International Research of Pharmacy and Pharmacology (ISSN 2251-0176) Vol. 1(9) pp.

228-236, December 2011 Available online Copyright © 2011 International Research Journals

Full Length Research Paper

Antimicrobial activity of lemongrass (Cymbopogon citratus) oil against microbes of environmental, clinical and food origin
Bhoj Raj Singh1*, Vidya Singh2, Raj Karan Singh2 and N. Ebibeni1

ICAR Research Complex for NEH Region, Jharnapani, Nagaland, India 2 NRC on Mithun, Jharnapani, Nagaland, India
Accepted 30 November, 2011

Out of the 1114 strains belonging to 29 genera and 105 species of microbes (molds, yeasts and bacteria) isolated from different sources [clinical cases, environment (water, air, soil, droppings of lizards and birds), food and healthy animals], 38.2% were sensitive to lemongrass oil discs containing 50 µg oil/disc. All molds, yeasts, Lactobacillus acidophilus, Morganella morganii, most of the Bacillus spp. strains (84.3%), aeromonads (78%), Edwardsiella spp. (73.9%), 53.6% pseudomonads, 53.1% streptococci and 50% of Budvicia aquatica and Leminorella ghirmontii strains were sensitive to lemongrass oil (LGO). On the other hand, all Hafnea alvei, Laclercia adecarboxylata, Xenorhabdus luminescens and majority of Salmonella enterica (98.3%), Citrobacter spp. (93.7%), Providencia spp. and Kluyvera cryocrescens (83.3%), Enterobacter spp. (78.2%), Proteus spp. (78%), Escherichia spp. (77.7%), enterococci (73.7%), Serratia spp. (75%) and Erwinia ananas (75%), Pragia fontium (70.6%), staphylococci (69.8%) and Klebsiella spp. (62.7%) strains were resistant to LGO. MIC of LGO for sensitive strains (tested against discs containing 50 µg LGO) varied from 1 µg to 32 µg /ml while none of the resistant strains had MIC <64 µg LGO/ ml. MIC for yeast strains was the least i.e., 1 µg LGO/ ml. LGO had microbicidal activity on E. coli, S. aureus and Candida albicans. LGO instantly killed C. albicans and E. coli, and S. aureus in 10 min at 1 mg/ ml concentration, indicating of its wide spectrum antimicrobial activity at easily achievable concentrations. Study also indicated that LGO is more effective on enterococci in aerobic instead of microaerophilic growth conditions, it is indicative that in-vivo sensitivity results may differ from in-vitro tests. Keywords: Lemongrass oil, Antimicrobial activity, Microbes INTRODUCTION Of more than 400,000 spp. of tropical flowering plants, varieties of several thousands species have been used for their medicinal properties in traditional medicine (AliShtayeh and Abu Ghdeib, 1999; Odugbemi, 2006). Lemongrass (Cymbopogon citratus), a tall perennial grass comprising of about 55 species, is native to warm region and grows in almost all tropical and subtropical countries (Cheel et al., 2005). The biologically active constituent of lemon grass is citral constituting more than 75% (w/w) of its essential oil (Huynh et al., 2008). Lemongrass is widely used as an essential ingredient in Asian cuisines because of its sharp lemon flavour. Herbal tea of lemongrass is used as sedatives, febrifuge and immunostimulant in India (Pearson, 2010; Brian and Ikhlas, 2002) while, lemongrass essential oil
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is applied for its medicinal value to cure acne, oily skin, scabies, flatulence, headaches, blood circulation problems (Pearson, 2010) and excessive perspiration due to its antimicrobial and antibacterial activities (Lawless, 1995). It has also been used as carminative, stimulant, emmenagogue, diuretic and antiseptic (Ghani et al., 1997). In Nigeria, lemon grass is used for stomach problem and it is also used in combination with few other plants for effective treatment of malaria (Aibinu et al., 2007) and typhoid (Depken, 2011). Although, in a few preliminary antimicrobial screenings, LGO had shown no activity against four Gram positive (Bacillus subtilis, Corynebacterium diphtheriae, Streptococcus pyogenes and Staphylococcus aureus) and three Gram negative (Salmonella paratyphi A, Escherichia coli and Pseudomonas aeruginosa) bacterial cultures (Saify et al., 2000), later on several studies have shown that the

.3 83. 1991. (3) Staphylococcus spp.0 50.3 46. LGO’s antimicrobial properties make it an effective drug for bacterial and fungal infections.0 100. soil. Abu-Seif. 2000. Bacterial. 1999) and fungi (Pratt and Hudson.3 30. birds) and maintained at Microbiology Laboratory. water. 1994.1 50.7 0. fish. Nagaland Centre.0 78.0 84.0 26.0 0. It can be used in cleaning wounds and treatment of skin diseases such as ringworm. flavus..Singh et al. Besides.6 53.7 22. (3) Hafnea alvei Leclercia adecarboxylata Xenorhabdus luminescens lemon grass has antibacterial and antifungal properties (Ushimaru et al. 2010). India.8 16.0 0. Syed et al.9 66. cattle.. 2007). 2.9 50.0 0.0 % resistant strains 0. six A.7 77. Saikia et al. MATERIALS AND METHODS Lemongrass oil (LGO) Light yellow colored pure lemongrass oil was obtained as free gift from Naga Fragrance Pvt.0 100. et al. (1) Bacillus spp. 2003. pig. (4) Proteus spp..0 100. 1989.. (8) Budvicia aquatica Leminorella ghirmontii Klebsiella spp. (15) Aeromonas spp. ICAR Research Complex for NEH Region.0 100.4 25.0 100.0 26. lizards.. Nagaland. Jharnapani. Onawunmi. (3) Salmonella enterica spp. 229 Table 1. staphylococcal infections. Antimicrobial effect of lemongrass oil on strains of different genera of microbes Microbial strains (Number of species) tested Strains tested 11 7 1 3 3 115 91 23 3 28 32 8 2 110 43 17 12 12 213 112 41 55 6 6 95 59 4 1 1 Strain resistant 0 0 0 0 0 18 20 6 1 13 15 4 1 69 30 12 9 9 157 87 32 43 5 5 89 58 4 1 1 Strains sensitive 11 7 1 3 3 97 71 17 2 15 17 4 1 41 13 5 3 3 56 25 9 12 1 1 6 1 0 0 0 % sensitive strains 100. Lactobacillus acidophilus Morganella morganii Penicillium spp.7 53. (8) Edwardsiella spp.0 Aspergillus spp.3 22.7 69. ponds. air. Alam et al.4 46.0 0.7 6. 1995.8 70. Fungal and bacterial strains Five Aspergillus niger. 2009). (2) Micrococcus agilis Pseudomonas spp.0 25. 1993.2 83. 1996) and malaria. (5) Pragia fontium Ervinia ananas Serratia spp.7 16.0 75..0 62. 3) isolated (from clinical cases. the present study was undertaken to elucidate the antimicrobial spectrum of LGO through testing it against 21 isolates of three genera of fungi and 1085 isolates of common bacteria belonging to 26 genera.6 75.0 73.. Therefore. (4) Enterobacter spp.0 21.7 78.3 1.0 15. only little is known about its action on field strains of clinical. (2) Citrobacter spp. Sharma et al. three Penicillium spp. yeast and . India.3 22. (5) Enterococcus spp.1 33. (15) Escherichia spp.2 29.0 37.0 0.7 98. the Plasmodium spp (Pearson. Nagaland. environmental and food origin.0 73. molds and yeasts. (2) Candida spp. Although many studies have proved antimicrobial effect of LGO using reference strains of variety of bacteria (Chao and Young.3 78. LGO has been reported to effectively control growth of agent of American foulbrood disease (AFD). bacteria.3 100. and urinary tract. Nieto et al. It can also be used in food poisoning.0 50.0 100. (9) Kluyvera cryocrescens Providencia spp.. stomach. (3) Streptococcus spp. Dimapur.3 93. seven Candida albicans strains and 1093 bacterial strains of 26 genera (Tables 1. the Paenibacillus larvae (Alippi et al. were revived and checked for purity. and other common infections of the colon.0 0. Ltd.

0 25. Pharm.3 83.0 66.6 16.0 77.0 0.0 100.0 60.0 100.0 100.2 0.0 7.0 0.4 83.0 100.0 0. salmonicida A.0 100.3 39.7 0.0 0.0 0. pneumoniae Klebsiella terrigena Kluyvera cryocrescens Leclercia adecarboxylata Leminorella ghirmontii Morganella morganii Proteus mirabilis Proteus myxofaciens Proteus penneri Proteus vulgaris Pragia fontium Providencia heimbachae Providencia rettgeri Pseudomonas aeruginosa Pseudomonas fluorescens Pseudomonas spp Salmonella enterica ssp.0 100.8 60.7 0.1 0.0 100.0 100.5 22.0 0.0 0. houtenae Salmonella enterica ssp.3 100. Pharmacol Table 2.0 100.0 50.7 60.0 100.0 0.0 0.0 0.0 0.8 100.9 50. veronii Budvicia aquatica Citrobacter amalonaticus C.6 0.8 70.7 0.3 19.0 100.3 100.0 0.0 7.9 100.3 100.0 0.1 50. J.0 92.0 0.0 100. hydrophila A.8 50. Antimicrobial effect of lemongrass oil on strains of Gram negative bacteria Microbial strains tested (Number species) Aeromonas caviae A. salamae Serratia fonticola Serratia marcescens Serratia odorifera Serratia plymuthica Serratia rubidiae Xenorhabdus luminescens of Strains tested 12 18 18 9 3 5 1 8 3 14 8 11 6 78 1 22 23 9 3 1 5 11 1 1 1 12 6 96 8 2 4 9 95 6 6 1 2 3 12 1 19 9 17 1 5 2 1 25 3 45 11 1 2 5 1 3 1 Strain resistant 2 10 3 0 2 2 0 0 0 1 4 11 6 72 1 5 14 9 1 1 5 11 1 1 0 9 4 77 4 2 4 7 57 5 5 1 1 0 8 0 17 7 12 0 5 2 1 10 3 44 11 1 2 5 1 0 1 Strains sensitive 10 8 15 9 1 3 1 8 3 13 4 0 0 6 0 17 9 0 2 0 0 0 0 0 1 3 2 19 4 0 0 2 38 1 1 0 1 3 4 1 2 2 5 1 0 0 0 15 0 1 0 0 0 0 0 3 0 % sensitive strains 83.7 0.0 33.3 100.0 66.7 16. diversus C.0 100.0 0.0 16.7 80. salmonicida ssp.0 83. indica Salmonella enterica ssp.3 44.0 100.0 0.3 60. Erwinia ananas Escherichia blattae Escherichia coli Escherichia furgusonii Escherichia vulneris Hafnea alvei Klebsiella oxytoca K.0 100.0 75.0 40. freundii Edwardsiella hoshiniae Edwardsiella.0 33.0 100.0 92. schubertii A. eucranophila A. tarda Enterobacter agglomerans Enterobacter.7 0.3 100.0 100.0 77. smithia A.230 Int.0 2.2 50.0 66.0 89. amnigenus I Enterobacter amnigenus II Enterobacter cancerogenus Enterobacter cloacae Enterobacter gregoviae Enterobacter hormaechei Enterobacter sakazaki Enterobacter spp.0 % resistant strains 16.0 22. salmonicida ssp.0 0.0 97.0 0.7 55.0 .0 66.0 10.0 100.0 100.5 77. achromogenes A.0 100.4 100.0 50.0 0.0 33. pnumoniae ssp. salmonicida ssp.2 29.7 40.0 100.0 100.0 100. Res. sobria A. media A.2 40.0 0.0 0.0 22.0 0.0 0.0 33.

0 100.0 33.9 66.0 25.0 0.0 100.0 0.0 100.8 100.1 100.6 100.0 0.0 100.7 50.3 0.Singh et al.2 65.3 78.0 100.0 100.4 100.0 100.0 0.3 50.9 0.0 53.0 66.0 100.1 33.0 42.3 50.0 0.0 61.0 100.0 100.0 0.0 57.0 0.7 100. 231 Table 3.6 0.0 0.0 75.8 21.0 19.0 38.0 0.5 100.0 33.0 0.0 100.3 Aspergillus flavus Aspergillus niger Bacillus anthracoides Bacillus badius Bacillus brevis Bacillus circulans Bacillus coaggulans Bacillus laterosporus Bacillus lentus Bacillus licheniformis Bacillus marcerans Bacillus mycoides Bacillus pentothenticus Bacillus stearothermophilus I Bacillus stearothermophilus II Bacillus subtilis Bacillus spp.0 100.0 93.7 % resistant strains 0.5 0.0 6.3 0.8 34. Antimicrobial effect of lemongrass oil on strains of Gram positive bacteria and fungi Microbial strains (Number of species) tested Strains tested 6 5 3 7 4 4 51 1 8 6 4 2 16 1 4 3 1 7 1 13 32 32 29 2 13 11 16 42 3 7 5 1 6 1 3 3 13 2 23 2 3 2 3 1 1 1 21 3 Strain resistant 0 0 0 0 1 0 10 0 0 6 0 0 1 0 0 0 0 0 0 6 21 26 26 1 8 11 13 33 3 4 5 0 0 0 1 0 8 0 19 2 1 1 3 0 1 1 8 1 Strains sensitive 6 5 3 7 3 4 41 1 8 0 4 2 15 1 4 3 1 7 1 7 11 6 3 1 5 0 3 9 0 3 0 1 6 1 2 3 5 2 4 0 2 1 0 1 0 0 13 2 % sensitive strains 100.0 0. Candida albicans Eenterococcus asacchrolyticus Eenterococcus avium Eenterococcus caecorum Eenterococcus casseliflavus Eenterococcus dispar Eenterococcus durans Eenterococcus faecalis Eenterococcus faecium Eenterococcus gallinarum Eenterococcus hirae Eenterococcus malodoratus Eenterococcus mundatii Eenterococcus raffinosus Eenterococcus solitarius Enterococcus spp.0 0.0 66. Streptococcus gallinarum Streptococcus milleri Streptococcus agalactiae Streptococcus alactolyticus Streptococcus caseolyticus Streptococcus mobilis Streptococcus spp.0 100.0 17.0 100.0 80. Lactobacillus acidophilus Micrococcus agilis Penicillium spp.4 0.7 50.4 0.0 100.0 0.0 0.0 100.4 18.5 100.0 0.0 38.0 61.0 82.0 100.0 18.0 100.0 100.0 81.6 81.0 0.0 0.5 0. .0 38.8 10.0 0.0 61.0 0.6 100.3 89.0 46. Staphylococcus aureus Staphylococcus epidermidis Staphylococcus sciuri Staphylococcus xylosus Staphylococcus spp.0 100.0 0.

LGO discs with standard positive control disc (50µg mercuric chloride) and negative control disc (disc soaked in methanol and dried) was placed on the MHA plate. 1. M10. India. Mueller Hinton agar (MHA. Edwardsiella tarda (26P. was sensitive to all antimicrobial drugs and was used as control to determine the MIC of LGO. Bacillus coagulans (CB1. Minimum inhibitory concentration of lemongrass oil for different microbes Type of strain Candida albicans Enterococcus faecalis Strain number CV1PD ABY42 SV7 SV20 E31 CV14NC SV11 SV27NC SV12 SV36NC SK10S2 SK5S1 SK6S1 SKE111 CB1 CB6 A12 B17 CP62 M10 LT81 LT121 26P 1BCY 56LT1 59LT3 E382 (Control) C91 P82 P86 Results with disc diffusion method Sensitive Sensitive Sensitive Sensitive Resistant Resistant Sensitive Sensitive Resistant Resistant Sensitive Sensitive Resistant Resistant Sensitive Sensitive Resistant Resistant Sensitive Sensitive Resistant Resistant Sensitive Sensitive Resistant Resistant Sensitive Sensitive Resistant Resistant Minimum inhibitory concentration of LGO in µg/ ml 1 1 16 32 64 128 16 32 64 64 1 8 64 64 1 4 64 64 16 32 64 124 4 32 128 64 1 8 128 128 Streptococcus mobilis Staphylococcus aureus Bacillus coagulans Klebsiella pneumoniae Edwardsiella tarda Escherichia coli mold strains were confirmed according to Holt et al. For determination of MIC of selected LGO disc sensitive and resistant strains (Table 4) of Klebsiella pneumoniae (CP62. LT 81. received from National Salmonella Centre. Hi-Media. the inhibition zone around discs was measured in mm. Bareilly. Germany) using gas generating kit. To determine the effect of growth condition on disc diffusion assay. 59LT3). Plates were incubated overnight at 37° C for bacteria and for 48-72 hours at 22° C for yeast/fungi. Izatnagar. 56LT1. P82. P86). C91. Mumbai) plates were swabbed with 6-8 hour growth of test bacteria in tryptic soy broth (TSB. Pharm. coli (E382). IVRI.0001.232 Int. Res. Barnett et al. J. LT121).16275. (1986). CB6. sterile disks of five mm diameter were soaked in methanolic solution of LGO and dried at room temperature to contain 50µg of the oil. plates were allowed to dry. Plates were incubated for 24 h and zone of inhibition was recorded as for the aerobic plates. . 8 strains of Enterococcus avium were tested under aerobic and microaerobic growth conditions simultaneously. Hi-Media) medium or with overnight Sabrauds’ broth (Hi-Media Mumbai) growth of yeast and mold strains. plates were incubated in an anaerobic culture jar (Merck. For disk diffusion test. Determination of Antimicrobial activity of LGO The antibacterial activity was determined by disk diffusion method and minimum inhibitory concentration (MIC) determination assays methods of National Committee for Clinical Laboratory Standards (NCCLS) and Clinical and Laboratory Standards Institute (CLSI). For microaerophilic condition. A reference strain of E. Anaeocult® C (Merck) Cat No. respectively. (2000) and Raper and Fennell (1977). Escherichia coli (E382. 1BCY. Pharmacol Table 4.

Xenorhabdus luminescens (1) and majority of Salmonella enterica (98. agar dilution susceptibility test was performed based on modified method of NCCLS and CLSI. fonticola. SV12. All Serratia including S. Pragia fontium (70. E31. licheniformis strains were resistant to LGO discs. indica. the only one sensitive strain belonged to S. and 50% of Budvicia aquatica and Leminorella ghirmontii strains were sensitive to LGO (Table 3).1% of 23) and two of the three strains of E. of the 41 strains of four species of Proteus.3% strains of Citrobacter freundii and all strains of C. aeruginosa and P.Singh et al. Staphylococcus aureus (SK10S2. washed (with NSS) cells of overnight grown bacteria (S. coagulans (19. coli. 7). B. marcescens. Among Klebsiella species strains.2% of 1114 strains of different microbes were sensitive. However. terrigena (5 of 6) and K. SKE111). Plates were poured and after solidification. haembachii were resistant to LGO. 64. On the other hand. E. To determine that LGO is either microbiostatic or microbicidal.01 mg/ ml. SK5S1. and Erwinia ananas (75% of 12 each). Providencia spp. rubidiae were sensitive to LGO. 128. Zone of inhibition also reduced significantly under microaerobic growth conditions. and E. Streptococcus mobilis (SV11. Among the members of Enterobacteriaceae majority of Edwardsiella (73. A. E. caviae (10 of 12). faecium (11). The only strains of Edwardsiella hoshiniae and 77.3% of 22 E.7% of 112). 33. Out of 213 strains of enterococci. Serratia spp. 8. excepting a few strains of Enterobacter agglomerans (39. Similarly. All molds (Apergillus spp. 2 and 1 µg /ml concentration of essential oil in 0 molten (at 45 C) MHA. SV20. amalonaticus were resistant to LGO discs (Table 2). 157 were resistant to LGO including all strains of E. (62.. 32 were resistant to LGO without much variation among different species except the only strain tested of P. Escherichia spp. Citrobacter spp. SV36NC). coli 382) and yeast (C. Similarly.3% of 6 each). A. Although 78% aeromonads were sensitive to LGO. S. 19.6% of 17). raffinosus (5). 1024 µg /ml) was taken as standard and two fold dilutions were made to achieve 256. salmonicida ssp. vulneris were resistant to LGO. All five strains of Providencia rettgeri but no strains of P. yeasts (Candida albicans. B. Laclercia adecarboxylata (1). however. strains of K.4%) were sensitive but all strains of P. A. SK6S1. myxofaciens (Table 2). enterica ssp. Penicillium spp. On the same lines. Among the Gram negative bacteria there was a wide variation in sensitivity of bacterial strains to LGO discs among different genera and different species of a genus (Table 2). (93. 87 were resistant to LGO but 50% of E. 233 A12. veronii (13 of 14).3%) and all six B. LGO dissolved in sterilized dimethylsulphoxide (DMSO. salmonicida (3 of 5) were sensitive to LGO discs. Aliquots were plated in triplicate for counting the cfu/ ml after serial dilution in NSS. A. The effect of reduced oxygen and enhanced carbon-di-oxide in incubating chamber was also evident.. RESULTS Results of antimicrobial activity of LGO using disc diffusion method revealed that 38. the MIC was determined.3% of 59). Lactobacillus acidophilus (1) and Morganella morganii (3) strains tested were sensitive to LGO (Table. only 92. 16. enterica ssp. the plates were spot inoculated with loopfull (2 µl) of overnight grown bacterial/ yeast cultures. The test was carried out in triplicates and plates were incubated overnight at 37° C for bacteria and 22° C for yeast. all Hafnea alvei (4). 11.8% of E. (78. In LGO containing BHI medium or NSS.3% of E. In contrast. out of 59 salmonellae 58 were resistant to LGO. All tests were repeated thrice for conformity. 100 mg /ml) was mixed with sterilized normal saline solution (NSS) or with brain hear infusion (BHI) medium (Hi-Media) to the final concentration of 1 mg/ ml and 0. none of the S. (77. oxytoca (7 of 9) were resistant to LGO. salmonicida ssp. None of the strains belonging to 11 Bacillus spp. 3). brevis (25%). hydrophila (15 of 18). A. blattae and one of the two strains of E. fergusonii strains were sensitive. variation in LGO sensitivity pattern was evident in Gram positive bacteria too (Table 3). After 18 to 24 hours. majority of the strains of A. pentothenticus (6. enterica ssp.2% of 55).3%) and many of the streptococci (53. fluorescens were resistant to LGO. Only three stains were resistant under aerobic incubation while six turned resistant under microaerobic incubation. houtenae and S. A. ABY42) were added at concentration of 42000 colony forming units per ml. and Kluyvera cryocrescens (83. (78% of 41). achromogenes (2 of 3) and A. of the 8 Enterococcus avium strains tested simultaneously under aerobic and microaerobic conditions. media (9). 3). eucranophila (10 of 18) were resistant to LGO. malodoratus (3) and majority of . aureus SKE111.. diversus and C. 1) while for other bacteria results varied with species of the microbes (Table 2. odorifera and S. LGO dissolved in sterilized dimethyl-sulphoxide (DMSO. S.1%) were sensitive to LGO while majority of enterococci (73. Similar to Gram negative strains. Proteus spp. Out of 112 strains of Escherichia. Enterococcus faecalis (SV7. pneumoniae were the most sensitive (40% of 95) while majority of K. majority of the strains of A. sobria (3). Many of the pseudomonads (46. (Table 3) was resistant to LGO. and Klebsiella spp. tarda were sensitive to LGO however. Briefly. SV27NC.9%).8%) were resistant. schubertii (8).7% of 110) strains were resistant to lemongrass oil (Table 1). albicans. amnigenus group II along with one unidentified Enterobacter strain all Enterobacter strains belonging to other six species (Table 2) were resistant to LGO. 32.6%). Aliquots were drawn at an interval of 1 min for first 10 min and then at an hour interval for 30 h. Most of the Bacillus species strains (84. salamae strain was sensitive to LGO.7%) and staphylococci (69. plymuthica strains were resistant to LGO but all the three strains belonging to S. B17). species wise analysis (Table 2) revealed that all strains of A.7% of 95). ABY42). salmonicida ssp. a few strains of B. 4. Enterobacter spp. smithia (1). CV14NC) and Candida albicans (CV1PD.

M10. albicans MIC was determined to be 1µg/ ml. albicans) strains were sensitive to LGO. and Klebsiella spp.01 mg/ ml level was only bacteriostatic for E. asacchrolyticus and E. it may be explained either on the basis of strain variation or the differences in LGO extracted from lemongrass in Nagaland and elsewhere. alactolyticus and S.01 mg/ ml of LGO it took 18 h to kill C. and Erwinia ananas (75% of 12 each).6% of 17)..7% of 95). CB6. SV27NC. 1993). 2008). aureus (SKE111) started to increase after a bacteriostatic period of 3 h. In earlier studies LGO is reported to possess potent bactericidal activity against Gram positive and Gram negative bacteria (Chao and Young. epidermidis were sensitive. Sue et al. sciuri (82. P86).. (62. C91..8%) were resistant. 2009. aureus.7% of 110) strains were resistant. and our findings are in concurrence to earlier reports (Abd-El Fattah et al. At higher concentration one may not find any strain resistant but at lower levels of LGO even the most sensitive may appear as resistant. Both the strains of Staphylococcus xylosus were resistant but both the strains of S. Alam et al. LTLT121). J. 1999) but in most cases the concentration used to kill the bacteria was too high (1 to 100mg/ml) varying for different organisms (Ferdinand et al. coli (E382) and C. Pharm. 1BCY. E31. However we have not tested fungal strains for MIC of LGO but by analogy (that all strains tested sensitive with disc diffusion assay had MIC not more than 32 µg/ ml) we may predict that the isolates of Penicillium.1%) were sensitive to LGO while majority of enterococci (73.. 1994. Pragia fontium (70. most of the strains of S. Syed et al.6% of 23) and S. dispar (26 of 29). SV20. 2003. Both the C. Both the sensitive cultures were killed within a minute while resistant S. 73. Our observations revealed that all 14 fungal (Aspergillus spp. SK5S1. Saikia et al. it might be due to use of different strains in earlier studies (Botrytis cinerea) which might be more resistant than the strains of C. indicating the microbicidal action of LGO. aureus (SKE111) and LGO sensitive E. most of the S... ABY42) revealed (Table 4) that all the strains tested sensitive to LGO (with disc diffusion method) had MIC ≤32 µg/ ml while those resistant had MIC ≥64 µg/ ml.234 Int. and Kluyvera cryocrescens (83. however. Therefore we need to standardize the cut off limit for the concentration according to the tolerance of LGO. Serratia spp. solitarus were sensitive to LGO. (78% of 41). coli (E382) and C. (78. albicans. SKE111).2% were sensitive and clear zone of growth inhibition (≥8 mm) was evident.7%) and staphylococci (69. Escherichia coli (E382. SV12. Escherichia spp. Sharma et al.. LT 81. hirae (33 of 42) were resistant but all the unidentified enterococci and sole strains of E. CV14NC) and Candida albicans (CV1PD. LGO was effective against several Gram positive and Gram negative bacteria but its effectiveness cannot be generalized beyond certain levels of concentration. when cultures were suspended in NSS containing 0. 2003). In our study. 38. E. Enterobacter spp.. But this resistance or sensitiveness is comparative and results vary with the concentration of LGO used.. Although in earlier studies antimicrobial activity of LGO has been reported higher against bacteria than fungi and yeast (Helal et al. SV36NC). Ohno et al. Abu-Seif. in our study with C. Penicillium species) and 7 yeast (C. 2009). Staphylococcus aureus (SK10S2. Due to LGO’s antifungal activity it has been claimed as an effective fungi control agent suitable for protection of food (Patker et al. 1989. coli strains but had no bactericidal effect on S. 2006) with a MIC for yeasts ~2 µl/ ml. 56LT1. Abu-Seif et al. agalactiae.7 % of non-classified streptococcal strains. flavus and A.9%) and 50% of Budvicia aquatica and Leminorella ghirmontii strains were sensitive to LGO while majority of Salmonella enterica (98.7% of 112). Determination of MIC through agar dilution method against resistant and sensitive strains of Klebsiella pneumoniae (CP62. da Silva et al. A. Pharmacol the strains of E. on cultures of LGO resistant S. aureus (61.3% of 6 each). revealed that LGO was more active while bacteria were in NSS than they were in BHI. 2006).. 2010. 1995. Similarly among Gram negative bacteria 78% aeromonads. albicans (ABY42). caseslliflavus (26 of 32). mobilis (61. B17).. LGO sensitive bacteria could be detected for 6 h and resistant strains was present up to 18 h of exposure.. no strains of S. milleri. however..9%) and 66. 1993. Majority of the strains of streptococci were sensitive to LGO including all strains of S. (93. albicans strain had MIC 1 µg/ ml while for bacterial strains sensitive to LGO discs it ranged from 1 µg/ ml to 32 µg/ ml. Onawunmi. Streptococcus mobilis (SV11.2% of 55).5% of 13) were resistant to LGO. niger also had MIC ≤32 µg/ ml which is much lower than that reported earlier 1. Studies to determine that the action of LGO on microbes is either microbiostatic or microbicidal. albicans and 24 h for killing E. 59LT3). 2009) and phenolic compounds (Abu-seif et al. Observations revealed that all bacterial strains of a genus may not be equally sensitive as 115 Bacillus species strains (84. DISCUSSION Of the 1114 strains of microbes tested for sensitivity to LGO discs (50µg LGO/ disc). Providencia spp.. 2009. In BHI. LGO at 0. On the other hand in BHI. Edwardsiella tarda (26P.3% of 59). et al. Proteus spp. Citrobacter spp. Bacillus coagulans (CB1.5 µl/ ml (Helal et al.3%) and many of the streptococci (53. aureus (SKE111) was detected even after 5 minutes but not at 10 minute of exposure of microbes to 1mg / ml in NSS. E. SK6S1.. at 1mg/ ml concentration all the microbes tested including the resistant S.9% Edwardsiella (73... aureus were killed within 5 minutes while only those which were sensitive with disc diffusion method could be eliminated at 10 µg/ ml concentration. 1991. gallinarum (13 of 16) and E. albicans (ABY42) while number of S. (77. P82. Similar results have been reported earlier . A12. S. Antifungal activity of LGO is proved to be due to its flavonoids (Pratt and Hudson. However.. 2000. Enterococcus faecalis (SV7. Res. 2008. Nieto et al. caseolyticus was sensitive to LGO.

Dudley MN (2010). ACKNOWLEDGEMENTS Authors are thankful to Naga Fragrance Pvt. Jharnapani. 34: 219-229. U. Antimicrobial activity of some essential oil against Paenibacillus larvae. Theoduloz C. Barnett JA. Ali-Shtayeh MS. Ramadan MM (2009). Agric.ehow. 3rd ed. 32: 396-399. Am. It can be concluded from the observations that LGO is bactericidal and fungicidal at higher concentration (1mg/ ml) while bacteriostatic at lower concentrations (<10µg/ ml). aureus. the confusion regarding antimicrobial activity among different study in relation to MIC might be due to difference in the medium and the method used. Infect. J. pyogenes and S. Re MS. Evaluation of the antimicrobial properties of unripe banana (Musa sapientum L. Citron DM. Yeasts: characteristics and identification. 2. Antimicrobial activity of some essential oils against microorganisms deteriorating fruit juices. pneumoniae. their uniform sensitivity was indicative of wide spectrum of LGO’s antimicrobial action. and aztreonam under anaerobic conditions (King et al. Cerimele EL. vol. Sharpee ME (1986). All microbes are not equally susceptible to LGO as Bacillus spp. Shaaban HA. Abu Ghdeib SI (1999). Ogunsanya T. fungi and viruses. Inouye S. approved standard . Weisheimer V. CLSI document M07-A8. Comparison of extraction methods for marker compounds in the essential oil of lemongrass by GC. Rodríguez J. J. Payne RW. Performance standards for antimicrobial susceptibility 235 for Hemophillus influenzae. The Phillippine Agric. Maridable J. Wayne. 4: 9-16. more studies are required to understand effect of microaerophilic growth conditions on sensitivity of microbes and to have a broad idea about utility of general diffusion assay for facultative anaerobes and microaerophilic microbes. amikacin. and 50 µg / disc concentration is quite feasible option to explore the affectivity of probable antimicrobial herbs. Further. Huynh KP. Antimicrob. 8: 1176-1182. Sci. Method for dilution antimicrobial susceptibility tests for bacterial that grow aerobically.). Baltemore. Animal and Mineral Origin in Pakistan. seagrass and lemongrass oil from Papua New Guinea for antimicrobial and antifungal activity. Chao SC. Authors also thankful to Joint Director. Alippi MA.K.) on pathogens. Guterres SS. Saeed A. U. University of Karachi. Omotoyin A (2009). J. J. Preliminary screening of seaweeds. Abu Shahla ANK. J. tarda among Gram negative bacteria are comparatively more susceptible to LGO than most of the other potentially pathogenic bacteria. Burrows I. Diagnostic . Bayoum HM. 50: 1345-1349. J Herbs Spices Med. Clinical and Laboratory Standards Institute (2008). Ferdinand FJ. Although number of yeast and mold strains was less (21) in the study. The effect of oxygen deficient and CO2 rich environment as expected under in vivo conditions was highly significant in reduction of the sensitivity of eight E. Lomovskaya O. 6: 582-594. Agric. Food Chem. Therefore. Traditional Medicine of Herbal. Biotechnol. avium strains tested. Clinical and Laboratory Standards Institute (2009). http://www. inhibited at at 100 µg/ ml concentration (Inouye et al.A. for providing LGO free of cost as and when required. Variation in LGO activity (MIC) on different strains of bacteria is inevitable as for most of the antimicrobials. lemon grass (Cymbopogon citratus S. 2004). Alam K. El-Khai EKA (2006). Essent. Sarhan MM. Tradit.. Director of the ICAR Research Centre for NEH Region and Director NRC on Mithun. Abo Sreia YH. Agua T. 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