Safety Assessment of Two New Lactobacillus Plantarum Strains Isolated from Traditional Fermented Food of Northeast China
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Abstract: This study aimed to evaluate the safety of previously isolated Lactobacillus strains (L. Plantarum L12 and L20) by in vitro and in vivo methods. Antibiotic sensitivity, bile salt hydrolysis activity, D-lactic acid content, and hemolytic activity were analyzed in vitro. Then, a short-term oral toxicity study was conducted in mice, which received 0.2 mL (109 CFU/mL) of experimental bacterial solutions orally for 30 days. The results showed that both strains had good antioxidant activity, high sensitivity to antibiotics, high bile salt hydrolase activity, low D-lactic acid content, and gamma hemolysis. Intragastric administration had no adverse effects on growth, behavior, food intake, organ weight, or histopathological analysis of the treated mice. Compared with control mice, the serum malondialdehyde concentration and liver glutathione content of probiotic-treated mice were statistically different (P<0.05). These results suggest that strains L12 and L20 are nonpathogenic and likely to be safe for human consumption.
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Keywords:
- probiotics /
- safety /
- oral toxicity /
- Lactobacillus plantarum /
- mice
摘要: 针对已分离的植物乳杆菌L12和L20通过体外和体内两种方法进行安全性评价。通过体外实验测定乳酸水解活性抗生素敏感性，胆盐水解酶活性，D-乳酸含量，γ溶血等指标；体内实验是对小鼠进行短期口服毒性的研究，口服0.2 mL（109 CFU/mL）的实验性细菌溶液30 d。结果表明，体外实验中，两株菌株均具有良好的抗氧化活性，对抗生素敏感性高，胆盐水解酶活性高，D-乳酸含量低，γ溶血；体内实验中灌胃给药对治疗小鼠的生长、行为、摄食量、器官重量或组织病理学分析均无不良影响。与对照组相比，益生菌组小鼠血清MDA浓度和肝脏GSH含量差异有统计学意义(P<0.05)。这些结果表明，菌株L12和L20是非致病性的，人直接食用是安全的。 -
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Figure 3. Photomicrographs of liver paraffin section of mice (a) control (b) L12, and (c) L20
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Figure 4. Serum MDA concentration (a) and total Liver GSH concentration (b) in control and probiotic-treated mice
Note: Values are means with standard deviations shown by vertical bars (n=4). Bars with a different lowercase letter (s) in the same treatments indicate significant differences at P<0.05 among the treatments.
Table 1 Antibiotic susceptibility of Lactobacillus strains L12 and L20
Antibiotic L12 L20 Diameters (mm) Sensitivity Diameters (mm) Sensitivity Rifampin 22±0.2g S 31±0.3b S Vancomycin 15±0.1j I 8.3±0.03j R Kanamycin 7±0.5ll R 27±0.4d S Ceftazidime 30±0.1c S 14±0.05i I Sulphamethoxazole/Trimethoprim 30±0.02c S 17±0.2h I Erythromycin 32±0.04b S 27±0.03d S Ampicillin 24±0.07f S 26±0.05e S Penicillin G 33±0.0a S 31±0.2b S Streptomycin 26±0.03e S 21±0.1g S Gentamicin 29±0.04d S 29±0.03c S Tetracycline 16±0.04i I 32±0.1a S Cefotaxime 2.9±0.05m R 23±0.2f S Ciprofloxacin 20±0.2h S 2.3±0.03k R Norfloxacin 14±0.01k S 1.6±0.6l R Note: The different lowercase letters in the table indicate a significant difference (P<0.05);R=is low -sensitive or resistant, S=is highly sensitive, I=is moderately sensitive. 
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Table 2 Test results of D-lactic acid content and bile salt hydrolase activity of strains
Strain Bile salt hydrolase activity D-lactic acid content Absorbance values Cholecystolytic enzyme (μmol/L) Absorbance values D-lactic acid concentration (μmol/L) L12 43.51±0.25a 227.32±1.89b 4.99±0.25a 18.87±15.36b L20 59.78±0.38b 245.92±2.0a 6.47±0.38a 21.53±20.25a Control 0.11±0.01b 0.62±2.20c Note: The different lowercase letters in the table indicate a significant difference (P<0.05).
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Table 3 Treatment-related sign of toxicity during the 30-days study period
Organ system Observed & inspected items Performance after gavage Central nervous system Behaviour No abnormal calls, no abnormal posture and repeated scratching Neuromuscular system Action Normal movement, no movement disorder Autonomic nervous system Pupil size Normal, no reduction or expansion Secretion No salivation, no tears Respiratory system Nostrils No flow Respiration rate Normal breathing Digestive system Belly type Abdominal type is normal Fecal shape and color Stool formation, dark yellow Urogenital system Perineum No secretions Mammary gland No swelling Skin and coat Color, tension Normal, no sagging, no rash Completeness No hair removal, no erect Eye Eyelid No upper eyelid droop Eyeball No protrusion, no corneal opacity, no congestion of the conjunctiva Transparency Not turbid
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Table 4 Organ relative weights of control and probiotic-treated mice
Strain Concentration (CFU/mL) Gavage dose (mL) Visceral index (organ-body ratio) Spleen Heart Liver Kidney L12 109 0.2 0.008±0.00028 0.008±0.00005 0.05±0.0022 0.06±0.001 L20 109 0.2 0.009±0.0003 0.008±0.00025 0.06±0.004 0.06±0.003 Control 109 0.2 0.009±0.0005 0.008±0.00022 0.06±0.003 0.06±0.0013 Note: There is no significant difference between the experimental group and the control group (P>0.05).
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[1] An Y, Wang Y, Liang X, et al. Purification and partial characterization of M1-UVs300, a novel bacteriocin produced by Lactobacillus plantarum isolated from fermented sausage[J]. Food Control,2017:S0956713517302724.
[2] Gibson Glenn R, Fuller Roy. Aspects of in vitro and in vivo research approaches directed toward identifying probiotics and prebiotics for human use [J]. The Journal of Nutrition,2000,130(2):391s−395s. doi: 10.1093/jn/130.2.391S
[3] Belicová A, Mikulášová M, Dušinský R. Probiotic potential and safety properties of Lactobacillus plantarum from slovak bryndza cheese[J]. Biomed Research International,2013(2013-9-4):760298.
[4] Berbegal C, Peña N, Russo P, et al. Technological properties of Lactobacillus plantarum strains isolated from grape must fermentation[J]. Food Microbiology,2016,57(aug.):187−194.
[5] Abushelaibi A, Al-Mahadin S, El-Tarabily K, et al. Characterization of potential probiotic lactic acid bacteria isolated from camel milk[J]. LWT - Food Science and Technology,2017,79:316−325. doi: 10.1016/j.lwt.2017.01.041
[6] Jiang M, Zhang F, Wan C, et al. Evaluation of probiotic properties of Lactobacillus plantarum WLPL04 isolated from human breast milk[J]. Journal of Dairy Science,2016,99(3):1736−1746. doi: 10.3168/jds.2015-10434
[7] R Azat, Y Liu, LI Wei, et al. Probiotic properties of lactic acid bacteria isolated from traditionally fermented Xinjiang cheese[J]. Journal - Zhejiang University Science B,2016,17(8):597.
[8] Donohue D C, Salminen S. Safety of probiotic bacteria[J]. Asia Pacific Journal of Clinical Nutrition,1996,5(1):25−8.
[9] Ren D Y, Yan W, Bin A N, et al. Screening and in vitro tolerance analysis of Lactobacillus plantarum with high antioxidant activity in traditional fermented food of northeast China[J]. Science and Technology of Food Industry, 2019.
[10] Savardi, Mattia, Ferrari, et al. Automatic hemolysis identification on aligned dual-lighting images of cultured blood agar plates[J]. Computer Methods & Programs in Biomedicine,2018,156(13−24).
[11] Collignon P C, Conly J M, Antoine A, et al. World health organization ranking of antimicrobials according to their importance in human medicine: A critical step for developing risk management strategies to control antimicrobial resistance from food animal production[J]. Clinical Infectious Diseases,2016(8):8.
[12] Maheshwari S U, Amutha S, Anandham R, et al. In vitro evaluation of antimicrobial activity of lactic acid bacteria isolated from fermented fruit mix - indian traditional fermented foods, against selected food borne pathogens[J]. International Journal of Current Microbiology and Applied Sciences,2019,8(1):3122−3126. doi: 10.20546/ijcmas.2019.801.333
[13] Khan I, Kang S C. Probiotic potential of nutritionally improved Lactobacillus plantarum DGK-17 isolated from Kimchi – A traditional Korean fermented food[J]. Food Control,2016,60:88−94. doi: 10.1016/j.foodcont.2015.07.010
[14] Dixt G, Samarth D, Tale V, et al. Comparative studies on potential probiotic characteristics of Lactobacillus acidophilus strains[J]. EurAsian Journal of Biosciences, 2013, 7: 1-9.
[15] Kim J M, Guo X H, Nam H M, et al. Screening lactic acid bacteria from swine origins for multistrain probiotics based on in vitro functional properties[J]. Anaerobe, 2010, 16(4): 321-326.
[16] Amraii H N, Abtahi H, Jafari P, et al. In vitro study of potentially probiotic lactic acid bacteria strains isolated from traditional dairy products[J]. Jundishapur Journal of Microbiology,2014,7(6).
[17] Lee M, Song J H, Lee S H, et al. Effect of seasonal production on bacterial communities in Korean industrial kimchi fermentation[J]. Food Control,2018:S0956713518301865.
[18] Honi U, F Sabrin, Islam T, et al. Enzymatic activity and antibiotic resistance profile of Lactobacillus paracasei ssp. paracasei-1 isolated from regional yogurts of bangladesh[J]. Journal of Microbiology, Biotechnology and Food Sciences,2013,3(3):235−239.
[19] Bustos A Y, Saavedra L, Valdez G, et al. Relationship between bile salt hydrolase activity, changes in the internal pH and tolerance to bile acids in lactic acid bacteria[J]. Biotechnology Letters,2012,34(8):1511−1518. doi: 10.1007/s10529-012-0932-5
[20] Pohanka M. D-lactic acid as a metabolite: Toxicology, diagnosis, and detection[J]. BioMed Research International,2020,2020(2):1−9.
[21] Seddik H A, Bendali F, Gancel F, et al. Lactobacillus plantarum and its probiotic and food potentialities[J]. Probiotics & Antimicrobial Proteins,2017,9(2):111−122.
[22] Huseby M, Shi K, Brown C K, et al. Structure and biological activities of beta toxin fromStaphylococcus aureus[J]. Journal of Bacteriology,2007,189(23):8719−8726. doi: 10.1128/JB.00741-07
[23] Kaktcham P M, Zambou N F, Tchouanguep F M, et al. Antimicrobial and safety properties of Lactobacilli isolated from two cameroonian traditional fermented foods[J]. Scientia Pharmaceutica,2012,80(1):189−203.
[24] Debashis H, Manisha M, Shiv C, et al. Indigenous probiotic Lactobacillus isolates presenting antibiotic like activity against human pathogenic bacteria[J]. Biomedicines,2017,5(4):31. doi: 10.3390/biomedicines5020031
[25] Saharan B, Chauhary A. Probiotic properties of Lactobacillus plantarum[J]. Journal of Pure & Applied Microbiology,2019,13(2):933−48.
[26] Abotsi W, Ainooson G K, Boakye-Gyasi E. Acute and sub-acute toxicity studies of the ethanolic extract of the aerial parts of hilleria Latifolia (Lam.) H. Walt. (Phytolaccaceae) in rodents[J]. West African Journal of Pharmacy , 2011, 22 (1) 27 - 35.
[27] Shokryazdan P, Jahromi M F, Liang J B, et al. Safety assessment of two new Lactobacillus Strains as probiotic for human using a rat model[J]. PLoS One,2016,11(7):e0159851. doi: 10.1371/journal.pone.0159851
[28] Ding W, Shi C, Chen M, et al. Screening for lactic acid bacteria in traditional fermented Tibetan yak milk and evaluating their probiotic and cholesterol-lowering potentials in rats fed a high-cholesterol diet[J]. Journal of Functional Foods,2017,32:324−332. doi: 10.1016/j.jff.2017.03.021
[29] Adnan M T, Amin M N, Uddin M G, et al. Increased concentration of serum MDA, decreased antioxidants and altered trace elements and macro-minerals are linked to obesity among Bangladeshi population[J]. Diabetes & Metabolic Syndrome: Clinical Research & Reviews,2019,13(2):933−938.
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1. 潘玲，冮洁，薛晨光，杨新鹏. 东北少数民族特色发酵食品研究进展. 大连民族大学学报. 2022(01): 8-11+17 . 
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