Efficacy of the incorporation between self-encapsulation and cryoprotectants on improving the freeze-dried survival of probiotic bacteria.

AIMS: This study aimed to improve the viability of probiotic bacteria during freeze-drying by the combination of self-encapsulation and cryoprotectants. METHODS AND RESULTS: Lactiplantibacillus plantarum VAL6 and Lactobacillus acidophilus VAR1 were exposed to environmental stresses including temperature, pH and increased CO2 concentration before performing freeze-drying with the addition of cryoprotectants. The results proved that tested stresses can stimulate the bacteria to synthesize more extracellular polymeric substances to form self-encapsulation that increases their freeze-dried viability. In combination with cryoprotectants to form double-layered microencapsulation, L. plantarum VAL6 stressed at pH 3.5 in combination with whey protein isolate could achieve the highest Improving Cell Viability of 4361-fold, while L. acidophilus VAR1 stressed at 25o C in combination with alginate gave a maximum Improving Cell Viability of 73.33-fold. CONCLUSIONS: The combination of self-encapsulation and cryoprotectants significantly improves the freeze-dried viability of probiotics. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report that uses environmental stress to stimulate extracellular polymeric substance synthesis for self-encapsulation formation combined with the addition of cryoprotectants to enhance the freeze-dried survival of probiotics. This could be a novel approach in improving the viability of probiotic strains for various applications.

Correlation Between the Amount of Extracellular Polymeric Substances and the Survival Rate to Freeze-Drying of Probiotics.

To demonstrate that the amount of extracellular polymeric substances (EPS) and the freeze-dried viability of probiotics are correlated. Three strains of probiotics including Lactiplantibacillus plantarum, Lactobacillus acidophilus, and Bifidobacterium bifidum were subjected to environmental challenges, such as temperature, pH, and carbon dioxide. The results indicated that the challenges could stimulate the EPS synthesis of the probiotics. The experimental correlation between the amount of synthesized EPS and the freeze-dried survival rate was also analyzed, and the viability of each of the three strains was represented by the following functions in which the equation of L. plantarum is y = - 0.0336x2 + 2.7059x - 14.849 with R2 = 0.9699, the B. bifidum's equation is y = - 0.0554x2 + 2.6243x - 13.654 with R2 = 0.9554, and the L. acidophilus's one was y = 0.0346x2 + 0.5862x - 9.1339 with R2 = 0.9733. This could be a new approach to determining the freeze-dried viability of probiotic strains based on the measured EPS content.

Identification of a hormogonium polysaccharide-specific gene set conserved in filamentous cyanobacteria.

Cyanobacteria comprise a phylum defined by the capacity for oxygenic photosynthesis. Members of this phylum are frequently motile as well. Strains that display gliding or twitching motility across semisolid surfaces are powered by a conserved type IV pilus system (T4P). Among the filamentous, heterocyst-forming cyanobacteria, motility is usually confined to specialized filaments known as hormogonia, and requires the deposition of an associated hormogonium polysaccharide (HPS). The genes involved in assembly and export of HPS are largely undefined, and it has been hypothesized that HPS exits the outer membrane via an atypical T4P-driven mechanism. Here, several novel hps loci, primarily encoding glycosyl transferases, are identified. Mutational analysis demonstrates that the majority of these genes are essential for both motility and production of HPS. Notably, most mutant strains accumulate wild-type cellular levels of the major pilin PilA, but not extracellular PilA, indicating dysregulation of the T4P motors, and, therefore, a regulatory interaction between HPS assembly and T4P activity. A co-occurrence analysis of Hps orthologs among cyanobacteria identified an extended set of putative Hps proteins comprising most components of a Wzx/Wzy-type polysaccharide synthesis and export system. This implies that HPS may be secreted through a more canonical pathway, rather than a T4P-mediated mechanism.

Protective anti-chlamydial vaccine regimen-induced CD4+ T cell response mediates early inhibition of pathogenic CD8+ T cell response following genital challenge.

We have demonstrated previously that TNF-α-producing CD8+ T cells mediate chlamydial pathogenesis, likely in an antigen (Ag)-specific fashion. Here we hypothesize that inhibition of Ag-specific CD8+ T cell response after immunization and/or challenge would correlate with protection against oviduct pathology induced by a protective vaccine regimen. Intranasal (i.n.) live chlamydial elementary body (EB), intramuscular (i.m.) live EB, or i.n. irrelevant antigen, bovine serum albumin (BSA), immunized animals induced near-total protection, 50% protection, or no protection, respectively against oviduct pathology following i.vag. C. muridarum challenge. In these models, we evaluated Ag-specific CD8+ T cell cytokine response at various time-periods after immunization or challenge. The results show protective efficacy of vaccine regimens correlated with reduction of Ag-specific CD8+ T cell TNF-α responses following i.vag. chlamydial challenge, not after immunization. Depletion of CD4+ T cells abrogated, whereas adoptive transfer of Ag-specific CD4+ T cells induced the significant reduction of Ag-specific CD8+ T cell TNF-α response after chlamydial challenge. In conclusion, protective anti-chlamydial vaccine regimens induce Ag-specific CD4+ T cell response that mediate early inhibition of pathogenic CD8+ T cell response following challenge and may serve as a predictive biomarker of protection against Chlamydia -induced chronic pathologies.

Electrochemical Scanning Tunneling Microscopic Study of the Potential Dependence of Germanene Growth on Au(111) at pH 9.0.

Germanene is a 2D material whose structure and properties are of great interest for integration with Si technology. Preparation of germanene experimentally remains a challenge because, unlike graphene, bulk germanene does not exist. Thus, germanene cannot be directly exfoliated and is mostly grown in ultrahigh vacuum. The present report uses electrodeposition in an aqueous HGeO3- solution at pH 9. Germanene deposition has been limited to 2-3 monolayers, thus greatly restricting many applicable characterization methods. The in situ technique of electrochemical scanning tunneling microscopy was used to follow Ge deposition on Au(111) as a function of potential. Previous work by this group at pH 4.5 suggested germanene growth, but no buffer was used, resulting in change in surface pH. The addition of borate buffer to create pH 9.0 solution has reduced hydrogen formation and stabilized the surface pH, allowing systematic characterization of germanene growth versus potential. Initial germanene nucleated at defects in the Au(111) herringbone (HB) reconstruction. Subsequent growth proceeded down the face-centered cubic troughs, slowly relaxing the HB. The resulting honeycomb (HC) structure displayed an average lattice constant of 0.41 ± 0.06 nm. Continued growth resulted in the addition of a second layer on top, formed initially by nucleating around small islands and subsequent lateral 2D growth. Near atomic resolution of the germanene layers displayed small coherent domains, 2-3 nm, of the HC structure composed of six-membered rings. Domain walls were based on defective, five- and seven-membered rings, which resulted in small rotations between adjacent HC domains.