Effect of honey on multidrug resistant organisms and its synergistic action with three common antibiotics.S Karayil, SD Deshpande, GV Koppikar
Department of Microbiology, T.N.M.C. & B.Y.L. Nair Charitable Hospital, Mumbai.
A total of 15 bacterial strains (7 Pseudomonas & 8 Klebsiella species) isolated from various samples which showed multi-drug resistance were studied to verify in vitro antibacterial action of honey on the principle of Minimum Inhibitory Concentration (MIC) & its synergism with 3 common antibiotics--Gentamicin, Amikacin & Ceftazidime. The MIC of honey with saline for both organisms was found to be 1:2. The synergistic action was seen in the case of Pseudomonas spp. and not with Klebsiella spp.
Keywords: Amikacin, therapeutic use,Antibiotics, therapeutic use,Ceftazidime, therapeutic use,Drug Resistance, Multiple, Drug Synergism, Gentamicins, therapeutic use,Honey, Klebsiella, drug effects,Microbial Sensitivity Tests, Pseudomonas, drug effects,
The emergence of multi-drug resistant organisms has created a lot of chaos in the medical field recently. The organisms responsible for nosocomial infections have also shown an increased trend to be multi-drug resistant. Hence there is a need to find an alternative to counter these multi-drugs resistant organisms.
Honey has been used as an adjuvant method of accelerating wound healing from ancient times & has been sporadically used in treatment of burns,,,. It is reported to prevent infection & promote healing, since it has ingredients very similar to antibiotics.
In the present study emphasis has been laid to find the Minimum Inhibitory Concentration (MIC) of honey & its synergistic action with three common antibiotics - Gentamicin, Amikacin, Ceftazidime using multi-drug resistant Pseudomonas and Klebsiella species.
Collection of organism
The multi-drug resistant organisms isolated from samples like bile, blood, pus, ascitic fluid etc. were collected and categorized as A, B and C as shown in [Table - 1]. The multi-drug resistance was determined by Kirby - Bauer disk diffusion method.
These organisms were then subcultured on a nutrient agar slant. The classification of organisms is given in [Table - 2].
Selection of honey
The brand of honey used had the Agmark seal. It was packed and marketed by Phondaghat Pharmacy. The honey was of standard grade. The standard grade of honey has the following properties. Specific gravity-1.37 at 27 degrees, moisture -25% max, reducing sugars-65% min, sucrose-5% max, fructose:glucose-1.0 min, acidity (as formic acid)-0.2% max, ash-0.5% max.
Studies carried out
Three different studies were carried out with honey. These were: - MIC of honey with physiological saline, MIC of honey with Soybean Casein Digest broth (SCD), Synergistic action of honey with different antibiotics by dilution method.
Different concentrations of honey were prepared in SCD as well as physiological saline to give two sets of 1:2, 1:4, 1:8 & 1:16 dilutions each. One tube containing 1ml of undiluted honey was also used.
For synergistic action, three common antibiotics -- Gentamicin, Amikacin, & Ceftazidime were selected. The MIC values of these antibiotics as per NCCLS standards were 8mcg/ml, 32mcg/ml & 16mcg/ml respectively. The dilution method was followed for determination of synergistic action. Honey & the antibiotics were mixed in 1:1 ratio.
24 hours old cultures of Pseudomonas and Klebsiella species were inoculated in saline & the bacterial concentration adjusted to 0.3 x 108 cells/ml with the help of Brown’s Opacity tube no: 16. The cultures were inoculated in the dilutions and incubated at 37?C. After 1hour, 2hours, 3hours, 4hours, 5hours, & 24hours, the samples were spot inoculated with a standard loop of 3mm on MacConkey’s agar plate & the plates incubated at 37?C for 24hours & observed for growth.
A positive control --> (1) SCD + inoculum, (2) Saline + inoculum & a negative control --> (1) SCD only, (2) Saline only was maintained throughout the experiment. The same procedure of spot inoculation as that of the test samples was carried out for the controls. The positive control was incubated at 37?C & the negative control at 4?C.
For this study 50 samples were screened and fifteen organisms were selected. Out of these 7 were of Pseudomonas species & 8 were of Klebsiella species. These organisms showed resistance to Gentamicin, Amikacin & Ceftazidime and hence these antibiotics were used to study the synergistic action with honey on selected strains.
The results obtained from the tests are given in [Table - 3], & [Table - 4]. A positive control & negative control was maintained for each of the tests.
Total number of Pseudomonas species studied was 7. Of these two belonged to category A and five belonged to category B.
Total number of Klebsiella species studied was 8. Of these five belonged to category A, 1 belonged to category B and two belonged to category C.
The present study was undertaken to determine the antibacterial action of honey & its synergistic action with three common antibiotics on multi-drug resistant organism. The main objective underlying this study was to ascertain whether honey could be given orally to the patients along with the antibiotics in case of infection from multi-drug resistant organisms.
The antibacterial properties of honey can be attributed to its low pH, a thermolabile substance called inhibine and its hygroscopic properties,,,. Honey also contains enzymes such as catalase that may aid the healing process,,. Honey serves as an important medicine because of its mild laxative, bactericidal, sedative and antiseptic characteristics. Honey is also known to act as highly viscous barrier preventing bacterial penetration and colonization of wound surface,,. Honey works quicker than many antibiotics because it is easily absorbed into the blood stream. Honey also shows antivibriocidal activity.
The studies conducted shows that undiluted honey as well as dilutions prepared in saline inhibited both Pseudomonas as well as Klebsiella spps. whereas dilutions prepared in SCD showed inhibition only of Pseudomonas spps. The synergism was observed only in Pseudomonas spps. and not in Klebsiella spps.
The studies conducted so far by various workers have shown that honey has excellent wound healing properties and can be used effectively for topical applications,,,. In this study we have tried to show its antibacterial properties in vitro and from the results obtained we can suggest that it may be possible to give honey orally along with the antibiotics in case of infections from multi-drug resistant organisms. This is only a pilot study limited to 15 strains of organisms nevertheless it forms a foundation for future research in this field.
[Table - 1], [Table - 2], [Table - 3], [Table - 4]