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Evaluation of Efficiency and Extent of Inhibitory Characteristics of Lactobacillus spp. (Lactobacillus plantarum) against Selected Pathogenic Bacteria

Peerzada Rouf Ahmad1, Mudasir Ali Rather1, Burhan Nabi2,*, Omer Mohi U Din Sofi3, Mahrukh Hafiz4, Iyman Binti Fayaz1, Mohammad Mateen Zehgeer5
1Division of Veterinary Public Health, Faculty of Veterinary Sciences and Animal Husbandry, SKUAST, Shuhama-190 006, Kashmir, Jammu and Kashmir, India.
2Division of Veterinary Medicine, Faculty of Veterinary Sciences and Animal Husbandry, SKUAST, R.S. Pura-181102, Jammu, Jammu and Kashmir, India.
3Division of Veterinary Parasitology, Faculty of Veterinary Sciences and Animal Husbandry, SKUAST, R.S. Pura-181102, Jammu, Jammu and Kashmir, India.
4Division of Veterinary Microbiology and Immunology, Faculty of Veterinary Sciences and Animal Husbandry, SKUAST, Shuhama-190 006, Kashmir, Jammu and Kashmir, India.
5Veterinary Officer, Central Animal House Facility, Jamia Hamdard, Delhi-110 019, India.

ABSTRACT

Background: Despite the improved modern technologies to inactivate pathogens, there has been a continuous surge of food-borne diseases involving many bacterial pathogens like Staphylococcus aureus, Escherichia coli, Bacillus cereus, Salmonella, Campylobacter, Listeria etc. Physical methods of reduction of microbial load in raw foods have been known to negatively impact the organoleptic properties of the products hence reducing their acceptability. Therefore, knowing that food may be laden with many pathogens, the biocontrol methods may be an effective alternative to chemical and physical methods of preservation for eliminating pathogens.

Methods: The pathogens to be tested in the experiment were recovered from various food sources and clinical samples viz., milk, raw chicken and a faecal sample, chicken. For evaluation of inhibitory characteristics of Lactobacillus spp. (Lactobacillus plantarum) against selected pathogenic bacteria the grown culture of pathogens (S. aureus, E. coli, B. cereus) was added with Lactobacillus spp. at different concentrations and incubated at 24, 36 and 48 hrs and mean CFU units were evaluated. All of the treated groups were compared with the concurrent untreated groups of respective bacteria.

Result: After 24 hrs of incubation at 37°C, the mean colony forming units (CFU/ml) of S. aureus, E. coli, B. cereus in the cultured groups treated with 108, 107, 106, 105 and 104/ml of L. plantarum were 2.56 x 108, 2.78 x 108, 2.90 x 108, 3.03 x 108 and 39.19 x 108; 0.81 x 108, 1.09 x 108, 1.36 x 108, 1.85 x 108 and 24.62 x 108; 0.31 x 108, 0.43 x 108, 0.60 x 108, 0.69 x 108 and 7.12 x 108, respectively. Henceforth, the mean colony forming counts of all the three pathogens increased when the concentration of L. plantarum decreased.

INTRODUCTION

Despite the improved modern technologies to inactivate pathogens, there has been a continuous surge of food-borne diseases involving many bacterial pathogens like Staphylococcus aureus, Escherichia coli, Bacillus cereus, Salmonella, Campylobacter, Listeria etc. These pathogens come into contact with foods during harvest or slaughtering, processing, storage and packaging. For reducing pathogenic bacteria physical treatments such as UV light, high pressure, dry heat and steam have been encouraged as the use of antibiotics has been restricted over the years due to the risk of antibiotic-resistant bacteria entering the human food chain and leading to negative impact on human antimicrobial treatment. However, such methods have been known to negatively impact the organoleptic properties of the products thus reducing their acceptability (Ravi et al., 2017). Many of the chemical preservatives are known potential carcinogens and have many other ill effects on human health. Thus there has been an increasing need to develop novel strategies to reduce bacterial pathogens in foods and still satisfy consumer demand for minimally processed foods with low concentrations of chemical preservatives (Garcia et al., 2008). Therefore, knowing that food may be laden with many pathogens, the biocontrol methods may be an effective alternative to chemical and physical methods of preservation for eliminating pathogens.
       
Because of the wide spread association of bacteriocins, nisin and pediocin with foods and is generally recognized as safe (GRAS) status, the use of Lactobacilli spp. and/or their metabolites as natural drugs or antimicrobial agents have attracted considerable interest in recent years (Reis et al., 2012). Lactobacillus plantarum is one of the most important species of LAB, with proven health benefits and antagonistic properties against the various pathogen (Arena et al., 2016). In this regard, the present study has been undertaken to determine the efficiency and extent of inhibitory characteristics of Lactobacillus spp. (Lactobacillus plantarum) against selected pathogenic bacteria.
 

MATERIALS AND METHODS

In the current study, the reference strain of Bacillus cereus (NCTC 11145) was available with the Division of Veterinary Public Health, Faculty of Veterinary Science and Animal Husbandry (F.V.Sc and A.H), Alusteng, SKUAST-K. The standard strains of Staphylococcus aureus (ATCC 25293) and Lactobacillus plantarum (ATCC 8014) used in the study were procured from Hi-media, Mumbai. The characterized shiga toxin Escherichia coli was available with the Division. The strains were maintained on nutrient agar slants by subculturing every fortnight and were tested for purity.
       
A total of 60 samples comprising of 30 samples of milk (15 mastitic milk, 15 raw milk), 15 samples of raw chicken and 15 faecal samples from sheep were collected randomly from the different areas of district Srinagar. The milk and faecal samples were collected in sterile vials. The raw chicken samples were collected aseptically in sterile zip lock polythene bags and transported in ice to the Division of Veterinary Public Health, F.V.Sc and A.H, Shuhama and processed for isolation and identification of Staphylococcus aureus, Bacillus cereus and Escherichia coli. Five samples from each source were collected separately for the isolation of the pathogens, the details of the samples are provided in Table 1.

Table 1: Type and Nature of samples collected for isolation of S.aureus, B. cereus and E. coli.


 
Isolation and identification of Staphylococcus aureus
 
Briefly, for isolation of S. aureus, the samples were enriched in buffered peptone water (BPW) and incubated at 37°C for 18-24 hrs. A loop full of inoculum from the enriched medium was directly streaked on Baird-Parker Agar (BPA) plates containing egg-yolk tellurite emulsion. The inoculated plates were incubated at 37°C for 24 hrs. The typical jet-black colonies surrounded by white halo on BPA were considered as presumptive positive for Staphylococcus spp. These presumptive positive isolates were preserved on nutrient agar slants, stored at 4°C till further processing and were identified and confirmed based on cultural, morphological and various biochemical characteristics as per the standard protocol (Cruickshank et al., 1970).
       
The presumptive Staphylococcus isolates were subjected to morphological and biochemical characteristics for further confirmation. The biochemical tests included catalase reaction, oxidase testing, indole reaction, methyl red test, voges-proskauer test and coagulase test (Cruickshank et al., 1970).
 
Isolation and identification of Escherichia coli
 
Isolation and identification of E. coli was done by using standard bacteriological procedures (Cowan and Steel, 1974). The milk, meat and faecal samples were enriched overnight at 37°C in buffered peptone water and were subsequently inoculated on MacConkey’s agar (Hi- Media, Mumbai) and incubated aerobically for 24 hr. The presumptive colonies (lactose fermenting pink colonies) were picked up and re-cultured on Eosine Methylene Blue (EMB) agar and again incubated aerobically for 24 hr. Colonies producing characteristic greenish metallic sheen colour were considered as E. coli.
       
The well-separated colonies were picked up on nutrient agar slants as pure culture and subjected to standard morphological and biochemical tests such as oxidase test, catalase test, tryptophan degradation (indole test), glucose degradation (Methyl red/Voges-Proskauer test), citrate and urease utilization test and ability to ferment triple sugar iron for further confirmation.
 
Isolation and identification of Bacillus cereus
 
Polymyxin-pyruvate-egg yolk-mannitol-bromothymol blue agar (PEMBA) media was used for isolation of B. cereus. The samples were processed as per the method described by Shinagawa (1990) and Tallent et al., (2012) with suitable modifications. Approximately 10 ml/gm of the sample, was inoculated in 50 ml of brain heart infusion broth (BHIB) containing polymyxin B (10,000 U per 100 ml). The raw chicken was homogenized with BHIB. The enrichment of samples in BHIB was carried out at 35°C for 16-18 h. After enrichment, a loopful was streaked on PEMBA plates and incubated at 35°C for 24 h. The plates showing crenate to fimbriate peacock blue colored colonies (3-5 mm) surrounded by a blue zone of egg yolk hydrolysis against a green background and other related species were purified and taken on nutrient agar slants
       
The confirmation of B. cereus was carried out as described by Rhodehamel and Harmon (2001). Briefly, the presumptive colonies of B. cereus and closely related species were collected and subjected to morphological and biochemical tests for identification of species. Gram reaction, cell morphology, spore production, motility, reduction of nitrate, Christensen’s citrate utilization, Voges-proskauer reaction, aerobic and anaerobic utilization of glucose, mannitol fermentation and hemolysis were the tests employed for identification and conformation.

Lactobacillus plantarum as a biocontrol agent against food-borne pathogens
 
The procedure consisted of the following steps:
 
Probiotic preparation
 
This step included the preparation of 0.5 Mac Farland solution (0.05 ml of 1.175% barium chloride dihydrate and 9.95 ml of 1% sulfuric acid) of Lactobacillus spp. in sterile normal saline solution. Briefly, 0.5 Mac Farland solution (Appendix-I) was taken and Lactobacillus plantarum colonies from the plates were dissolved in 9 ml of sterile NSS one by one till turbidity matched with standard 0.5 Mac Farland solution.  The number of Lactobacillus plantarum organisms at 0.5 Mac Farland were calculated by plating on selected agar (ManRogosa Sharpe agar) by serial dilution and the total viable count was evaluated. The concentration of L. plantarum equivalent to 0.5 Mac Farland was approx. 108 per ml.
       
The aliquots of this tube were prepared by serial dilution to get different concentrations of L. plantarum solution (108/ml, 107/ml, 106/ml, 105/ml and 104/ml). The efficacy of these five concentrations of L. plantarum was tested against standard concentrations of selected pathogenic microorganisms (S. aureus, E. coli and B. cereus).
 
Efficacy of Lactobacillus plantarum against pathogenic bacteria      
         
The efficacy of L. plantarum was observed against three foodborne pathogens (S. aureus, E. coli and B. cereus) recovered naturally from various foods. To determine the efficiency of L. plantarum against S. aureus, briefly, the inoculums of S. aureus were prepared as given in Table 2. Briefly, a standard 0.5 Mac Farland solution of S. aureus was prepared in sterile normal saline solution thereby containing approximately 1.5x108 microorganisms per ml. Then six tubes containing 15 ml of nutrient broth were taken and all the six tubes were inoculated with 1 ml of the standard solution of S. aureus (108/ml) and 1 ml of sterile NSS. The first tube was kept as a control with 1ml of S. aureus inoculum and 1 ml of NSS. In the rest of the five tubes, 1ml each of different standard concentrations (108/ml, 107/ml, 106/ml, 105/ml and 104/ml) of L. plantarum (mentioned above in probiotic preparation) were added. Then, all the six tubes were incubated at 37°C and viable counts of Staphylococcus aureus on selective media (Baired Parker Agar) was determined after 24, 36 and 48 hours to see the degree of reduction/increase in growth/number of organisms brought about by presence of different concentrations of Lactobacillus spp. as compared to the control.

Table 2: Effect of different concentrations of L. plantarum on counts of standard concentration of S. aureus (108/ml).


       
A similar procedure was adopted to determine the inhibition of the growth of B. cereus and E. coli by L. plantarum.
 
Antibacterial characteristics of L. plantarum against selected pathogenic micro-organisms
 
An evaluation of the antibacterial activity of L. plantarum against the selected pathogenic bacteria was evaluated as per the standard protocol (Tagg and McGiven, 1971). Briefly, the isolated L. plantarum strain was grown in MRS broth (pH 6.5) inoculated with 1% of an overnight culture and incubated at 37°C for 18-24 hours. After incubation, cells were removed from the broth by centrifugation (6000 rpm for 20 min, 4°C). The cell free supernatant was sterilized by filtering through a 0.22 milli pore filter. The antimicrobial spectrum of the cell free supernatant of different L. plantarum strain was determined using the disc diffusion method.
 

RESULTS AND DISCUSION

The present study was undertaken to determine the efficiency of Lactobacillus plantarum as a biocontrol agent against some foodborne pathogens viz. Staphylococcus aureus, Escherichia coli and Bacillus cereus. The extent of antibacterial activity of L. plantarum against these pathogens was also studied.
 
Occurrence of S. aureus, E. coli and B. cereus in foods and clinical sample
 
Out of 20 samples comprising of raw milk (5), mastitic milk (5), raw chicken meat (5) and sheep faecal samples (5) that were screened for the presence of S. aureus, 9 turned out to be positive for Staphylococcus spp. and of theses 9 isolates, 6 were identified as S. aureus (Table 3) based on morphological and biochemical characteristics. One isolate each was recovered from raw chicken and sheep faecal samples and two isolates each were recovered from raw milk and mastitis milk samples, respectively. The isolates of S. aureus on Baird-Parker Agar (BPA) showed typical jet black colored colonies surrounded by white halo zone (Lecithinase activity). On Gram staining, the gram-positive cocci arranged in bunches were seen under a microscope. The Staphylococcus isolates characterized biochemically gave a positive reaction for catalase, methyl red test and Voges-Proskauer while as negative reaction was observed for oxidase activity and indole reaction. On the basis of these tests, out of 20 samples, 9 isolates of Staphylococcus spp. were identified. These 9 isolates were subjected to coagulase test and 6 coagulase positive isolates were recognized, thereby confirmed as S. aureus.

Table 3: Occurrence of S. aureus, E. coli and B. cereus in various samples.


       
Similarly, for isolation of E. coli, raw milk (5), mastitic milk (5), raw chicken meat (5) and sheep faecal samples (5) were screened and 7 isolates were recovered (Table 3). The isolates of E. coli were recovered from raw milk (2), meat (2) and faecal samples (3) and from the mastitis milk none of the isolates of E. coli could be recovered. Of the 7 isolates recovered 4 isolates were selected for experimental testing of which one each were from raw milk and chicken and two isolates included those recovered from sheep faecal samples. All the isolates that produced a characteristic greenish metallic sheen on Eosine Methylene Blue (EMB) were presumptively considered as E. coli and were small rod shaped when seen under a microscope. The E. coli isolates characterized biochemically revealed positive reactions for catalase, indole and methyl red. While as isolates were negative for oxidase, voges- proskauer, citrate and urease utilization and triple sugar iron fermentation activity.
       
For isolation of B. cereus, 5 samples each of raw milk, mastitic milk, raw chicken meat and sheep faecal samples were screened and in total 5 isolates were recovered. The isolates were recovered from raw milk (2), raw chicken (2) and faecal samples (1). Of the 5 isolates, 4 were tested in the experiment of which, 2 were recovered from raw milk and 1 isolate each was taken from raw chicken and faecal samples. All presumptive isolates of B. cereus showed typical fimbriate or crenate peacock blue colored colonies with egg yolk reaction (Lecithinase activity) on PEMBA medium. On morphological characterization all the isolates were found to be Gram positive, rod shaped, spore formers and were devoid of toxin crystals. All the isolates were motile and strongly hemolytic (β-hemolytic) on 5 per cent sheep blood agar. The presumptive isolates of B. cereus were further characterized using biochemical tests. The isolates reduced nitrate, were also positive for VP and citrate utilization tests. All the isolates fermented glucose both aerobically and anaerobically and none of the isolates fermented mannitol.
       
The inhibitory effect of L. plantarum was observed on in vitro growth of selected pathogens (S. aureus, B. cereus and E. coli). The enumeration of these pathogens was carried out after different incubation periods (24 hrs, 36 hrs and 48 hrs). The standard concentration of pathogens (108/ml) was grown along with varying concentration of L. plantarum (104 to 108/ml). The mean colony forming counts of S. aureus in the controls after 24 hrs of incubation were found to be 622.66 x 108 CFU/ml and in treatment groups the counts were 2.56 x 108 CFU/ml, 2.78 x 108 CFU/ml, 2.90 x 108 CFU/ml, 3.03 x 108 CFU/ml and 39.19 x 108 CFU/ml treated with 108, 107, 106, 105 and 104/ml of L. plantarum, respectively. The mean counts of S. aureus in control groups were statistically higher compared to treatment groups. After 36 hrs of incubation the mean counts of S. aureus in different treatment groups with concentrations of L. plantarum as 108, 107, 106, 105 and 104/ml the counts were 2.04 x 108 CFU/ml, 2.44 x 108 CFU/ml, 2.39 x 108 CFU/ml, 2.97 x 108 CFU/ml and 74.60 x 108 CFU/ml, respectively. In the control group, the counts after 36 hrs of incubation were 7466.66 ´ 108 CFU/ml. Therefore, the difference in the mean counts of control group and all the treatment groups was statistically significant. The mean counts of S. aureus in the control group after 48 hrs of incubation were 902.66 x 108 CFU/ml. The mean counts of S. aureus after 48 hrs in different treatment groups with concentration of L. plantarum as 108, 107, 106, 105 and 104/ml were 0.11 x 108 CFU/ml, 0.14 x 108 CFU/ml, 0.17 x 108 CFU/ml, 0.23 x 108 CFU/ml and 3.87 x 108 CFU/ml, respectively. Again the difference in the counts of S. aureus between all the treatment groups and the control group were statistically significant.
       
For E. coli the mean colony forming units in the control group after 24 hours of incubation were 446.66 x 108 CFU/ml and were 0.81 x 108 CFU/ml, 1.09 x 108 CFU/ml, 1.36 x 108 CFU/ml, 1.85 x 108 CFU/ml and 24.62 x 108 CFU/ml, when grown with 108, 107, 106, 105 and 104/ml, of L. plantarum, respectively. The counts in the control group were significantly higher compared to all the treatment groups. After 36 hrs of incubation mean colony forming units of the control group was 5720.00 x 108 CFU/ml. The treatment groups with concentration of L. plantarum as 108, 107, 106, 105 and 104/ml, was 1.13 x 108 CFU/ml, 1.64 x 108 CFU/ml, 2.20 x 108 CFU/ml, 2.54 x 108 CFU/ml and 24.84 x 108 CFU/ml, respectively. A significant difference was there in the mean counts of E. coli between the control and treatment groups at 36 hrs of incubation. The mean colony forming counts of E. coli in control group after 48 hrs of incubation was 624.66 ´ 108 CFU/ml. The counts of E. coli in the treatment groups after 48 hrs of incubation were 0.10 x 108 CFU/ml, 0.12 x 108 CFU/ml, 0.14 x 108 CFU/ml, 0.18 x 108 CFU/ml and 2.26 x 108 CFU/ml when grown with 108, 107, 106, 105 and 104/ml of L. plantarum, respectively. The difference in the counts of treatment and control was statistically significant.
       
After 24 hrs of incubation the mean colony forming counts of B. cereus in the control group was 83.60 x 108 CFU/ml and in the treatment groups the counts of B. cereus were 0.31 x 108 CFU/ml, 0.43 x 108 CFU/ml, 0.60 x 108 CFU/ml, 0.69 x 108/ CFU/ml and 7.12 x 108 CFU/ml, when grown with L. plantarum with concentration of 108, 107, 106, 105 and 104/ml, respectively. Statistically, it was seen that the mean colony forming units of control group of B. cereus varied significantly with B. cereus treated with different concentrations of L. plantarum. Similarly, the counts of B. cereus were highest in the control group (1330 x 108 CFU/ml) compared to all the treatment groups. The counts of treatment groups with the concentration of L. plantarum as 108, 107, 106, 105 and 104/ml were 1.22 x 108 CFU/ml, 1.31 x 108 CFU/ml, 1.60 x 108 CFU/ml, 1.73 x 108 CFU/ml and 6.86 x 108 CFU/ml, respectively. All the counts of treatment groups were statistically lower compared to control groups. The counts of B. cereus in the control group after 48 hours of incubation were 1434.66 x 108 CFU/ml. In the treatment groups with different concentration of L. plantarum viz. 108, 107, 106, 105 and 104/ml the counts of B. cereus were 1.30 x 108 CFU/ml, 1.31 x 108 CFU/ml, 1.46 x 108 CFU/ml, 1.57 x 108 CFU/ml and 7.56 x 108 CFU/ml, respectively. The difference in the counts of B. cereus in the control group and all the treatment groups was statistically significant.
       
Food-borne diseases are of global concern and the World Health Organization estimated that diarrheal diseases are responsible for around 1.9 million child deaths every year (WHO, 2008). Despite improved hygiene and sanitation, there is an increased incidence of food-borne diseases around the globe and more so in developing countries. Many technologies have been employed to combat the food-borne pathogens involving physical and chemical methods, but they are with many disadvantages. These conventional preservative methods may be at the cost of food quality, for example, heat treatments are associated with deterioration of organoleptic properties, extensive use of antimicrobials have led to the development of resistant bacteria and chemical preservatives have a negative effect not only on sensory parameters but also on health as many of them are carcinogenic. Therefore, the consumers prefer organic food which is free from all sorts of chemicals and the addition of chemical preservatives prevents consumers to buy these products. Hence, there is a need for new strategies that fulfill consumer demand and ensure food safety. One such promising approach is the use of a natural antagonist towards pathogenic bacteria to control food-borne diseases as well as bacterial contamination in foods by a process called as biocontrol. The biocontrol methods may tackle the drawbacks of current processing and preservation technologies and is likely to be acceptable to consumers. One such method involves the use of beneficial bacteria to control harmful bacteria, the classical example being Lactic Acid Bacilli (LAB).
       
The main properties of beneficial/probiotic microorganisms involve equilibrating the endogenous microflora, in protecting the gut from pathogen invasion by competitive exclusion and production of antimicrobial molecules and in stimulating mucosal immunity. Of the various species of LAB, Lactobacillus plantarum is one of the most versatile species with valuable use in milk industry and recognized probiotic features (Da Silva Sabo, 2014; Guidone et al., 2014). Concurrently, because of the increasing attention of consumers for healthy and natural food the food industry is prompted towards scientific research to investigate the application of natural compounds for the processing of food products, in order to eliminate or reduce chemical additives used as antimicrobial agents. Thus, in recent decades, several lines of research have tried to find the natural solution to the chemical problem. Among these, the selection of microbial molecules or bacterial strains able to produce such compounds to be used as antimicrobials and preservatives, proved that Lactic Acid Bacteria (LAB) could be suitable candidates for biocontrol (Da Silva Sabo, 2014; Suskovi et al., 2010).          
       
The present study was carried out to study In-vitro antagonistic effect of Lactobacillus plantarum on the growth of some important food-borne pathogens viz. Staphylococcus aureus, Escherichia coli and Bacillus cereus recovered from foods of animal origin and clinical sources. The isolation and identification of these pathogens were carried out by standard microbiological protocols. The pathogenic isolates recovered from various foods of animal origins were also characterized for virulence properties involving molecular methods. The experiment was also carried out on these pathogens, in order to assess the extent of antibacterial effect of L. plantarum on the pathogenic isolates involving disc diffusion assay.
       
Staphylococcus aureus is widely distributed across the globe and is linked to an array of infections in humans and animals. The organism is profoundly known for its pathogenicity and ability to diminish the impact of antimicrobials. The organism causes infections, ranging from mild superficial skin to severe and fatal diseases. In the present study, of the 20 samples comprising of raw milk (5), mastitis milk (5), raw chicken (5) and sheep faecal samples (5) screened for isolation of S. aureus, 6 turned out positive making an overall occurrence of 30%. The results concur with the observations of Philip et al., (2006) reporting an occurrence of 31.6% from foods of animal origin. Among all the categories of the samples processed, the highest occurrence (40%) of S. aureus was found in mastitis milk samples. Similar results from mastitic milk were reported by Sharma et al., (2015) and Awad et al., (2017), with 33.7% and 42% occurrence of S. aureus, respectively. The highest recovery of S. aureus from mastitis milk samples indicates that the pathogen is one of the prime causes of mastitis in bovines in this part of the world. The reports of S. aureus being one of the leading causes of mastitis in cattle has been reported by other authors as well (Reshi et al., 2015, Wells et al., 1998). Of the raw milk samples, 2 (40%) were positive for S. aureus. The results are in agreement with the findings of Suelam et al., (2012), reporting 30% occurrence of S. aureus in raw milk samples.
 
 

CONCLUSION

From the present study it could be concluded that bio-control and bio-preservation strategies can be adopted against food-borne pathogens to cope with the problems related to the chemical preservatives and antibiotics use in animal farming and food processing. Searching and developing novel probiotics could prove better for the treatment of food-borne bacterial pathogens. This antimicrobial/antagonistic ability of probiotics can be efficiently used as antimicrobials as well as antagonist against pathogenic bacteria. This may pave way for the use of beneficial bacteria as biocontrol agents and as preservatives in food industry with immense health benefits.
 

Conflict of interest

There is no conflict of interest among the authors.
 

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