The complex world of kefir: Structural insights and symbiotic relationships.

Kefir milk, known for its high nutritional value and health benefits, is traditionally produced by fermenting milk with kefir grains. These grains are a complex symbiotic community of lactic acid bacteria, acetic acid bacteria, yeasts, and other microorganisms. However, the intricate coexistence mechanisms within these microbial colonies remain a mystery, posing challenges in predicting their biological and functional traits. This uncertainty often leads to variability in kefir milk's quality and safety. This review delves into the unique structural characteristics of kefir grains, particularly their distinctive hollow structure. We propose hypotheses on their formation, which appears to be influenced by the aggregation behaviors of the community members and their alliances. In kefir milk, a systematic colonization process is driven by metabolite release, orchestrating the spatiotemporal rearrangement of ecological niches. We place special emphasis on the dynamic spatiotemporal changes within the kefir microbial community. Spatially, we observe variations in species morphology and distribution across different locations within the grain structure. Temporally, the review highlights the succession patterns of the microbial community, shedding light on their evolving interactions.Furthermore, we explore the ecological mechanisms underpinning the formation of a stable community composition. The interplay of cooperative and competitive species within these microorganisms ensures a dynamic balance, contributing to the community's richness and stability. In kefir community, competitive species foster diversity and stability, whereas cooperative species bolster mutualistic symbiosis. By deepening our understanding of the behaviors of these complex microbial communities, we can pave the way for future advancements in the development and diversification of starter cultures for food fermentation processes.

Enhanced photodynamic inactivation against Escherichia coli O157:H7 provided by chitosan/curcumin coating and its application in food contact surfaces.

Sterilisation technologies are essential to eliminate foodborne pathogens from food contact surfaces. However, most of the current sterilisation methods involve high energy and chemical consumption. In this study, a photodynamic inactivation coating featuring excellent antibacterial activity was prepared by dispersing curcumin as a plant-based photosensitiser in a chitosan solution. The coating generated abundant reactive oxygen species (ROS) after light irradiation at 420 nm, which eradicated ≥99.999 % of Escherichia coli O157:H7. It was also found that ROS damaged the cell membrane, leading to the leakage of cell contents and cell shrinkage on the basis of chitosan. In addition, the production of ROS first excited the bacterial antioxidant defence system resulting in the increase of peroxidase (POD) and superoxide dismutase (SOD). ROS levels exceed its capacity, causing damage to the defence system and further oxidative decomposition of large molecules, such as DNA and proteins, eventually leading to the death of E. coli O157:H7. We also found the curcumin/chitosan coating could effectively remove E. coli O157:H7 biofilms by oxidative of extracellular polysaccharides and proteins. All the contributors made the chitosan/curcumin coating an efficient detergent comparable with HClO.

The Role of GmSnRK1-GmNodH Module in Regulating Soybean Nodulation Capacity.

SnRK1 protein kinase plays hub roles in plant carbon and nitrogen metabolism. However, the function of SnRK1 in legume nodulation and symbiotic nitrogen fixation is still elusive. In this study, we identified GmNodH, a putative sulfotransferase, as an interacting protein of GmSnRK1 by yeast two-hybrid screen. The qRT-PCR assays indicate that GmNodH gene is highly expressed in soybean roots and could be induced by rhizobial infection and nitrate stress. Fluorescence microscopic analyses showed that GmNodH was colocalized with GsSnRK1 on plasma membrane. The physical interaction between GmNodH and GmSnRK1 was further verified by using split-luciferase complementary assay and pull-down approaches. In vitro phosphorylation assay showed that GmSnRK1 could phosphorylate GmNodH at Ser193. To dissect the function and genetic relationship of GmSnRK1 and GmNodH in soybean, we co-expressed the wild-type and mutated GmSnRK1 and GmNodH genes in soybean hairy roots and found that co-expression of GmSnRK1/GmNodH genes significantly promoted soybean nodulation rates and the expression levels of nodulation-related GmNF5α and GmNSP1 genes. Taken together, this study provides the first biological evidence that GmSnRK1 may interact with and phosphorylate GmNodH to synergistically regulate soybean nodulation.

Functional characterization of wild soybean (Glycine soja) GsSnRK1.1 protein kinase in plant resistance to abiotic stresses.

Protein kinases play crucial roles in the regulation of plant resistance to various stresses. In this work, we determined that GsSnRK1.1 was actively responsive to saline-alkali, drought, and abscisic acid (ABA) stresses by histochemical staining and qRT-PCR analyses. The wild-type GsSnRK1.1 but not the kinase-dead mutant, GsSnRK1.1(K49M), demonstrated in vitro kinase activity by phosphorylating GsABF2. Intriguingly, we found that GsSnRK1.1 could complement the loss of SNF1 kinase in yeast Msy1193 (-snf1) mutant, rescue growth defects of yeast cells on medium with glycerol as a carbon resource, and promote yeast resistance to NaCl or NaHCO3. To further elucidate GsSnRK1.1 function in planta, we knocked out SnRK1.1 gene from the Arabidopsis genome by the CRISPR/Cas9 approach, and then expressed GsSnRK1.1 and a series of mutants into snrk1.1-null lines. The transgenic Arabidopsis lines were subjected to various abiotic stress treatments. The results showed that GsSnRK1.1(T176E) mutant with enhanced protein kinase activity significantly promoted, but GsSnRK1.1(K49M) and GsSnRK1.1(T176A) mutants with disrupted protein kinase activity abrogated, plant stomatal closure and tolerance to abiotic stresses. In conclusion, this study provides the molecular clues to fully understand the physiological functions of plant SnRK1 protein kinases.

Quantitative phosphoproteomics reveals the role of wild soybean GsSnRK1 as a metabolic regulator under drought and alkali stresses.

Drought and alkali stresses cause detrimental effects on plant growth and development. SnRK1 protein kinases act as key energy and stress sensors by phosphorylation-mediated signaling in the regulation of plant defense reactions against adverse environments. To understand SnRK1-dependent phosphorylation events in signaling pathways triggered by abiotic factors, we employed quantitative phosphoproteomics to compare the global changes in phosphopeptides and phosphoproteins in 2kinm mutant Arabidopsis (SnRK1.1 T-DNA knockout and SnRK1.2 knockdown by ß-estradiol-induced RNAi) complemented with wild soybean GsSnRK1(wt) or dominant negative mutant GsSnRK1(K49M) in response to drought and alkali stresses. Among 4014 phosphopeptides (representing 2380 phosphoproteins) identified in this study, we finalized 74 phosphopeptides (representing 61 phosphoproteins), and 75 phosphopeptides (representing 57 phosphoproteins) showing significant changes in phosphorylation levels under drought and alkali treatments respectively. Function enrichment and protein-protein interaction analyses indicated that the differentially-expressed phosphoproteins (DPs) under drought and alkali stresses were mainly involved in signaling transduction, stress response, carbohydrate and energy metabolism, transport and membrane trafficking, RNA splicing and processing, DNA binding and gene expression, and protein synthesis/folding/degradation. These results provide assistance to identify bona fide and novel SnRK1 phosphorylation substrates and shed new light on the biological functions of SnRK1 kinase in responses to abiotic stresses. SIGNIFICANCE: These results provide assistance to identify novel SnRK1 phosphorylation substrates and regulatory proteins, and shed new light on investigating the potential roles of reversible phosphorylation in plant responses to abiotic stresses.

Genome-wide identification and expression analyses of nitrate transporter family genes in wild soybean (Glycine soja).

Nitrate transporters (NRTs) are important channel proteins facilitating cross-membrane movement of small molecules like NO3- which is a critical nutrient for all life. However, the classification and evolution of nitrate transporters in the legume plants are still elusive. In this study, we surveyed the wild soybean (G. soja) genomic databases and identified 120 GsNRT1 and 5 GsNRT2 encoding genes. Phylogenetic analyses show that GsNRT1 subfamily is consisted of eight clades (NPF1 to NPF8), while GsNRT2 subfamily has only one clade. Gene chromosomal location and evolutionary historic analyses indicate that GsNRT genes are unevenly distributed on 19 out of 20 G. soja chromosomes and segmental duplications may take a major part in the expansion of GsNRT family. Investigations of gene structure and protein motif compositions suggest that GsNRT family members are highly conserved in structures of both gene and protein levels. In addition, we analyzed the spatial expression patterns of representative GsNRT genes and their responses to exogenous nitrogen and carbon supplies and different abiotic stresses. The qRT-PCR data indicated that 16 selected GsNRT genes showed various expression levels in the roots, stems, leaves, and pods of young G. soja plants, and these genes were regulated by not only nitrogen and carbohydrate nutrients but also NaCl, NaHCO3, abscisic acid (ABA), and salicylic acid (SA). These results suggest that GsNRT genes may be involved in the regulation of plant growth, development, and adaptation to environmental stresses, and the study will shed light on functional dissection of plant nitrate transporter proteins in the future.