminated soil. Int. J. Syst. Evol. Microbiol. 62 : 2997-3002.

Kaur, J., Moskalikova, H., Niharika, N., Sedlackova, M., Hampl, A., Damborsky, J., Prokop, Z and Lal, R . 2012. Sphingobium baderi sp. nov., isolated from a hexachlorocyclohexane (HCH) dumpsite in Spolana. Int. J. Syst. Evol. Microbiol . 63 : 673-678.

Niharika, N., Moskalikova, H., Kaur, J., Sedlackova, M., Hampl, A., Damborsky, J., Prokop, Z and Lal, R . 2012. Novosphingobium barchaimii sp. nov., isolated from a hexachlorocyclohexane (HCH) contaminated soil. Int. J. Syst. Evol. Microbiol . 63 : 667-672.

Jindal, S., Dua, A. and Lal, R . 2012. Sphingopyxis indica sp. nov., isolated from a high dose point hexachlorocyclohexane (HCH) contaminated dumpsite. Int. J. Syst. Evol. Microbiol. 63: 2186-2191.

Garg, N., Bala, K., and Lal, R . 2011. Sphingobium lucknowense sp. nov., a hexachlorocyclohexane (HCH) degrading bacterium isolated from HCH contaminated soil. Int. J. Syst. Evol. Microbiol . 62 : 618-623.

Anand, S., Bala, K., Saxena, A., and Lal, R . 2011. Microbacterium amylolyticum sp. nov ., N5T a bacterium isolated from industrial waste soil. Int. J. Syst. Evol. Microbiol. 62 : 2114-2120.