Faculty Recognitions & Awards
Delhi U creates Kit to diagnose Sexually Transmitted diseases
-Prof. Daman Saluja
Sexually transmitted diseases are emerging as a major threat to reproductive health of men and women at large. Several Sexually transmitted diseases (STDs) are caused by bacteria that can grow and multiply easily in the warm, moist areas of the reproductive tract, including the cervix, uterus, and fallopian tubes in women, and in the urethra in women and men. The most common among these are Chlamydia & Neisseria. The usual method of detection is Gram-staining followed by confirmation using methods like antigen detection or biochemical assay. Both methods are highly unsatisfactory, especially in asymptomatic patient (where the infection load is low) which largely contributes to the persistence and transmission of disease to the sexual partner. Unavailability of a reliable and cost effective test for detection of the infection is a growing concern especially in developing countries.
Prof Daman Saluja and her group at Dr B R Ambedkar Center for Biomedical Research, University of Delhi have designed and established an in-house PCR assay for detection of Chlamydia & Neisseria. The performance of the prototype kit was evaluated against currently used diagnostic methods and has been found to be cost-effective, highly specific, sensitive and user friendly. The research was funded by Department of Biotechnology, Government of India. The prototype kit has been patented in India and is under process for patent in USA. University of Delhi and Department of Biotechnology, Government of India has signed a Memorandum of Agreement on 6th November 2012 for transfer of technology on ‘Development of kit for detection of Neisseria and Chlamydia’ to DSS Tech Pvt Ltd. The industry is expected to bring it into the market within the next two years. This is the first technology transferred jointly by Dr. B.R. Ambedkar Center, University of Delhi and Department of Biotechnology. The University has also received a signing amount of Rs. 5 Lakh as the first installment out of Rs. 15 Lakh and royalty offered by the Industry. The newly developed prototype kit for detection of Neisseria & Chlamydia is PCR-based, which has internal and positive control in order to confirm the success of the reaction. Detection is done in dark reader using fluorescent-labelled probe, which successfully reduces the time of detection.
Most of the tests currently available in the market range from US$ 250 to US$ 1,800 for 100 tests. The new product offers a highly inexpensive alternative to these tests in developing countries. The test does not involve the laborious microscopic examination required in many other diagnostic kits available in the market, thus enabling its operation by relatively unskilled staff. The other benefits include a non-invasive swab collection method for the sample which is safe for the patient, and low capital investment as no special equipment is used other than a PCR machine and a dark reader for the diagnostic assay. The method is highly sensitivity, specific and is eco friendly as it doesn’t involve any carcinogenic or non-biodegradable chemical. Most importantly, the results are available in less than 4 hours. The diagnostic test is easy to handle. Keeping in mind the cost and facilities in peripheral clinical laboratories, the assay reagents can be stored under refrigeration for more than 6 months. Thus, the kits can be easily transported to remote areas.
Bacterial Life in Hot Water Springs atop the Himalayan Ranges
- Prof. Rup Lal
Hot water springs harbour unique microbial diversity, which is naturally selected to survive under high temperature conditions. Ever since the discovery of first reference bacterial strain, Thermus aquaticus, from the hot springs in Yellow Stone National Park, USA, several bacterial strains have been isolated from such hot water springs around the world. These bacteria are of special biotechnological interest as they produce thermostable enzymes and have been used for eradicating heavy metal and organic pollutants in ground water and clearing clogged wells. For example, the Taq DNA polymerase enzyme (for DNA amplification) was isolated from Thermus aquaticus and this enzyme has become a household word in molecular biology. The market for this enzyme is growing into hundreds of millions of dollars every year. While in India there are about 303 thermal springs located in different states, the major ones gathering attention are in Yamunotri, Gangnani, Kedarnath and Rudraprayag in Uttrakhand, Bakreshwar and Bhuri in West Bengal, Tantloi in Jharkand, Gangtok in Sikkim, and Manikaran in Himachal Pradesh. However, these hot water springs have not been explored for bacterial diversity and biotechnological applications. We initiated a project to explore bacterial diversity from one of the hot springs located in the Himalayan ranges at an altitude of 1760 m in Manikaran (Figure 1) under the DU/DST Purse (Promotion of University Research and Scientific excellence ) Program.
Figure 1: Hot water spring alongside the Parvati River in Manikaran; Water gushes out (arrow) from the rocks underneath with a lot of pressure. These are the hottest springs (90°C to 98°C) in the country and contain low levels of helium from where the bacterium Thermus sp. RL was isolated. We have now performed random sampling in and around the hot water springs and isolated DNA from these samples. The DNA samples are being sequenced to understand the microbial diversity existing at this high temperature by using culture independent molecular approaches.
The hot water springs in Manikaran are the hottest (90°C to 98°C) in India and the place is popular for its scenic beauty and spiritual sanctity. The water in these hot springs contains low levels of helium and calcium carbonate, the hard crust of which can be seen sticking to the rocks or at the bottom of the springs.
As a first step, we isolated a bacterium Thermus sp. RL (Figure 2) from one of the springs. While taxonomical characterization of this bacterium is still underway, this bacterium was found to grow at a temperature range of 80 to 90 °C. Thermus sp. RL also showed better protease activity (proteases are responsible for protein digestion) that was qualitatively determined by zone of clearance assay on milk-casein plates (at 70oC) and compared with its neighbouring strain Thermus thermophilus HB8, isolated from a hot spring located in Japan (Figure 2(II)).
Figure 2: (I); Transmission electron microscopy of Thermus sp. RL. (II); Zone of clearance due to protease activity of Thermus grown on a medium containing milk casein; (A) Thermus sp. RL T and (B) Thermus thermophilus HB8T (Larger zone of clearance as in the case of Thermus sp RL means more protease production) .
In order to understand the mechanism or biogeochemical processes that enable this organism to survive under such extreme temperatures, we have sequenced the genome of Thermus sp. RL that was published in July issue of Journal of Bacteriology (A publication of American Society for Microbiology) (Dwivedi, V., Sangwan, N., Nigam, A., Garg, N., Niharika, N., Khurana, P., Khurana, J,P., and Lal, R. J. Bacteriol. 194, 3534, 2012). The site from where the bacterium has been isolated was featured on the cover page of the Journal of Bacteriology http://jb.asm.org/content/194/13.cover-expansion. The draft genome sequence revealed a genome size of 20,36,600 bp with an average GC content of 68.77%. Genome annotations predicted 1,986 protein-coding genes and 710 hypothetical proteins. Strain RL has two rRNA operons and 47 tRNA genes. A total of 111 tandem repeats, 2,825 CpG islands, and a single clustered regularly interspaced short palindromic repeat (CRISPR) element. Interestingly, Taq DNA polymerase gene sequence showed 98% similarity with its neighbouring strains, suggesting further experimentations for exploitation of this enzyme in DNA amplification. These studies and data have now set the stage to carry out metagenomic analysis (culture independent approach) to understand the bacterial life and biochemical processes at higher temperatures apart from exploitation of Thermus sp. RL for thermo-stable enzymes.