Potential Antibacterial Efficacy of Garlic Extract on Staphylococcus aureus, Escherichia coli, and Klebsiella pneumoniae: An In vitro Study

Abstract

Background:

Garlic has been recommended by many ancient medicines such as the Chinese and the Indian medicine to cure respiratory and digestive issues along with treating microbial infestation and leprosy. The therapeutic effects encompass many advantages in the field of cardiovascular system, antibiotics, anticancer, anti-inflammatory, and hormone-like effects.

Aims and Objective:

The present study was carried out to evaluate the garlic antibacterial effect against clinical isolates of Staphylococcus aureus, Escherichia coli, and Pneumoniae from patients attending referral hospital.

Materials and Methods:

The isolation of bacteria was done from pus sample collected from referral hospital, Bedar, Karnataka, with sterile swabs. The study samples were inoculated under aseptic conditions on culture media such as nutrient agar, blood agar, and MacConkey agar plates and isolated the pathogen bacteria such as E. coli, Klebsiella pneumoniae, and S. aureus. The garlic bulbs were peeled off and then ligated using pestle simultaneously with addition of minor quantity of H₂O for preparation of plant extract and study the antimicrobial effect of garlic on these bacteria.

Results:

The result showed that garlic extracts have a high range of antibacterial effect against both Gram-negative (E. coli and K. pneumoniae) and Gram-positive bacteria Staphylococcus.

Conclusion:

The present study observations revealed that garlic makes large clear zones in comparison to the currently available antibiotics used in the study. The potentiality of the garlic can be utilized in the field of antibacterial agents. It can be prepared in the form of tablets in the best concentrations and affordable dosages so that it can be used as medicine against these pathogenic organisms.

INTRODUCTION

Garlic is an herb which is grown across the globe and known well for its anti-infective properties. It falls in family: Amaryllidaceae, order, Asparagales, and Kingdom: Plantae.[1,2] The botanical name of garlic is “Allium sativum” deriving it from the Celtic word “all,” which stands for burning or stinging, and the Latin “sativum” stands for planted or cultivated.[2] It has a long traditional history as medicinal plant, started with a direction of preparing a medicinal remedy written in a cuneiform character in about 3000 BC to present date formulation of tablets. Scientific investigations of various garlic preparations began in 1939.[1]

Naturally occurring plants play a pivotal role in developing the ancient medicine as it has many therapeutic properties which in previous time in the absence of good diagnostic tools contribute immensely in alleviating patient's pain and improving the social well-being.[3] The therapeutic properties range from the beneficial effects on the cardiovascular system, central nervous system, anticancer effect, anti-inflammatory, antibiotics, and immunity-boosting effect.[4] These extracts are nothing but the by-products which are released during the secondary metabolism of the plants. The availability of these herbal medicine is in many forms such as the powder, liquid, or mixtures which are formulated in paste and ointment forms.[2] The ancient traditional science utilizes these properties and adopts indigenous methods to maintain health system.[5] They also used it for prevention, diagnosis, and treatment of the diseases by accumulating the knowledge, skills, and practices derive from the concepts of different cultures that are acknowledged to maintain health.[6] In underdeveloped and developing countries, a big segment of population put their belief in the traditional form of medicine to cure any kind of illness and improve the health.[7] Garlic (A. sativum) is one of the very selected variety of plant species on which lot of research has been carried out to explore its medical propertied qualities and apply it for cure of various health issues including cancer.[3,4,7,8]

Garlic growth is by vegetative reproduction rather than sexual reproduction (seed) producing the individual cloves which contain bulbs having the same genetic properties as the original clove. The earliest evidence of these medicinal plant extract was mention in Avesta, a Zoroastrian book complied during the 6^th century along with its detail description in the civilization of Sumerian and Egypt. Indian and Chinese's ancient civilization also recommended the use of garlic for curing respiratory ailment, digestive issues, leprosy, and parasitic infestation.[7] Al Qanoon Fil Tib (The canon of medicine) suggested that the garlic can be powerful curative effect in a condition such as arteritis, toothache, chronic cough, constipation, insect bites, and gynecologic disease.[9] There is ample of epidemiologic instances that elaborates many good health benefits of garlic based on the experimental and clinical investigations.[10,11]

Latest studies carried out in Ethiopia suggested that garlic has been a common medicine used in its traditional medicine for treating infectious diseases such as tuberculosis, sexually transmitted disease, and wounds. Apart that, it serves as a main culinary preparation and applications.[8,9,10] We carried out this study to evaluate the antibacterial effect of garlic against clinical isolates of Staphylococcus aureus, Escherichia coli, and Pneumoniae.

MATERIALS AND METHODS

Isolation of bacteria

For bacteria isolation, pus samples were collected by sterile swabs from inpatients and outpatients of different wards of referral Hospital, Bedar, Karnataka, for a period of 6 months from July 2019 to December 2019 in accordance with standard protocols and ethical guidelines. Skin, nasal wounds, ears, legs, internal organs (lungs, kidney, and bladder), and catheters served as samples for collection of pus samples.

Pus samples were preserved in Cary Blair transport medium and transported to the S. B, Patil Dental College, Department of Oral Pathology and Microbiology, Bedar, Karnataka, for the Gram stain and culturing procedure. The samples were inoculated under aseptic conditions on nutrient agar, blood agar (5% sheep blood), and MacConkey agar plates, incubated aerobically at 35°C–37°C for 24–48 h. For primary identification and characterization of isolates were performed on the basis of Gram staining, microscopic characteristics, colony characteristic, and secondary identification were done with the help of biochemical tests such as tripal sugar iron agar, hydrogen sulfide test, carbohydrate fermentation test, phenylalanine deaminase test, methyl red test, nitrate reduction test, urease test, Vogesproskauer, citrate utilization test, and indole test using standard microbiological methods.

Preparation of plant extract

Fresh garlic extract preparation from the plant's bulbs was taken from the local market. The garlic bulbs were peeled, weighed (50 g), and cleaned. The peeled garlic bulbs were then ligated using pestle with simultaneously addition of small quantity of water. This extract was considered as the 100% concentration of the extract and used for antimicrobial effect on isolated bacteria.

Antibacterial sensitivity test

The antibacterial activity test of the crude extract of garlic against clinical isolated bacteria such as E. coli, Klebsiella pneumoniae, and S. aureus was carried out by the Agar diffusion method.[11,12] E. coli, K. Pneumoniae, and S. aureus were inoculated on a nutrient agar plate with the help of sterile cotton swab. Then prepared the wells in the center of Petri plate with the help of puncher. After inoculation of bacteria, 10 μl garlic extract was added with the help of micropipette, and then incubated at 35°C–37°C for 24 h. After 24 h the diameter of the ring was measured.

Statistical analysis

One-way ANOVA was used to compare the mean values as a measure of test of significance. A P < 0.05 was considered statistically significant.

RESULTS

The results of the susceptibility of the test organisms against the garlic extracts showed that isolates of S. aureus, E. coli, and K. pneumoniae were sensitive to the concentration of 10 μl garlic of agar media in using diffusion method. In addition, larger clear zones were observed against Staphylococcus its 28 mm and then 2^nd largest clear zone on E. coli that is 27 mm and K. pneumoniae also show good clear zone 22 mm at 10 μl concentration against among microorganisms [Table 1, Graph 1 and Figures 1–3]. A P = 1.000 was found, and there was no statistically significant difference between the three groups.

Table 1.

Antibiotic sensitivity test results

Concentration of garlic (ml)	Diameter of clear zone (in mm) on nutrient agar and macConkey agar			P
	Escherichia coli	Klebsiella pneumoniae	Staphylococcus aureus
10	27	22	28	1 (NS)

NS: Not significant

Graph 1.

Antibiotic sensitivity test results

Figure 1.

Effect of garlic on Staphylococcus aureus

Figure 3.

Effect of garlic on Escherichia coli

Figure 2.

Effect of garlic on Klebsiella pneumonia

DISCUSSION

Naturally occurring species such as garlic and other herbs produce secondary metabolites which are useful for health, but simultaneously these bioactive compounds can cause adverse reactions in the body such as allergy, cardiovascular system (CVS) problem, dermatitis, and bleeding unless there are used under controlled protocol and guidelines.[11,12] However, to confirm the therapeutic use, further studies are needed to find out its efficacy and potential side effects.[6,9]

This research mainly concentrates on the antibacterial effect against multidrug-resistant human pathogen E. coli, K. pneumoniae, and S. aureus with their respective inhibition zone.[12,13] Minimum inhibitory concentration or high zone of inhibition elaborates the bacterial susceptibility or bacterial reaction to the antibiotic used.[14,15]

The extract of the garlic contains a varied range of antimicrobial/antibacterial potential which are effective against Gram-negative organisms (E. coli and K. pneumoniae) and Gram-positive bacterial Staphylococcus.[12,13,14,15] The organism which is very stubborn such as the antibiotic-resistant bacterial and their toxic by-product can also be countered with the help of these garlic extracts.[13] The components which bring about this effect are the allicin which mainly inhibits the growth of bacteria by inhibiting the DNA and protein synthesis partially along with RNA inhibition synthesis as the primary target.[16] In addition, there are various studies that have been carried out which suggest the allicin potential to stop the RNA synthesis speed by trapping the RNA peptides chain reaction and amplify the antibacterial.[17]

S. aureus organism membrane consists of lipid, which gives it protection.[7] Allicin, a component of garlic, has the ability to penetrate this membrane and consequently influences the RNA mechanism, lysis of membrane along with bactericidal effect.[12,13,14,15,16] The effect of inhibition against the growth of these microorganisms relates to the fact that E. coli and K. pneumoniae are made up of 20% of lipid while S. aureus 2% lipid.[7,18]

Findings of the current study are concordant with the studies conducted by Jehan et al.,[2] Onyeagba et al.,[3] Shaloo et al.,[4] Nkang et al.,[14] Deresse,[15] Abebe,[16] Yadav et al.,[17] Aliy,[18] and Shokrzadeh and Ebadi.[19]

Therefore, the garlic extract is more potent against S. aureus and the resistant species of S. aureus. As the antibiotic resistance has become challenging scenarios in current medical practice, so in such cases, these findings of garlic and its extracts come as a boon to the patients and medical field.[19]

CONCLUSION

The present study results suggest that the garlic exhibits a large clear zone compare to the currently available antibiotics used in the study. Garlic can be used as an effective antibacterial agent formulating it in the form of tablets in the best concentrations and affordable dosages so that it can serve as a medicine against these pathogenic microorganisms. In this era of drug-resistant bacteria, we need to get our research strengths in usage of alternative medicine that has a past strong record in treating various disease pathogens with the help of these naturally occurring herbs.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.