ANTIBACTERIAL SCREENING OF CERTAIN TRADITIONALLY USED INDIAN MEDICINAL PLANTS AT

microbial pathogens. In the present study, we have selected 10 medicinal plants to be screened against human pathogenic bacteria.

Ethanolic and aqueous extracts of 10 Indian medicinal plants traditionally used in medicine at Vidisha district were studied for their antimicrobial activity against human pathogenic bacteria of clinical origin. Of these, 09 ethanolic extracts and 08 aqueous extracts of medicinal plant showed varied levels of antibacterial activity against one or more tested human pathogenic bacterial strains. Overall, broadspectrum antibacterial activity was observed in 05 Indian medicinal plants (

…………………………………………………………………………………………………….... Introduction:-
The discovery, development, and clinical use of antibiotics during the 20 th century have decreased substantially the morbidity and mortality from bacterial infections. The antibiotic era began with the therapeutic application of sulfonamide drugs in the 1930s, followed by a golden period of discovery from approximately 1945 to 1970, when a number of structurally diverse, highly effective agents were discovered and developed. However, since the 1980s the introduction of new agents for clinical use has declined, reflecting both the challenge of identifying new drug classes and a declining commitment to antibacterial drug discovery by the pharmaceutical industry. The same period with a reduced rate of introduction of new agents has been accompanied by an alarming increase in bacterial resistance to existing agents, resulting in the emergence of a serious threat to global public health. The resistance problem demands that a renewed effort be made to seek antibacterial agents effective against pathogenic bacteria resistant to current antibiotics (Chopra et al., 1997).
Today, the situation is alarming in developing as well as developed countries due to indiscriminate use of antibiotics. However, rapid emergence of antimicrobial resistance among pathogenic microorganisms has led to a renewed search for new antimicrobial agents. Severe infections caused by bacteria that are resistant to commonly used antibiotics have become a major global healthcare problem in the 21 st century (Alanis, 2005). Infectious diseases are the world's leading cause of premature deaths, killing almost 60,000 people per day despite remarkable advances in Medical research and treatment during the 20 th century, infectious diseases remain among the leading cause of death worldwide. Of these, nosocomial infections comprise about 5 to 10% (Culver et al., 1985). It has been estimated that one third of all nosocomial infections may be preventable and are frequently caused by organisms acquired within the hospital environment (Hughes, 1988). In recent years, drug resistance to human pathogenic bacteria has been commonly reported from all over the world ( Infectious disease caused by bacteria, viruses, fungi and parasites are still a major threat to public health, despite the tremendous progress in human medicine (Cosa et al. 2006). Such situation stimulates the development of new antimicrobial agents in order to treat the infectious disease in an effective manner. So this matter continued to an era to identify the potential antimicrobial agent from the natural resources. The edible plants that used for traditional medicine contain a wide range of substance that can be used to treat abundant of infectious disease with reduced side effects ( In the present scenario of emergence of multiple drug resistance to human pathogenic organisms, this has necessitated a search for new antimicrobial substances from other sources including plants. Therefore, it is imperative to search for new, efficacious and safe antibiotics from natural sources to combat the menace of drugresistant infections. It is expected that plant extracts showing target sites other than those used by antibiotics will be active against drug-resistant microbial pathogens. In the present study, we have selected 10 Indian medicinal plants to be screened against human pathogenic bacteria.

Plant Material
Ten authenticated plants namely Beta vulgaris L., Coriandrum sativum L., Emblica officinalis Gaerth., Eucalyptus sp., Lantana camara L., Lawsonia inermis L., Mentha arvensis L., Ocimum sanctum L., Syzgium aromaticum L. and Terminalia indica L. were collected different areas of Vidisha district. The taxonomic identities of these plants were confirmed by relevant data and were further identified by a senior taxonomist Dr. S. K. Jain, Dept. of Botany, S. S. L. Jain P. G. College, Vidisha and voucher specimens have been deposited in the Department of Botany, St. Mary's P. G. College, Vidisha. The details of medicinal plants along with their specimen code number are listed in Table 1.

Preparation of Plant Extracts
Plant extracts were prepared by the method of Harborne (1984). Briefly, 100 grams of each powered plant sample was extracted with 100 ml of ethanol and distilled water for ethanolic and aqueous extracts respectively using the Soxhlet apparatus. After extraction, an excess was evaporated under reduced pressure in vacuum evaporator. The dried crude extracts were sterilized overnight by UV radiation and then stored at 4ºC in amber color glass vials until further use.

Preparation of Plants Derived Antibiotic Discs
Both the crude extracts (ethanolic and aqueous) each 100 mg, were dissolved in 1 ml of dimethyle sulphooxide (DMSO) and were filtered by using membrane (pore size 0.47 μm). The discs of 6 mm diameter (Sterile blank, HiMedia, Bombay, India) were impregnated into the concentration of the each extracts. The final impregnated discs used for the sensitivity test were 100 mg disc -1 . These impregnated discs were dried in incubator at 37 ºC for 18 -24 hours and after this stored in an amber colour glass bottle at room temperature until further use.

Microorganisms Used
The test organisms were used Staphylococcus aureus, Bacillus subtilis, Escherichia coli, Salmonella typhi and Streptococcus pneumoniae. The test bacterial strains were collected and isolated from the Department of Microbiology, Gandhi Medical College, Bhopal, M P., India All the clinical isolates were biochemically and serologically characterized by standard method (MacFaddin, 1985).

Antibiotics Resistance of test strains
The antibiotic sensitivity of all the test strains was determined by the standard disk diffusion method of Bauer et al., 1966 against a number of antibiotics. The potency of antibiotics discs are: Ciprofloxacin (05 µg disc -1 each), Gentamycin and Streptomycin (10 µg disc -1 each), Erythromycin, Chloramphenicol and Vancomycin (30 µg disc -1 each). All antibiotic discs were purchased from the Hi-Media, Bombay, India. The details of antibiotics resistance of test strains are listed in Table 2.

Antimicrobial Assay of Plants Derived Antibiotic Discs
For this, the standard disk diffusion method of Bauer et al., 1966 was used. 0.1 ml of diluted inoculum (10 5 CFU / ml) of test organisms was spread on Muller-Hinton agar plates. The plant derived antibiotics discs were placed on the agar plates. DMSO was used as a control. The plates were incubated at 37ºC for 18 -24 hours. The antibacterial activity was evaluated by measuring the zone of inhibition (ZOI) in mm against the test organisms.

Results And Discussion:-
Emergence of multidrug resistance in human and animal pathogenic bacteria as well as undesirable side effects of certain antibiotics has triggered immense interest in the search for new antibiotics or antimicrobial drugs of plant origin. In the present study ethanolic and aqueous extracts of 10 traditionally used Indian medicinal plants have been tested against 05 human pathogenic bacterial strains.
Ethno-botanical data, plant parts used along with their voucher specimen number are given in Table 1. The bacterial susceptibilities to the tested antibiotics are shown in Table 2. Most of the antibiotics were inhibited the growth of at least two bacterial strains accept chloramphenicol that inhibited four bacterial strains. The antibacterial screening of the extracts and their potency was quantitatively assessed by the presence or absence of inhibition zone and zone diameter as given in Table 3. The alcoholic and aqueous extracts were tested, as alcohol was found to be a better solvent for extraction of anti-bacterially active substances compared to water (Ahmad et al., 1998). The results of screening are encouraging as out of the 10 plants 09 ethanolic and 08 aqueous extracts showed antibacterial activity against one or more test bacteria. In the case of test bacteria, the basis for their differences in susceptibility might be due to the differences in the cell wall composition of Gram + ve and Gram -ve bacteria (Grosvenor et al., 1995). B. subtilis was least sensitive compared to other test bacteria, which may be due to their ability to form highly resistant resting stages called endospores. Drug-resistant strains of bacteria were found to be sensitive to the tested plant extracts. This has clearly indicated that antibiotic resistance does not interfere with the antibacterial action of plant extracts and these extracts might have different modes of action on test organisms.
Major factor limiting the long-term use of antibiotic agents is resistance. Before antibiotics era, many people died of bacterial infections caused by pathogens as Staphylococcus aureus, Streptococcus pyogenes and Streptococcus pneumonia. Use, abuse or misuse of antimicrobial agents has encouraged the evolution of bacteria towards resistance that often results in therapeutic failure (Straut et al., 1995). Prescribing practice of specific class of antibiotics to certain organisms has been found to play a critical role in development of resistance against that antibiotic (Metz-Gercek et al., 2009; Costelloe et al., 2010). Thus, antibacterial resistance findings and understanding are necessary to help minimize the emergence of multi drug resistant organisms by promoting prudent use of antibiotics, for this purpose, the need for general public to be appropriately informed on use of antibiotics has been emphasized (Euro surveillance editorial team, 2010).