Aims: This study aimed to evaluate the effect of electroporation on the growth characteristics and antimicrobial activity of lactic acid bacteria (LAB) including Bifidobacterium longum ATCC 15707, Lactobacillus acidophilus ATCC 314, Lactobacillus casei ATCC 393 and Lactobacillus fermentum ATCC 14931. Methodology and results: Electroporation with the strength of electric field at 1.0–3.0 kV/cm for 2-4 millisecond were applied on the bacterial cultures. All bacterial cultures showed significant (P<0.05) increased in cell viability (40%-325%) upon electroporation. Such treatment also increased the acidity of the cell where the pH of cells decreased upon treatment. In tandem with the increased viability, electroporated bacterial cultures also showed higher proteolytic activity compared to the control (P<0.05). The electroporation treatment also increased (P<0.05) the bacteriocin activity of treated cells compared to the control. However, the molecular weight of bacteriocins produced were not affected by electroporation. Treated cells also possessed better antimicrobial activity. According to the results collected, all treated LAB strains showed 11.5%-113.8% higher (P<0.05) inhibitory activity compared to untreated control against tested pathogenic bacteria, Escherichia coli and Listeria monocytogenes that commonly associated with food contamination. Microarray data analysis showed that electroporation regulated the entities encoding for surface protein and transporter. Conclusion, significance and impact of study The results from this study suggested that electroporation could enhance the growth characteristics and antimicrobial activity of LAB by modifying the surface regions of the cells. This result may serve as the reference for food manufacturers to opt for effective biopreservation method and produce food with extended shelf life.
Electroporation ; Lactobacillales

Aims: The aim of this study was to screen lactic acid bacteria (LAB) isolates with probiotic properties for tilapia (Oreochromis niloticus) from the intestines of mackerel and to analyze its ability to produce hydrolase enzymes (amylase, protease, cellulase and lipase). Methodology and results: Ten samples of mackerel were collected from the Fish Auction Place, Lewoleba, Nubatukan District, Lembata Regency, East Nusa Tenggara Province, Indonesia. The process for screening were antimicrobial activity, pH tolerance test of 1, 3 and 5, bile salt concentrations of 2.5%, 5% and 7.5%, and water salinity (0.5%, 3.5% and 6.5%). The autoaggregation and semi-qualitative test of hydrolase enzymes were also carried out. In total, 21 LAB isolates were derived from mackerel intestines. Isolates that have the stronger ability to inhibit the pathogen Aeromonas hydrophila ATCC 7966 are KBP 3.3, KBP 3.3.1 and KBP 6.3, while KBP 1.3, KBP 1.1.1 and KBP 3.3.1 were able to inhibit Streptococcus agalactiae ATCC 13813. The LAB isolates that survived at the tested pH were KBP 3.3.1, KBP 3.3, KBP 1.1.1 and KBP 1.3. Results also suggest that the four isolates were tolerant to bile salt concentrations and water salinity with good survival rates (>94.7%) and had proteolytic, lipolytic and cellulolytic activities. The isolates KBP 3.3, KBP 3.3.1 and KBP 1.1.1 were identified as Weissella confusa 6250, while the isolate KBP 1.3 was identified as W. confusa C5-7. Conclusion, significance and impact of study The characteristics of four selected isolates indicate their potential as a probiotic. These potential probiotic isolates can be applied directly to fish by utilizing microencapsulation technology or adding to the feed.
Lactobacillales ; Perciformes ; Indonesia

Aims: The aim of this study was to screen lactic acid bacteria (LAB) isolates from fermented Sumbawa mare‘s milk that meet the requirements as starter cultures, and to evaluate the effect of the selected starter culture in improving the organoleptic quality of mare‘s milk fermentation. Methodology and results: The LAB isolates (13 isolates) derived from naturally fermented Sumbawa mare‘s milk were firstly screened for acidification activity. Afterwards, the selected isolates were evaluated for the starter culture criteria such as technological properties (proteolytic test, lipolytic test, and exopolysaccharide production), food safety test (hemolytic test and antibiotic sensitivity test), antimicrobial activity test. The selected culture (SC) together with yogurt starter cultures (YC) and combination between the selected isolate and a mixture of both (MC) were used to ferment fresh mare’s milk. Six LAB isolates (DB7, BC10, DC4, BC9, DC10, and BC7) were obtained from the acidification screening. Isolate BC10 was the most potential isolate as starter culture due to its ability in terms of acidification and proteolytic activity, lack of lipolytic activity, no indication of pathogenic potency, as well as able to inhibit the growth of Escherichia coli ATCC 25922. However, this isolate was resistant to antibiotics kanamycin, trimethoprim, and cinoxacin. The isolate BC10 presented 99.99% sequence similarity with respect to Lactobacillus plantarum. Conclusion, significance and impact of study The selected starter culture (isolate BC10) was able to improve the organoleptic quality of fermented mare‘s milk especially aroma compared to the other starter cultures. Therefore, L. plantarum BC10 is a potential isolate to be used as starter culture for mare’s milk fermentation.
Lactobacillales--isolation & ; purification

Aims: Researchers are focusing more on the isolation of new probiotic bacteria to increase varieties for the growing market demand. This study aimed to isolate lactic acid bacteria (LAB) strains from sugarcane waste materials and evaluate its characteristic. Methodology and results: In the present study, two strains of LAB (Isolates A and B) were isolated from sugarcane waste and investigated in vitro for their characteristics as potential probiotics. These isolates were evaluated on their characteristics based on four biochemical tests (acid tolerance, bile tolerance, microbial adhesion, and phenol resistance), with the commercial strain Lactobacillus isolated from Yakult ® served as a positive control. Both isolated strains (>8 log10 CFU/mL) displayed higher survivability than control (>6 log10 CFU/mL) in simulated gastrointestinal conditions at pH 2.0 and pH 6.9 after 24 h. Furthermore, both isolated LABs were resistant to inhibitory substances which are 0.05-0.3% bile and 0.4% phenol. For bile tolerance, isolate A (OD 6.83) had a higher absorbance at 0.3% bile concentration as compared to isolate B (OD 2.20). However, isolate B (7.49 log10 CFU/mL) showed higher resistance towards 0.4% phenol than isolate A (7.11 log10 CFU/mL) after 24 h. Both isolate A and isolate B displayed low cell surface hydrophobicity, strong electron donor, and basic characteristic. Conclusion, significance and impact of study Both isolates were able to survive under gastrointestinal stress conditions, implying their potential as probiotics. This study demonstrated that valuable products such as probiotic strain could be isolated from sugarcane wastes to use in food production or medical treatment.
Lactobacillales ; Waste Management

Aims: Lactic acid bacteria (LAB) biofilms constitute one of the most remarkable breakthroughs in the field of food biopreservatives and can be employed to prevent foodborne disease. The purposes of this study were to investigate the efficacy of inhibitory LAB biofilms against foodborne pathogens and evaluate their tolerance to acidic pH and bile salts, as well as their physicochemical properties. Methodology and results: Four strains of Lactobacillus brevis biofilms isolated from kimchi showed antipathogenic activity to the bacteria Staphylococcus aureus FNCC 0049 and Escherichia coli FNCC 0091. These biofilms were also tolerant to pH 2.5, 0.3% bile salt and strong adhesion. Two of the four L. brevis biofilms (L. brevis biofilm KA2 and KB1) produced the highest inhibitory activity against both pathogenic bacterial indicators, tolerance to acidic pH and bile salts, and the strongest adhesion. In addition, based on Scanning Electron Microscope-Energy Dispersion X-ray Spectroscopy (SEM-EDS) analysis, both biofilm strains had a smooth surface texture; the cell morphology was rod-shaped and consisted of several elements such as carbon, oxygen and nitrogen, which was built up of extracellular polymeric substances (EPS). Conclusion, significance and impact of study The presence of EPS as a constituent of LAB biofilms influenced their survival abilities in an acidic pH and bile salt environment. As a result, the characteristics of L. brevis biofilm KA2 and KB1 made them excellent candidates for use as antimicrobial packaging systems in food biopreservative applications.
Levilactobacillus brevis ; Lactobacillales ; Foodborne Diseases

Aims: To isolate and characterize the lactic acid bacteria (LAB) strains from the “mam chua ca ro” (sour fermented fish) in the South of Vietnam and investigate their potential anti-bacterial properties. Methodology and results: Four LAB strains (MCR1, MCR2, MCR3 and MCR4) were isolated from the "mam chua ca ro" product and their anti-bacterial activity was determined using the spot assay and the paper disc diffusion method. The isolated LABs can inhibit Escherichia coli ATCC 25922, Staphyloccocus aureus ATCC 25923 and Vibrio parahaemolyticus BV016 and produce bacteriocin to control the growth of E. coli ATCC 25922 and S. aureus ATCC 25923, except V. parahaemolyticus. MCR2 was chosen to sequence 16S rRNA of Pediococcus acidilactic. Conclusion, significance and impact of study On the basis of their prominent anti-pathogenic bacteria activity, LAB strains isolated from Vietnamese sour-fermented fish products were verified as prospective probiotics.
Lactobacillales--isolation & ; purification ; Pediococcus acidilactici

Aims: This study aims to isolate lactic acid bacteria (LAB) from various food sources to obtain a potent strain against Listeria monocytogenes. Methodology and results: A total of 68 LAB isolates were selected to evaluate their antimicrobial activity against L. monocytogenes, a foodborne pathogen and a causative agent of listeriosis. The selected isolate was identified and characterized. The isolate C23 from cabbage showed the highest antimicrobial activity against L. monocytogenes ATCC 7644 with inhibition ability of 73.94%. The isolate was closely related to Lactobacillus brevis by 16S rRNA sequencing and subsequently deposited in GenBank with an accession number of MN880215, named as L. brevis C23. The cell free supernatant (CFS) of L. brevis C23 had high tolerance in low pH and was able to withstand up to 60 °C. The proteinaceous nature of the antimicrobial agent was also confirmed through the enzymatic test. The CFS was stable on different detergents as well as bile salts. Under transmission electron microscopy (TEM), the inhibitory effect of CFS against L. monocytogenes was proven by causing cell lysis. Conclusion, significance and impact of study Bacteriocin-like inhibitory substances (BLIS) of L. brevis C23 showed very promising potential in food industrial application.
Lactobacillales ; Listeria monocytogenes ; Foodborne Diseases ; Sprains and Strains

Aims: This study aimed to isolate and identify lactobacilli strains that have antagonistic activity against multidrug resistant (MDR) isolates of Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa and evaluate their probiotic properties. Methodology and results: Twenty-one Lactobacillus isolates were collected, and their antimicrobial activity was assessed by agar well diffusion, broth microdilution and time-kill test. The probiotic potential of the isolates was evaluated as well. The bacterial culture and cell free supernatant (CFS) of all isolates exhibited antibacterial activity against all MDR isolates. Out of 21 isolates, 4 isolates (A31, B35, S20 and S25) displayed the highest antimicrobial activity and further evaluated. The minimum inhibitory percentages of CFS from selected isolates against pathogens ranged from 10 to 30% and the bactericidal percentages ranged from 20 to 50%. The inhibitory activity of CFS was not changed after heating but abrogated as the pH neutralized. The growth kinetic of the MDR pathogens was significantly reduced in the presence of the CFS of all isolates. The isolates had a less than 1-log reduction in their viability in acid tolerance test and could grow in the presence of 0.3% bile salts. Strains S20, S25 and B35 exhibited high coaggregation with E. coli (51.7-73.3%), P. aeruginosa (53.7-69.3%) and K. pneumoniae (49.7-65.3%). Molecular identification revealed that the isolates were Lactobacillus rhamnosus (B35) and Lactobacillus paracasei (S20, S25, A31). Conclusion, significance and impact of study The results suggest that these lactobacilli isolates may have potential applications for controlling and preventing colonization of infections caused by MDR pathogens.
Lactobacillales ; Kefir ; Probiotics ; Gram-Negative Bacteria

Lactic acid bacteria (LAB) are generally recognized as safe food-grade microorganisms and are widely used in food production, preservation, and as probiotics to promote human health. Given the need to develop effective drug delivery strategies, LAB have become attractive live vehicles for the oral, intranasal and vaginal delivery of therapeutic molecules. Being live and safe organisms, LAB are able to directly produce and deliver target proteins for therapeutic purpose, which remarkably reduces the cost for drug production. To date, LAB have been used to deliver a variety of functional proteins to mucosal tissues for the treatment of various diseases. This review summarized the development and application of LAB as mucosal delivery vectors in the last 20 years to provide references for future clinical research.
Drug Delivery Systems ; Humans ; Lactobacillales ; Mucous Membrane ; Probiotics ; Proteins

Aims: The menace of antibiotic resistance has led to the search for alternatives, which in turn has diverted the attention to bacteriocins, antimicrobial peptides (AMPs) produced by bacteria for their bactericidal properties. The aim of our study was to isolate and partially purify bacteriocin from lactic acid bacteria (LAB) and comparing its antimicrobial activity with antibiotics. Methodology and results: Among 38 LAB screened using agar spot assay, LAB 28D1 showed the highest antimicrobial activity against the test bacterial strains. The proteinaceous nature of the antimicrobial compound extracted from LAB 28D1 was confirmed by its inactivation after treatment with proteolytic enzymes. The crude bacteriocin was found to be stable over a wide range of temperatures (60-100 °C) and pH (4-9). The bacteriocin was partially purified by ammonium sulfate precipitation (ASP) and the activity units were 204,800 AU/mL. The total protein and the specific activity of partially purified bacteriocin were found to be 24.585 mg and 124,954.24 AU/mg respectively. The molecular weight of partially purified bacteriocin was determined to be 8.5 kDa approximately. The efficacy of the partially purified bacteriocin against indicator bacterial strains was compared with antibiotics by the disc diffusion method and minimum inhibitory concentration (MIC). According to our study, the hospital waste isolate Enterococcus spp. was found to be multidrug-resistant (MDR) but sensitive to bacteriocin from LAB (MIC 0.06 ± 0 µg/mL). Conclusion, significance and impact of the study Bacteriocin from LAB has potential in combating MDR enterococcal infections.
Bacteriocins--isolation & ; purification ; Lactobacillales ; Drug Resistance, Microbial