Allergen degradation of bee pollen by lactic acid bacteria fermentation and its alleviatory effects on allergic reactions in BALB/c mice

Food allergy as a global health problem threatens food industry.Bee pollen(BP)is a typical food with allergenic potentials,although it performs various nutritional/pharmacological functions to humans.In this study,lactic acid bacteria(LAB)were used to ferment Brassica napus BP for alleviating its allergenicity.Four novel allergens(glutaredoxin,oleosin-B2,catalase and lipase)were identified with significant decreases in LAB-fermented BP(FBP)than natural BP by proteomics.Meanwhile,metabolomics analysis showed significant increases of 28 characteristic oligopeptides and amino acids in FBP versus BP,indicating the degradation of LAB on allergens.Moreover,FBP showed alleviatory effects in BALB/c mice,which relieved pathological symptoms and lowered production of allergic mediators.Microbial high-throughput sequencing analysis showed that FBP could regulate gut microbiota and metabolism to strengthen immunity,which were closely correlated with the alleviation of allergic reactivity.These findings could contribute to the development and utilization of hypoallergenic BP products.

Preparation of lactic acid bacteria compound starter cultures based on pasting properties and its improvement of glutinous rice flour and dough

The effects of 5 lactic acid bacteria(LAB)fermentation on the pasting properties of glutinous rice flour were compared,and suitable fermentation strains were selected based on the changes of viscosity,setback value,and breakdown value to prepare LAB compound starter cultures.The results revealed that Latilactobacillus sakei HSD004 and Lacticaseibacillus rhamnosus HSD005 had apparent advantages in increasing the viscosity and reducing the setback and breakdown values of glutinous rice flour.In particular,the compound starter created using the two abovementioned LAB in the ratio of 3:1 had better performance than that using a single LAB in improving the pasting properties and increasing the water and oil absorption capacity of glutinous rice flour.Moreover,the gelatinization enthalpy of the fermented samples increased significantly.For frozen glutinous rice dough stored for 28 days,the viscoelasticity of frozen dough prepared by compound starter was better than that of control dough,and the freezable water content was lower than that of control dough.These results indicate that compound LAB fermentation is a promising technology in the glutinous rice-based food processing industry,which has significance for its application.

Screening of glucosinolates degrading lactic acid bacteria and their utilization in rapeseed meal fermentation

Rapeseed meal is a promising food ingredient, but its utilization is limited by the presence of some potentially harmful ingredients, such as glucosinolates. Fermentation is a cost-effective method of detoxication but a food-grade starter culture with glucosinolates degradation capacity is required. In this study, 46 strains of lactic acid bacteria from traditional paocai brines were screened for their ability to glucosinolate degradation. The results showed that more than 50% of the strains significantly degraded glucosinolates. Two strains of Lactiplantibacillus(p7 and s7) with high capacity of glucosinolates degradation through producing enzymes were identified. Then,an optimized condition for rapeseed meal fermentation by p7 was established to degrade glucosinolates, which can achieve about 80% degradation. UPLC/Q-TOF-MS analysis showed that the degradation rate of individual glucosinolates was different and the degradation rate of gluconapin and progoitrin in rapeseed meal can reach more than 90%. Meanwhile, fermentation with p7 can improve safety of rapeseed meal by inhibiting the growth of Enterobacteriaceae and improve its nutritional properties by degrading phytic acid. The in vitro digestion experiments showed that the content of glucosinolates in rapeseed meal decreased significantly during gastric digestion. Meanwhile, fermentation with p7 can greatly improve the release of soluble protein and increase the contents of free essential amino acids, such as lysine(increased by 12 folds) and methionine(increased by 10 folds).

Application of Next Generation Sequencing for Rapid Identification of Lactic Acid Bacteria

The rapid identification of lactic acid bacteria,which are essential microorganisms in the food industry,is of great significance for industrial applications.The identification of lactic acid bacteria traditionally relies on the isolation and identification of pure colonies.While this method is well-established and widely used,it is not without limitations.The subjective judgment inherent in the isolation and purification process introduces potential for error,and the incomplete nature of the isolation process can result in the loss of valuable information.The advent of next generation sequencing has provided a novel approach to the rapid identification of lactic acid bacteria.This technology offers several advantages,including rapidity,accuracy,high throughput,and low cost.Next generation sequencing represents a significant advancement in the field of DNA sequencing.Its ability to rapidly and accurately identify lactic acid bacteria strains in samples with insufficient information or in the presence of multiple lactic acid bacteria sets it apart as a valuable tool.The application of this technology not only circumvents the potential errors inherent in the traditional method but also provides a robust foundation for the expeditious identification of lactic acid bacteria strains and the authentication of bacterial powder in industrial applications.This paper commences with an overview of traditional and molecular biology methods for the identification of lactic acid bacteria.While each method has its own advantages,they are not without limitations in practical application.Subsequently,the paper provides an introduction of the principle,process,advantages,and disadvantages of next generation sequencing,and also details its application in strain identification and rapid identification of lactic acid bacteria.The objective of this study is to provide a comprehensive and reliable basis for the rapid identification of industrial lactic acid bacteria strains and the authenticity identification of bacterial powder.

3D printed lactic acid bacteria hydrogel:cell release kinetics and stability

In this study,a new type of 3 D printed living biological hydrogel was developed by integrating lactic acid bacteria(LAB)into biocompatible and non-toxic polymer materials.Interestingly,the living materials loaded with LAB can be freeze-dried and reused for more than 100 times.The bio-hydrogel can be used to co-culture different LAB and keep its fermentation performance stable in long-term use.The release kinetics model and response surface method were used to simulate and optimize the bacteria release mode in the bio-hydrogel.The results show that the release of bacteria from hydrogel is regulated by the coupling of Fickian diffusion and polymer swelling.The stability of LAB hydrogel was evaluated by reuse experiments.The images of confocal microscopy and scanning electron microscope showed that the bacteria with high cell viability were distributed in the hydrogel and intact structure of the living hydrogel was maintained after 100 times of reuse as yoghurt starter.In conclusion,the 3 D printed LAB bio-hydrogel developed in this study has the advantage of reuse and sustainability,which is expected to open up a new way for the preparation of food culture starter.

Application of Lactic Acid Bacteria(LAB) in Freshness Keeping of Tilapia Fillets as Sashimi

Aquatic products are extremely perishable food commodities. Developing methods to keep the freshness of fish represents a major task of the fishery processing industry. Application of Lactic Acid Bacteria(LAB) as food preservative is a novel approach. In the present study, the possibility of using lactic acid bacteria in freshness keeping of tilapia fillets as sashimi was examined. Fish fillets were dipped in Lactobacillus plantarum 1.19(obtained from China General Microbiological Culture Collection Center) suspension as LAB-treated group. Changes in K-value, APC, sensory properties and microbial flora were analyzed. Results showed that LAB treatment slowed the increase of K-value and APC in the earlier storage, and caused a smooth decrease in sensory score. Gram-negative bacteria dominated during refrigerated storage, with Pseudomonas and Aeromonas being relatively abundant. Lactobacillus plantarum 1.19 had no obvious inhibitory effect against these Gram-negatives. However, Lactobacillus plantarum 1.19 changed the composition of Gram-positive bacteria. No Micrococcus were detected and the proportion of Staphylococcus decreased in the spoiled LAB-treated samples. The period that tilapia fillets could be used as sashimi material extended from 24 h to 48 h after LAB treatment. The potential of using LAB in sashimi processing was confirmed.

Influence of lactic acid bacteria, cellulase, cellulase-producing Bacillus pumilus and their combinations on alfalfa silage quality

This study assessed the effects of lactic acid bacteria(LAB), cellulase, cellulase-producing Bacillus pumilus and their combinations on the fermentation characteristics, chemical composition, bacterial community and in vitro digestibility of alfalfa silage. A completely randomized design involving a 8(silage additives)×3 or 2(silage days) factorial arrangement of treatments was adopted in the present study. The 8 silage additive treatments were: untreated alfalfa(control); two commercial additives(GFJ and Chikuso-1); an originally selected LAB(Lactobacillus plantarum a214) isolated from alfalfa silage; a cellulase-producing Bacillus(CB) isolated from fresh alfalfa; cellulase(C); and the combined additives(a214+C and a214+CB). Silage fermentation characteristics, chemical composition and microorganism populations were analysed after 30, 60 and 65 days(60 days followed by exposure to air for five additional days). In vitro digestibility was analysed for 30 and 60 days. Compared with the other treatments, selected LAB a214, a214 combined with either C or CB, and Chikuso-1 had the decreased(P<0.001) pH and increased(P<0.001) lactic acid concentrations during the ensiling process, and there were no differences(P>0.05) among them. Fiber degradation was not significant(P≥0.054) in any C or CB treatments. The a214 treatment showed the highest(P=0.009) in vitro digestibility of dry matter(595.0 g kg–1DM) after ensiling and the highest abundance of Lactobacillus(69.42 and 79.81%, respectively) on days 60 and 65, compared to all of other treatments. Overall, the silage quality of alfalfa was improved with the addition of a214, which indicates its potential as an alfalfa silage inoculant.

Effect of lactic acid bacteria and propionic acid on conservation characteristics,aerobic stability and in vitro gas production kinetics and digestibility of whole-crop corn based total mixed ration silage

This study was conducted to evaluate the effect of lactic acid bacteria and propionic acid on the fermentation quality, aer- obic stability and in vitro gas production kinetics and digestibility of whole-crop corn based totalmixed ration （TMR） silage. Total mixed ration was ensiled with four treatments： （1） no additives （control）; （2） an inoculant （Lactobacillus plantarum） （L）; （3） propionicacid （P）; （4） propionic acid＋lactic acid bacteria （PL）. All treatments were ensiled in laboratory-scale silos for 45 days, and then subjected to an aerobic stability test for12 days. Further, four TMR silages were incubated in vitro with buffered rumen fluid to study in vitro gas production kinetics and digestibility. The results indicated that all TMR silages had good fermentation characteristics with low pH （〈3.80） and ammonia nitrogen （NH3-N） contents, and high lactic acid contents as well as Flieg points. Addition of L further improved TMR silage quality with more lactic acid production. Addition of P and PL decreased lactic acid and NH3-N contents of TMR silage compared to the control （P〈0.05）. After 12 days aerobic exposure, P and PL silages remained stable, but L and the control silages deteriorated as indicated by a reduction in lactic acid and an increase in pH, and numbers of yeast. Compared to the control, addition of L had no effects on TMR silage in terms of 72 h cumulative gas production, in vitro dry matter digestibility, metabolizable energy, net energy for lactation and short chain fatty acids, whereas addition of PL significantly （P〈0.05） increased them. L silage had higher （P〈0.05） in vitro neutral detergent fiber digestibility than the control silage. The results of our study suggested that TMR silage prepared with whole-crop corn can be well preserved with or without additives. Furthermore, the findings of this study suggested that propionic acid is compatible with lactic acid bacteria inoculants, and when used together, although they reduced lactic acid production of TMR silage, they improved aerobic stability and in vitro nutrients digestibility of TMR silage.

Effects of lactic acid bacteria isolated from fermented mustard on lowering cholesterol

Objective:To evaluate the ability of lactic acid bacteria(LAB)strains isolated from fermented mustard to lower the cholesterol in vitro.Methods:The ability of 50 LAB strains isolated from fermented mustard on lowering cholesterol in vitro was determined by modified o-phtshalaldehyde method.The LAB isolates were analyzed for their resistance to acid and bile salt.Strains with lowering cholesterol activity,were determined adherence to Caco-2 cells.Results:Strain B0007,B0006 and B0022 assimilated more cholesterol than BCRC10474 and BCRC17010.The isolated strains showed tolerance to pH 3.0 for 3h despite variations in the degree of viability and bile-tolerant strains,with more than 10~s CFU/mL after incubation for 24 h at 1%oxigall in MRS.In addition,strain B0007 and B0022 identified as Lactobacillus plantarum with 16S rDNA sequences were able to adhere to the Caco-2 cell lines.Conclusions:These strains B0007 and B0022 may be potential functional sources for cholesterollowering activities as well as adhering to Caco-2 cell lines.

The aflatoxin B1 isolating potential of two lactic acid bacteria

Objective:To determine lactic acid bacteria's capability to enhance the process of binding and isolating aflatoxin B1 and to utilize such lactic acid bacteria as a food supplement or probiotic products for preventing absorption of aflatoxin Bl in human and animal bodies.Methods:In the present research,the bacteria were isolated from five different sources.For surveying the capability of the bacteria in isolating aflatoxin Bl,ELISA method was implemented,and for identifying the resultant strains through 16S rRNA sequencing method,universal primers were applied.Results:Among the strains which were isolated,two strains of Lactobacillus pentosus and Lactobacillus beveris exhibited the capability of absorbing and isolating aflatoxin Bl by respectively absorbing and discharging 17.4%and 34.7%of the aforementioned toxin existing in the experiment solution.Conclusions:Strains of Lactobacillus pentosus and Lactobacillus beveris were isolated from human feces and local milk samples,respectively.And both strains has the ability to isolate or bind with aflatoxin Bl.

The effects of fermentation and adsorption using lactic acid bacteria culture broth on the feed quality of rice straw

To improve the nutritional value and the palatability of air-dried rice straw, culture broth of the lactic acid bacteria community SFC-2 was used to examine the effects of two different treatments, fermentation and adsorption. Air-dried and chopped rice straw was treated with either fermentation for 30 d after adding 1.5 L nutrient solution（50 m L inocula L–1, 1.2×1012 CFU m L–1 inocula） kg–1 straw dry matter, or spraying a large amount of culture broth（1.5 L kg–1 straw dry matter, 1.5×1011 CFU m L–1 culture broth） on the straw and allowing it to adsorb for 30 min. The feed quality and aerobic stability of the resulting forage were examined. Both treatments improved the feed quality of rice straw, and adsorption was better than fermentation for preserving nutrients and improving digestibility, as evidenced by higher dry matter（DM） and crude protein（CP） concentrations, lower neutral detergent fiber（NDF）, acid detergent fiber（ADF） and NH3-N concentrations, as well as higher lactic acid production and in vitro digestibility of DM（IVDMD）. The aerobic stability of the adsorbed straw and the fermented straw was 392 and 480 h, respectively. After being exposed to air, chemical components and microbial community of the fermented straw were more stable than the adsorbed straw.

The Role of Lactic Acid Bacteria in Milk Fermentation

Species of lactic acid bacteria (LAB) represent as potential microorganisms and have been widely applied in food fermentation worldwide. Milk fermentation process has been relied on the activity of LAB, where transformation of milk to good quality of fermented milk products made possible. The presence of LAB in milk fermentation can be either as spontaneous or inoculated starter cultures. Both of them are promising cultures to be explored in fermented milk manufacture. LAB have a role in milk fermentation to produce acid which is important as preservative agents and generating flavour of the products. They also produce exopolysaccharides which are essential as texture formation. Considering the existing reports on several health-promoting properties as well as their generally recognized as safe (GRAS) status of LAB, they can be widely used in the developing of new fermented milk products.

The effect of lactic acid bacteria inoculums on in vitro rumen fermentation, methane production, ruminal cellulolytic bacteria populations and cellulase activities of corn stover silage

The objective of this study was to investigate the effect of lactic acid bacteria(LAB) inoculums on fermentation quality and in vitro digestibility of corn stover silage.Corn stover was ensiled without(control) or with Lactobacillus plantarum(LP),Enterococcus faecalis(EF),and Enterococcus mundtii(EM) for 45 days.The fermentation characteristics were assessed,and subsequent in vitro dry matter digestibility(DM-D),neutral detergent fiber digestibility(NDF-D),volatile fatty acids(VFA),methane(CH4) production,cellulolytic bacteria proportions and their activities per corn stover silage were also determined.There was no significant difference(P>0.05) among the silage pH,lactic acid,crude protein(CP),water soluble carbohydrates(WSC) and lignocelluloses contents of different treatments.The relative proportions of Ruminococcus flavefaciens and Fibrobacter succinogenes,carboxymethyl-ocellulose and β-glycosidase activities,DM-D,NDF-D,and VFA production of in vitro incubation was higher(P<0.05) for silages inoculated with LP and EF than those of the control silage.Silage inoculated with LP showed the lowest(P<0.05) CH4 production per unit yield of VFA,which was positively corresponded to the lowest(P<0.05) ratio of acetate to propionate.In summary,the ensiling fermentation quality and subsequent utilization of corn stover silage were efficiently improved by inoculated with L.plantarum.

Comparative studies of versatile extracellular proteolytic activities of lactic acid bacteria and their potential for extracellular amino acid productions as feed supplements

Background:Increasing understanding on the functions of amino acids (AA) has led to new commercial applications and expansion of the worldwide markets.However,the current technologies rely heavily on non-food grade microorganism and chemical synthesis for the production of AA.Several studies reported that lactic acid bacteria (LAB) have the capability of producing AA owing to their well-established proteolytic system and amino acid biosynthesis genes.Hence,the objectives of this study were to explore the extracellular proteolytic activity of LAB isolated from various Malaysian fermented foods and their potential to produce AA extracellularly as feed supplements.Results:All the studied LAB isolates were versatile extracellular protease producers,whereby extracellular protease activities were detected from acidic to alkaline pH (pH 5,pH 6.5,pH 8) using qualitative and quantitative proteolytic assays.The highest proteolytic activity at pH 5 (15.76 U/mg) and pH 8 (19.42 U/mg) was achieved by Lactobacillus plantarum RG14,while Lactobacillus plantarum RS5 exhibited the highest proteolytic activity of 17.22 U/mg at pH 6.5.As for the results of AA production conducted in de Man,Rogosa and Sharpe medium and analysed by high pressure liquid chromatography system,all LAB isolates were capable of producing an array of AA.Generally,Pediococcus sp.showed greater ability for AA production as compared to Lactobacillus sp.Moreover,the studied LAB were able to produce a few major feed supplement AA such as methionine,lysine,threonine and tryptophan.P.pentosaceus TL-3 recorded the highest methionine and threonine productivity of 3.72 mg/L/h and 5.58 mg/L/h respectively.However,L.plantarum I-UL4 demonstrated a lysine productivity of 1.24 mg/L/h,while P.acidilactici TP-6 achieved up to 1.73 mg/L/h of tryptophan productivity.Conclusion:All the 17 studied LAB isolates possessed versatile extracellular proteolytic system and have vast capability of producing various amino acids including a few major feed supplement AA such as methionine,lysine,threonine and tryptophan.Despite AA production was strain dependent,the studied LAB isolates possessed vast potential and can be exploited further as a bio-agent or an alternative amino acids and bioactive peptide producers.

Effect of Cellulase and Lactic Acid Bacteria on Fermentation Quality and Chemical Composition of Wheat Straw Silage

The object of this study was to determine the effect of cellulase and lactic acid bacteria (LAB) on fermentation quality and chemical composition of wheat straw silage. Silages were prepared using a small-scale fermentation system and the moisture level was adjusted to 60% of fresh matter (FM) with deionized water. Treatments were designed as: control silage without additives, LAB inoculant Lactobacillus casei Z3-1 (1.0 × 106 cfu·g-1 of FM), commercial inoculant L. plantarum FG 1 (1.0 × 106 cfu·g-1 of FM), Z3-1 + cellulase and FG 1 + cellulase. The neutral detergent fiber (NDF), acid detergent fiber (ADF) and crude protein (CP) contents of the wheat straw prior to ensiling were 76.93%, 48.52% and 4.63% of dry matter (DM), respectively. After 30 days of fermentation, the silages treated with LAB and LAB + cellulase had a lower (P < 0.05) pH and higher (P < 0.05) lactic acid content than the control, and the coliform bacteria, yeast and mold were inhibited at the early stage of fermentation. Besides, silages treated with cellulase had lower (P < 0.05) values of ADF and NDF than the control. The results confirmed that the addition of cellulase and LAB contributed to improving the fermentation quality of wheat straw silage.

Current Limitations and Challenges with Lactic Acid Bacteria: A Review

Lactic acid bacteria (LAB) play a critical role in food, agricultural, and clinical applications. The fast growing characteristics of LAB and their metabolic activity have been the key in most applications including food production, agricultural industry, and probiotics. However, the biochemical and biophysical environments have significant effect on the growth and metabolic activity of LAB. While the biochemical conditions are most likely established, controlling and optimizing of biochemical conditions have many limitations and challenges. In addition to selecting the right strain, desirable metabolic processes required optimizing and controlling the available nutrients including sugars, peptides, free amino acids, minerals, and vitamins in addition to buffering agents. Thus, much of research was conducted to understand the impact of available nutrients on the growth and metabolic activities of LAB. However, only a few nutritional parameters could be controlled at a time while holding other parameters constant. The nutritional parameters may also interact with each other resulting in faulty results. Characteristics of LAB such as fastidiousness in their nutritional requirements, ability to produce acid and antimicrobial compounds, and variations in the nutritional requirements among strains have added additional limitations and challenges in this regard. Thus, chemically defined media (CDM) were suggested to deal with different limitations and challenges. However, due to differences in growth conditions, results obtained in CDM may face some obstacles when it comes to industrial applications. Thus, this paper aimed to review the recent data in regard to the role of the nutritional requirements of LAB in optimizing and controlling metabolic activities and to discuss the associated limitations and challenges.

Potential applications of lactic acid bacteria and bacteriocins in antimycobacterial therapy

Tuberculosis(TB) is a communicable disease caused by Mycobacterium tuberculosis(M. tuberculosis). WHO estimated that 10.4 million new(incident) TB cases worldwide in year 2016. The increased prevalence of drug resistant strains and side effects associated with the current anti-tubercular drugs make the treatment options more complicated. Hence, there are necessities to identify new drug candidates to fight against various sub-populations of M. tuberculosis with less or no toxicity/side effects and shorter treatment duration. Bacteriocins produced by lactic acid bacteria(LAB) attract attention of researchers because of its "Generally recognized as safe" status. LAB and its bacteriocins possess an effective antimicrobial activity against various bacteria and fungi. Interestingly bacteriocins such as nisin and lacticin 3147 have shown antimycobacterial activity in vitro. As probiotics, LAB plays a vital role in promoting various health benefits including ability to modulate immune response against various infectious diseases. LAB and its metabolic products activate immune system and thereby limiting the M. tuberculosis pathogenesis. The protein and peptide engineering techniques paved the ways to obtain hybrid bacteriocin derivatives from the known peptide sequence of existing bacteriocin. In this review, we focus on the antimycobacterial property and immunomodulatory role of LAB and its metabolic products. Techniques for large scale synthesis of potential bacteriocin with multifunctional activity and enhanced stability are also discussed.

Probiotic Lactic Acid Bacteria: A Review

Lactic acid bacteria (LAB) play a critical role in food production and health maintenance. There is an increasing interest in these species to reveal the many possible health benefits associated with them. The actions of LAB are species and strain specific, and depend on the amount of bacteria available in the gastrointestinal tract. Consumers are very concerned of chemical preservatives and processed foods. However, products with or processed with LAB are accepted as a natural way to preserve food and promote health. This paper aimed to review the recent data in regard to the role of probiotic LAB in the preservation of foods, in the immunomodulation in the gastrointestinal tract, and in its health

Effect of lactic acid bacteria isolated from fermented mustard on immunopotentiating activity

Objective: To investigate the effect of lactic acid bacteria isolated from fermented mustard on immunopotentiating activity Methods: One hundred and fifty nine strains of lactic acid bacteria isolated from traditional Taiwan fermented mustard were evaluated for their immunopotentiating activity on a murine macrophage cell line RAW 264.7.Results: Of the strains, pronounced increases in the levels of nitric oxide(NO), tumor necrosis factor-α and interleukin-6 were observed in strains B0040, B0110 and B0145. Among them,strain B0145 had the highest NO and tumor necrosis factor-α generation in RAW 264.7 cells;strains B0040 and B0110 were also superior to that of Lactobacillus casei. These results demonstrated that NO and cytokines were effectively induced when the bacterial stimulants were treated with macrophages. In addition, strains B0040 and B0110 were identified as Lactobacillus plantarum, and B0145 as Weissella cibaria using 16 S rDNA analysis.Conclusions: The results implicated selected strains may be regarded as a biological response modifier and had a broad application prospects in exploiting new functional food or as a feed additive.

Cloning of Bile Salt Hydrolase Gene and Its Expression in Lactic Acid Bacteria

According to the sequence of the bile salt hydrolase （BSH） gene of Bifidobacterium and the restriction enzyme cutting sites of expression vector pNZ8148, primers were designed and the bile salt hydrolase （BSH） gene was gotten from Bacillus bifidus ATCC 29521 by PCR. BSH gene was inserted into lactic acid bacteria expression vector pNZ8148 to construct the recombinant pNZ8148-BSH. The recombinant pNZ8148-BSH was transferred into lactic acid bacteria NZ9000 with electrotransformation method. And the recombinant which could express BSH protein was obtained. It was identified by SDS-PAGE electrophoresis and activity verification. The result could provide a rationale reference for expressing BSH in lactic acid bacteria.