Aims: Probiotics are claimed to confer many health effects upon consumption. However, the survivability of probiotic under the harsh conditions in the gastrointestinal tract has been a challenge. This study aimed to improve the survivability of Lactobacillus rhamnosus GG under gastrointestinal condition through co-extrusion microencapsulation and the addition of black bean extract. Methodology and results: Optimization was carried out on wall material formulation, types of pectin (low and high methoxyl pectin) and alginate: pectin ratio (2:1 and 3:1), and black bean extract concentration (0 to 1% w/v) to produce capsules with desired properties. The effect of L. rhamnosus GG microencapsulation with and without black bean extract on its survivability under simulated gastrointestinal conditions was also investigated. The optimal formulation that gives the highest microencapsulation efficiency (86.17%) was low methoxyl pectin, alginate: pectin ratio at 3:1, and 0.5% (w/v) of black bean extract. The inclusion of black bean extract into L. rhamnosus GG microencapsulation showed no significant effect (p >0.05) on the capsule diameter, with a mean diameter of 715.44 µm and a high microencapsulation efficiency of 97.4%. The viability of encapsulated L. rhamnosus GG increased with black bean extract after 6 h of sequential digestion with the final viable cell count of 12.47 log10 CFU/mL, which meet the minimum requirement of 10^6-10^7 log10 CFU/mL viable cells. Conclusion, significance and impact of study The high microencapsulation efficiency and survivability through sequential digestion showed that the optimized encapsulated L. rhamnosus GG with black bean extract has the potential to be a value-added ingredient in food application.
Lacticaseibacillus rhamnosus ; Alginates ; Phaseolus

ABSTRACT The antagonistic effect of probiotics against oral pathogens merits exploration because these bacteria are beneficial to the host’s health. The antimicrobial activity of two probiotic strains, Lactobacillus casei and Lactobacillus salivarius, as well as L. casei and L. salivarius combination (1:1), was investigated against Streptococcus mutans, Streptococcus sobrinus, Candida albicans, Candida glabrata and Candida tropicalis using agar-well diffusion, auto-aggregation and coaggregation assays. L. salivarius cell-free supernatant (CFS) alone exhibited greater inhibitory effect against Streptococci spp. compared to L. casei CFS alone and the combination. However, no inhibition was observed for Candida spp. L. salivarius alone exhibited significantly stronger auto-aggregation than L. casei alone (p ≤ 0.05) and L. casei and L. salivarius combination. L. salivarius exhibited strong coaggregation ability with Candida spp., followed by Streptococci spp. while L. casei exhibited coaggregation only with Streptococci spp. However, L. casei and L. salivarius combination did not display any coaggregation with all strains. L. salivarius alone exhibited a stronger antagonistic effect on the tested organisms than L. casei alone or in combination. Based on the results, both probiotic strains showed good antimicrobial activities against oral pathogens and should be further studied for their human health benefits.
Lacticaseibacillus casei--pathogenicity ; Ligilactobacillus salivarius--pathogenicity

The construction of efficient and stable Lactobacillus expression vector is critical for strain improvement and development of customized strains. In this study, four endogenous plasmids were isolated from Lacticaseibacillus paracasei ZY-1 and subjected to functional analysis. The Escherichia coli-Lactobacillus shuttle vectors pLPZ3N and pLPZ4N were constructed by combining the replicon rep from pLPZ3 or pLPZ4, the chloramphenicol acetyltransferase gene cat from pNZ5319 and the replicon ori from pUC19. Moreover, the expression vectors pLPZ3E and pLPZ4E with the promoter P_ldh3 of lactic acid dehydrogenase and the mCherry red fluorescent protein as a reporter gene were obtained. The size of pLPZ3 and pLPZ4 were 6 289 bp and 5 087 bp, respectively, and its GC content, 40.94% and 39.51%, were similar. Both shuttle vectors were successfully transformed into Lacticaseibacillus, and the transformation efficiency of pLPZ4N (5.23×10²-8.93×10² CFU/μg) was slightly higher than that of pLPZ3N. Furthermore, the mCherry fluorescent protein was successfully expressed after transforming the expression plasmids pLPZ3E and pLPZ4E into L. paracasei S-NB. The β-galactosidase activity of the recombinant strain obtained from the plasmid pLPZ4E-lacG constructed with P_ldh3 as promoter was higher than that of the wild-type strain. The construction of shuttle vectors and expression vectors provide novel molecular tools for the genetic engineering of Lacticaseibacillus strains.
Lacticaseibacillus ; Lacticaseibacillus paracasei ; Plasmids/genetics* ; Genetic Vectors/genetics* ; Lactobacillus/genetics* ; Escherichia coli/genetics*

To understand the effects of Lactobacillus rhamnosus GG (ATCC 53103) on intestinal barrier function in pre-weaning piglets under normal conditions, twenty-four newborn littermate piglets were randomly divided into two groups. Piglets in the control group were orally administered with 2 mL 0.1 g/mL sterilized skim milk while the treatment group was administered the same volume of sterilized skim milk with the addition of viable L. rhamnosus at the 1st, 3rd, and 5th days after birth. The feeding trial was conducted for 25 d. Results showed that piglets in the L. rhamnosus group exhibited increased weaning weight and average daily weight gain, whereas diarrhea incidence was decreased. The bacterial abundance and composition of cecal contents, especially Firmicutes, Bacteroidetes, and Fusobacteria, were altered by probiotic treatment. In addition, L. rhamnosus increased the jejunal permeability and promoted the immunologic barrier through regulating antimicrobial peptides, cytokines, and chemokines via Toll-like receptors. Our findings indicate that oral administration of L. rhamnosus GG to newborn piglets is beneficial for intestinal health of pre-weaning piglets by improving the biological, physical, and immunologic barriers of intestinal mucosa.
Administration, Oral ; Animals ; Animals, Newborn ; Cytokines/genetics* ; Female ; Gastrointestinal Microbiome ; Immunity, Innate ; Intestinal Mucosa/immunology* ; Lacticaseibacillus rhamnosus ; Male ; Probiotics/administration & dosage* ; Signal Transduction ; Swine ; Weaning