Detection and Identification of Probiotic Lactobacillus plantarum Strains by Multiplex PCR Using RAPD-Derived Primers

Lactobacillus plantarum 2035 and Lactobacillus plantarum ACA-DC 2640 are two lactic acid bacteria (LAB) strains that have been isolated from Feta cheese. Both display significant potential for the production of novel probiotic food products. The aim of the present study was the development of an accurate and efficient method for the molecular detection and identification of the above strains in a single reaction. A multiplex PCR assay was designed for each strain, based on specific primers derived from Random Amplified Polymorphic DNA (RAPD) Sequenced Characterized Amplified Region (SCAR) analysis. The specificity of the assay was tested with a total of 23 different LAB strains, for L. plantarum 2035 and L. plantarum ACA-DC 2640. The multiplex PCR assay was also successfully applied for the detection of the above cultures in yogurt samples prepared in our lab. The proposed methodology may be applied for monitoring the presence of these strains in food products, thus evaluating their probiotic character. Moreover, our strategy may be adapted for other novel LAB strains with probiotic potential, thus providing a powerful tool for molecular discrimination that could be invaluable to the food industry.     

恩替卡韦联合微生态制剂治疗乙肝肝硬化患者的临床疗效研究

目的: 观察恩替卡韦联合双歧三联活菌胶囊治疗乙型病毒性肝炎致肝硬化患者的临床应用效果。方法: 将2014年7月至2017年1月于宁波市第二医院肝病科收治的乙型肝炎病毒感染致肝硬化患者160例,按照随机数字表法分成对照组以及观察组,各80例。两组均予保肝基础治疗及恩替卡韦0.5 mg/次/d口服抗病毒治疗,观察组在此基础上加用双歧三联活菌胶囊一日2次,2~4粒/次。两组均持续治疗6个月,期间观察两组患者的临床表现、肝脏B超及肝功能、肝纤维4项、内毒素、外周血T细胞亚群水平。结果: 观察组及对照组治疗前后,丙氨酸转氨酶（ALT）、天冬氨酸转氨酶（AST）、总胆红素（TBIL）、血氨及内毒素、肝纤维化及免疫反应指标、门静脉内径及脾脏厚度均有显著改善,差异有统计学意义（P＜0.05）,而且观察组指标的改善情况优于对照组,差异有统计学意义（P＜0.05）,但两组并发症发生情况比较差异无统计学意义。结论: 恩替卡韦联合双歧三联活菌胶囊治疗能改善乙肝肝硬化患者肠道稳态,有效改善肝脏功能,减缓肝脏纤维化,临床效果显著。     

The effect of co-encapsulation of Lactobacillus rhamnosus GG ATCC 53103 with inulin on alginate/chitosan matrix: the viability in fermented soy blend and simulated digestive system

This research verified the ability of Lactobacillus rhamnosus encapsulated with inulin to tolerate the simulated digestive system and their viability in a soy blend. Probiotic encapsulated in alginate-chitosan matrix without inulin presented a better encapsulation efficiency (80.92%) than encapsulation with inulin (57.39%). On the 28th day, the count of probiotics decreased by 3.42 and 1.99 logarithmic cycles of free and encapsulated cells without inulin, respectively. In contrast, the microorganisms encapsulated with inulin showed an increase of 1.26 logs CFU g⁻¹. During gastrointestinal simulation, cell counts decreased by 0.78, 1.55 and 1.95 CFU g⁻¹ logs for encapsulated cells without inulin, free and encapsulated with inulin, respectively. Sensory panellists liked the fermented soy blend with encapsulated lactobacilli, and this result shows the possibility to create new probiotic foods of plant origin. Therefore, the alginate/chitosan matrix can be considered adequate for the encapsulation of L. rhamnosus. The inulin reduces the encapsulation efficiency and increases the cell loss in gastrointestinal simulation. Considering cellular losses, the best option for preparing a fermented soy blend is to use L. rhamnosus encapsulated without inulin.     

Organic whey as a source of Lactobacillus strains with selected technological and antimicrobial properties

Twenty‐five strains, isolated from raw, non‐pasteurised, organic whey samples, were identified phenotypically and genotypically. Biochemical tests were performed, and enzyme profiles, antibiotic resistance and antimicrobial properties were investigated. Sixteen strains were identified as genus Lactobacillus. Based on 16S rDNA gene sequence, the strains were identified as Lb. plantarum and Lb. fermentum. All of the strains had β‐galactosidase activity, and some of them reduced nitrate content. All strains utilised carbohydrates. The tested strains were characterised by low or average lipolytic and esterolytic activity. Moreover, the strains showed low proteolytic activity which is advantageous for their use as starter cultures for foods with low protein content. Strains Lb. fermentum S20, SM1, SM3, S2R and Lb. plantarum SM5 produced harmful N‐acetyl‐β‐glucosaminidase; moreover, the strain S20 produced also β‐glucuronidase. None of the strains produced α‐chymotrypsin. In phenotypic studies, most of the test strains were susceptible to gentamicin, ampicillin, tetracycline, chloramphenicol, penicillin and erythromycin. Strains Lb. plantarum S1 and Lb. fermentum S4, S7, S8, S10, SM1 and SM3 did not possess any transfer resistance genes. Antagonistic activity of the culture LAB strains was assessed as high or moderate in relation to the indicator strains, with the greatest zones of inhibition for E.coli and the smallest for L. monocytogenes ATCC 15313. This study reveals that the LAB strains isolated from organic whey have high potential for food application. Some strains of species Lb. fermentum (S4, S7, S8, S10) have been identified as the best candidates.     

酸奶粉产品稳定性研究

为确定产品的保质期,研究产品在加速试验、常温保存试验及低温保存试验的活菌数的变化及对发酵酸奶的影响.结果表明:经过3个月的加速试验,样品的活菌数下降3个数量级,发酵酸奶不成功.在常温试验中.活菌数下降1个数量级,样品发酵酸奶质量略有降低.而低温下,经过18个月保存活菌数才下降1个数量级,发酵酸奶的影响不大.初步评价产品在常温下的保质期为5个月～6个月.对于舍有益生菌的产品建议采用低温保存及冷链运输.     

Comparison of vacuum impregnation and soaking techniques for addition of the probiotic Lactobacillus acidophilus to minimally processed melon

This study aimed to evaluate the vacuum impregnation (VI) and soaking methods in the addition of Lactobacillus acidophilusLA‐3 to minimally processed melon (MPM). The melons were washed, sanitised in chlorine solution (200 mg L^?1), peeled and cutted into cubes. Lactobacillus acidophilusLA‐3 (1.4 × 10¹⁰ CFU g^?1) were added to the MPM through both techniques. The L. acidophilusLA‐3 count in MPM was similar to those commonly found in dairy products having probiotic claim, but VI was more efficient than soaking in maintaining the viability (8.61 and 7.98 Log CFU g^?1, respectively). The pH, acidity and soluble solids were not affected by probiotic culture and the incorporation technique; however, the VI affected the firmness of fruit. The MPM was within Brazilian standards for their microbiological characteristics. MPM may be used as a carrier of probiotic bacteria, being one more alternative for individuals who consume probiotic products.     

Recent advances on lactose intolerance: Tolerance thresholds and currently available answers

The genetically programmed reduction in lactase activity during adulthood affects 70% of the world adult population and can cause severe digestive disorders, which are the sign of lactose intolerance. Lactose intolerance symptoms vary depending on the residual lactase activity, the small bowel transit time, and especially the amount of ingested lactose. To formulate dairy products suitable for the vast majority of lactose intolerants, it is essential to define lactose intolerance threshold. A recent meta-analysis permitted to show that almost all lactose intolerants tolerate 12 g of lactose in one intake and approximately 18 g of lactose spread over the day. The prevalence and severity of lactose intolerance are probably overestimated by the general public. This misconception usually leads to an unnecessary reduction of dairy foodstuff consumption. Nevertheless, dairy products are essential for health mainly due to their calcium content and the positive influence of probiotic bacteria. The formulation of dairy products suitable for most intolerant and suspicious subjects seems necessary. The use of exogenous enzyme preparations, as well as the consumption of lactose-free products or products rich in probiotic bacteria are proposed as symptom-reducing strategies.     

Preparation and properties of probiotic concentrated yoghurt (labneh) fortified with conjugated linoleic acid

Milk of high conjugated linoleic acid (CLA) level (1.25 g per 100 g milk fat) was produced by inclusion of fish oil and rousted soy bean in the ration of Holstein cows as compared to 0.55 g per 100 g milk fat in the milk of animals receiving control diet. Milk of normal (control) and high CLA content (treatment) was spray‐dried. Labneh was made from 20 g L^?1 reconstituted milk using 3 mL per 100 mL yoghurt starter and 2 mL per 100 mL of probiotic cultures of Lactobacillus casei or Lactobacillus acidophilus. The control (C) and high CLA (T) labneh were analysed chemically and microbiologically, and their viscosities were determined during cold storage for 15 days. The fat content of labneh of high CLA was less than that of the control, but the total solids (TS) were unaffected by the CLA level. Labneh made with Lb. acidophilus had lower TS and higher acidity, exopolysaccharides and acetaldehyde contents and viscosity than that made with the use of Lb. casei. Labneh from the different treatments retained high counts of the added probiotic (>10⁸ cfu g^?1) throughout the storage period. The storage period had significant effects on all parameters determined.     

Oligosaccharides prepared by acid hydrolysis of polysaccharides from pumpkin (Cucurbita moschata) pulp and their prebiotic activities

As a rich source of polysaccharides, pumpkin pulp was used for the production of oligosaccharides. To investigate the prebiotic potential of the oligosaccharides, prepared by acid hydrolysis of polysaccharides from pumpkin pulp, the effect of the oligosaccharides on lactobacilli growth was studied. The oligosaccharides were composed of galactose (99.03%) and glucose (0.97%). The molecular weight distribution of the oligosaccharides showed a larger part of low molecular weight fraction in a range from 5000 to 324 Da than the polysaccharides, which gave the oligosaccharides higher solubility. The oligosaccharides showed good resistance to hydrolysis by artificial human gastric juice and α‐amylase, compared to the reference prebiotic inulin, and considerable capability to stimulate the growth of lactobacilli. In vitro studies showed that the number of lactobacilli was significantly greater than those in control medium when they were cultured in the medium supplemented with the oligosaccharides (P < 0.05), and there was no significant difference at the oligosaccharides group and the inulin group (P < 0.05).