In Vitro Probiotic Characteristics and Whole Genome Sequence Analysis of Lactobacillus Strains Isolated from Cattle-Yak Milk

Abstract

Cattle-yak milk is an important raw material and an indispensable source of high-quality food for local farmers and herdsmen to produce ghee, milk residue, yogurt, and other dairy products. In this study, Lactobacillus strains were isolated from cattle-yak milk for potential probiotic candidates using a series of in vitro tests, including probiotic characterization and safety evaluation (antibiotic susceptibility and hemolytic ability). The results found that the Lactobacillus rhamnosus CY12 strain showed a high survival rate in bile salts, under acid conditions, and in the gastrointestinal juice environment, as well as showing high antimicrobial activity and adhesive potential. The safety evaluation showed that all strains were considered non-hemolytic. In addition, the whole-genome sequencing indicated that the strain CY12 spanned 2,506,167 bp, with an average length of 881 bp; the GC content in the gene region (%) was 47.35, contained 1347 protein-coding sequences, and accounted for 85.72% of the genome. The genome annotation showed that genes mainly focused on the immune system process, metabolic process, carbohydrate utilization, carbon metabolism, galactose metabolism, and biological adhesion, etc. This study revealed that the Lactobacillus rhamnosus CY12 strain might be an excellent potential probiotic in the development of feed additives for animals and has the ability to promote health.

Assessment of the safety and probiotic characteristics of Lactobacillus salivarius CGMCC20700 based on whole-genome sequencing and phenotypic analysis

“…Probiotics are considered biotherapeutic agents owing to their potential to bestow various health benefits ( Al-Khalaifa et al, 2019 ). Numerous studies have shown that probiotics may adhere and survive in the gastrointestinal tract of humans and animals and contribute to maintaining a microecological balance of the gut microbiome, promoting digestive and metabolic processes, and modulating the immune response, thereby enhancing host immunity and improving human and animal health ( Kai et al, 2020 ; Zheng et al, 2021 ). However, since the effectiveness of probiotics is species or strain dependent, they should meet a series of specific characteristics, such as safety, functional and beneficial characteristics ( Li et al, 2020 ; Wu et al, 2022 ).…”

“…Antibiotic resistance was evaluated by adopting the method from a previous study ( Zheng et al, 2021 ). Briefly, susceptibility to the following 13 antibiotics was assessed using filter paper disks infused with the following: 30 μg each of ceftazidime, cefuroxime, cefazolin, vancomycin, and tetracycline; 10 μg each of penicillin, streptomycin, gentamicin, and amoxicillin; 100 μg of ampicillin; and 15 μg of erythromycin (Shanghai Yibaiju Economic and Trade Co., Ltd., Shanghai, China).…”

“…The hemolytic activity assays were performed by adopting the method from Zheng et al (2021) . Briefly, L. salivarius CGMCC20700 and Escherichia coli CMCC(B)44102 cultures were crossed, streaked on Columbia blood agar containing fresh sheep blood (Shanghai Yibaiju Economic and Trade Co., Ltd., Shanghai, China) and incubated at 37°C for 24 h. Hemolytic activity was determined according to the following rules: if the colony (strain) showed a grass-green ring on the plate, the strain was identified as α-hemolytic; if the colony showed a completely clear hemolytic ring on the plate, the strain was identified as β-hemolytic; and if no changes were observed, the colony was identified as nonhemolytic.…”

“…Probiotic genes were identified to evaluate the potential probiotic functions of L. salivarius CGMCC20700, as previously reported ( Goel et al, 2020 ; Zheng et al, 2021 ). Briefly, different probiotic genes were obtained using BLASTP in the NCBI database and compared with known probiotic genes.…”

Lactobacillus rhamnosus CY12 Enhances Intestinal Barrier Function by Regulating Tight Junction Protein Expression, Oxidative Stress, and Inflammation Response in Lipopolysaccharide-Induced Caco-2 Cells

“…However, in recent years, a growing body of research has revealed that the functions of probiotics are highly strain-specific, and their biological effects should be individually evaluated [ 22 ]. Our previous work reported the isolation of the Lactobacillus rhamnosus CY12 ( L. rhamnosus CY12) strain from cattle-yak milk that displayed potential probiotic characteristics such as a high survival rate in acidic conditions and bile salts, high antimicrobial activity, and high adhesive potential [ 31 ]. Considering the specificity of the strains, the protective effects of the novel L. rhamnosus CY12 strain isolated from cattle-yak milk on intestinal inflammation are worthy for further study.…”