A Single Plasmid of Nisin-Controlled Bovine and Human Lactoferrin Expressing Elevated Antibacterial Activity of Lactoferrin-Resistant Probiotic Strains

Abstract

Lactoferrin (LF) is a multifunctional protein found in mammals, and it shows broad-spectrum antimicrobial activity. To improve the functional properties of specific probiotics in order to provide both the beneficial characteristics of lactic acid bacteria and the biological activity of LF, cDNAs of bovine LF (BLF), human LF (HLF), or porcine LF (PLF) were cloned into a nisin-inducible plasmid. These were then transformed into the selected eight probiotics, which are LF-resistant hosts. Expression of recombinant LFs (rLFs) was analyzed via SDS-PAGE and Western blot analysis. Although the selected host strains may not contain the nisRK genes (NisK, the sensor kinase; NisR, the regulator protein), the components of autoregulation, a low level of LFs expression can be successfully induced by using nisin within bacterial cells in a time-dependent manner in three engineered clones, including Lactobacillus delbrueckii/HLF, L. delbrueckii/BLF, and L. gasseri/BLF. Lactobacillus delbrueckii and Lactobacillus gasseri originate from yogurt and human milk, respectively, and both strains are functional probiotic strains. Therefore, we further compared the antibacterial activities of disrupted recombinant probiotic clones, conventional strains (host control), and vector control ones by using agar diffusion and broth inhibition analysis, and the expression of rLFs in the above three clones considerately improved their antibacterial efficacies against four important food-borne pathogens, namely, Escherichia coli, Staphylococcus aureus, Enterococcus faecalis, and Salmonellaenterica. In conclusion, this study provides a simple strategy for the production of functional LFs (BLF and HLF) in both functional and LF-resistant hosts for applications in the field.

Therapeutic effects of orally administration of viable and inactivated probiotic strains against murine urinary tract infection

“…A collection of probiotic strains, including Lactobacillus casei (BCRC 10358), L. casei (BCRC 10697), Pediococcus pentosaceus Mees (BCRC 11064), Lactobacillus paracasei (BCRC 12193), L. fermentum (BCRC 12194), Bi fi dobacterium breve Reuter (BCRC 12584), Lactobacillus coryniformis (BCRC 12935), Lactobacillus elbrueckii (BCRC 14008), Lactobacillus acidophilus (BCRC 14065), C. tyrobutyricu m (BCRC 14535), Bi fi dobacterium angulatum (BCRC 14605), Bi fi dobacterium bi fi dum (BCRC 14615), L. reuteri (BCRC 14625), Lactobacillus rhamnosus (BCRC 16000), L. paracasei (BCRC 17483), and Lactobacillus paraplantarum (BCRC 17971) were obtained from BCRC Taiwan as well. In addition, this study utilized several recombinant LF-expression probiotic strains developed in our prior report [ 28 ], including Lactobacillus delbrueckii /HLF, L. delbrueckii /BLF, B. angulatum /BLF, B. angulatum /PLF, and Lactobacillus gasseri /BLF, as well as three strains of L. gasseri (HM-1, HM-3 and HM-4; laboratory stock) isolated from human milk. All probiotic bacterial strains were activated and cultured anaerobically in MRS broth (Lactobacilli MRS Broth, Difco, BD, USA) at 37 °C without agitation.…”

“…In our recent report, we developed and expressed recombinant human lactoferrin (rHLF), bovine lactoferrin (rBLF), and porcine lactoferrin (rPLF) in several lactobacillus or bifidobacterial strains resistant to BLF. Of particular significance, inactivated probiotic cell lysates containing functional rLFs can help considerably enhance antibacterial activity of host lactobacillus or bifidobacterial strains against food-borne pathogens in vitro [ 28 ]. Collectively, the present study was set up to determine whether oral intake of natural or recombinant LF-expressing probiotic strains could be a promising therapeutic intervention for UTIs.…”

Lactoferrin Can Attenuate SARS-CoV-2: An Analysis of Evidential Relations

“…However, certain bacteria can adjust to the changing circumstances and produce siderophores (bacterialderived iron-chelating chemicals) that strive with Lf for Fe 3+ ions 52 . Several bacteria, such as those in the Neisseriaceae family, are adaptive to changing circumstances by producing particular receptors that attach Lf and induce variations in the Lf molecule tertiary shape, resulting in iron dispersion 53 . Lf has also been shown to have bactericidal action (Figure 1).…”

“…Despite contradictory findings, Lf appears to have dual immunomodulatory and immunostimulatory properties (Figure 1). The capacity of Lf to attach endotoxin is thought to play a significant role in immunomodulation 53 . Gram-negative bacteria are subjected to different innate immune system proteins when they infect a human host.…”

Comparison of extraction methods and nutritional benefits of proteins of milk and dairy products: A review

“…Antibacterial effect. LF has shown antimicrobial properties by binding with the lipid of the lipopolysaccharides of the bacterial membrane and, as a result, increasing the permeability of the membrane (Liu et al 2021b).…”