Starch Degradation Research Articles

Transcription factor MdGTL1a accelerates starch degradation by promoting the MdBam8 expression in postharvest apple fruit.

Starch degradation plays a central role in fruit ripening. The β-amylase (Bam) catalysts starch degradation of apple fruit. However, the molecular mechanism regulating Bam gene expression in apples remains unclear. Using yeast one-hybrid library screening, we identified a transcription factor MdGTL1a that directly binds to the MdBam8 promoter. This bind was verified by using an electrophoretic mobility shift assay and confirmed to enhance the promotor activity of MdBam8 by promoter-β-glucuronidase transactivation assay. Transient over-expression of MdGTL1a activated MdBam8 expression and further enhanced starch degradation in apple flesh. Subcellular localization analyses in tobacco protoplasts demonstrated that the nucleus was the exclusive location of the MdGTL1a-GFP fusion protein. Exogenous salicylic acid treatment decreased MdGTL1a expression and resulted in higher starch content in apples, which was consistent with the fruit ripening. It is concluded that salicylic acid treatment could enhance apple storage quality by inhibiting starch degradation through a crucial transcription factor, MdGTL1a.

Possible role of autophagy in microbial volatile pollutant-induced starch degradation and expression of hypoxia responsive genes.

Autophagy is thought to be critically involved in the regulation of nutrient metabolism and gene expression. Nevertheless, little is known about its role in regulating starch metabolism and hypoxia responsive genes in plants exposed to microbial volatile pollutants. In the present study, we found that exposure of Arabidopsis to Enterobacter aerogene (E. aerogene) volatile pollutants induced autophagy, as indicated by autophagosome formation. The exposure also caused upregulation of autophagy-associated genes, such as ATGs, NBR1, ATI1, and ATG8e-regulating transcription factors. Additionally, exposure to E. aerogenes volatile pollutants induced starch degradation in the roots of Arabidopsis seedlings. Finally, we found that ATG7-deficiency negatively affected the expression of hypoxia-responsive genes (i.e HRE1, HRA1, and ADH1) and starch degradation induced by E. aerogenes volatile pollutants. Overall, our study reveals that microbial volatile pollutants can induce starch degradation and autophagy, which participates in the regulation of some hypoxia-responsive genes and starch metabolism. These findings help to define the role of autophagy in plant nutrient metabolism and regulation of gene expression under microbial volatile pollutant exposure. The insights gained may contribute to agricultural management when living organisms face challenges from microbial volatile pollutants.

Comparative miRNAome combined with transcriptome and degradome analysis reveals a novel miRNA-mRNA regulatory network associated with starch metabolism affecting pre-harvest sprouting resistance in wheat

BackgroundPre-harvest sprouting (PHS) is one of the most important problems associated with the severe decrease of yield and quality under disaster weather of continuous rain in wheat harvesting stage. At present, the functions and mechanisms related to the involvement of post-transcriptional regulation has not been studied very clearly in PHS resistance.ResultsThis study compared the differences of germinated seeds in miRNAome between the PHS-tolerant and PHS-susceptible white wheat varieties. A total of 1879 miRNAs were identified from three different stages during seed germination. In order to further obtain candidate miRNAs, the different datasets of differentially expressed miRNAs were excavated by using differential-expression and time-series analysis. Combined with degradome data, the miRNA-mRNA networks analysis was performed after genome-wide screening of target genes, and then KEGG enrichment highlighted that the starch and sucrose metabolism pathway related to PHS was specifically enriched in an especial target-gene dataset derived from R12R18-HE miRNAs. Based on transcriptome data, a network associated with starch metabolism was systematically and completely reconstructed in wheat. Then, the starch degradation pathway controlled by seven miRNA-RNA pairs were supposed to be the essential regulation center for seed germination in wheat, which also could play a critical role on the PHS resistance.ConclusionOur findings revealed the complex impact of the miRNA-mediated mechanism for forming intrinsic and inherent differences, which resulting in significant difference on PHS performance between white wheat varieties.

The Pseudoenzyme β-Amylase9 From Arabidopsis Activates α-Amylase3: A Possible Mechanism to Promote Stress-Induced Starch Degradation.

Starch accumulation in plants provides carbon for nighttime use, for regrowth after periods of dormancy, and for times of stress. Both ɑ- and β-amylases (AMYs and BAMs, respectively) catalyze starch hydrolysis, but their functional roles are unclear. Moreover, the presence of catalytically inactive amylases that show starch excess phenotypes when deleted presents questions on how starch degradation is regulated. Plants lacking one of these catalytically inactive β-amylases, BAM9, have enhanced starch accumulation when combined with mutations in BAM1 and BAM3, the primary starch degrading BAMs in response to stress and at night, respectively. BAM9 has been reported to be transcriptionally induced by stress although the mechanism for BAM9 function is unclear. From yeast two-hybrid experiments, we identified the plastid-localized AMY3 as a potential interaction partner for BAM9. We found that BAM9 interacted with AMY3 invitro and that BAM9 enhances AMY3 activity about three-fold. Modeling of the AMY3-BAM9 complex predicted a previously undescribed alpha-alpha hairpin in AMY3 that could serve as a potential interaction site. Additionally, AMY3 lacking the alpha-alpha hairpin is unaffected by BAM9. Structural analysis of AMY3 showed that it can form a homodimer in solution and that BAM9 appears to replace one of the AMY3 monomers to form a heterodimer. The presence of both BAM9 and AMY3 in many vascular plant lineages, along with model-based evidence that they heterodimerize, suggests that the interaction is conserved. Collectively these data suggest that BAM9 is a pseudoamylase that activates AMY3 in response to cellular stress, possibly facilitating stress recovery.

P1322 Gut microbiome and Virome dynamics between rural and urban population suggests the severity in ulcerative colitis patients

Abstract Background Ulcerative colitis (UC) is chronic, inflammatory disease of the colon mucosa. Gut microbiome dysbiosis is considered to play a significant role in UC pathogenesis. Apart from bacteria, viruses are the intricate part of the gut microbiome which are often neglected in 16s rRNA based metagenomic studies. Therefore, whole genome sequencing (WGS) based studies of gut meta-DNA should be implemented to identify the microorganisms of different domains (bacteria, and viruses) residing in the human gut. Methods Human gut microbiota composition was assessed at microbiome and virome profiling using WGS of fecal samples. The gut metagenomes were further evaluated for the human gut resistome study. Results The UC gut was enriched with the pathobionts such as Escherichia coli and Bacteroides sp. The beneficial microbe resides in perfect mutualism in the gut such as Prevotella sp CAG_279, Firmicutes bacterium CAG_129, Ruminococcus torques (degradation of resistant starch) were lower in abundance in UC. The gut microbiota composition of urbanized population was more similar to UC as they have lower abundance of Firmicutes, Faecalibacterium prausnitzii and Oscillibacter sp 57_20. The lower prevalence of Ruminococcus torques, a keystone taxa in UC urban participants, indicates the loss of physiological functions. The UC patients exhibited a dysbiotic virobiota, characterized by increase in viral abundance, particularly from the genera enterovirus (observed in enteric gut infections), lentivirus (observed in chronic gut diseases), and Betacoronavirus. In UC patients, meta-gut resistome was found to be dominated by antimicrobial resistant genes (ARGs) compared to healthy. The metagenome study revealed a higher prevalence of ARG genes in the rural population (378 in Healthy; 607 in UC) compared to the urban (340 in Healthy; 578 in UC). The maximum number of observed ARGs corresponds to E. coli corroborates with the presence of a higher number of E. coli bacteriophages. Bacteriophages were reported as the important contributor for the dissemination of ARGs. Conclusion Our findings suggest inevitable evidence that specific alterations of microbes residing in the gut are associated with the lifestyle changes; therefore, can be used for screening the UC cases. The pro-inflammatory nature of microorganisms may contribute to UC severity in the intestinal tract. The observed rural and urban differences are unlikely an artifact of the difference between their lifestyles; urbanization is actively transforming the gut microbiota of local communities at the expense of loss and decline of microbes associated with traditional lifestyles.

Physio-biochemical and molecular mechanisms of low nitrogen stress tolerance in peanut (Arachis hypogaea L.).

Nitrogen (N) is a major plant nutrient and its deficiency can arrest plant growth. However, how low-N stress impair plant growth and its related tolerance mechanisms in peanut seedlings has not yet been explored. To counteract this issue, a hydroponic study was conducted to explore low N stress (0.1mM NO3-) and normal (5.0mMNO3-) effects on the morpho-physiological and molecular attributes of peanut seedlings. Low-N stress significantly decreased peanut plant height, leaf surface area, total root length, and primary root length after 10days of treatment. Meanwhile, glutamate dehydrogenase, glutamine oxoglutarate aminotransferase activities, chlorophyll, and soluble protein contents were substantially decreased. Impairment in these parameters further suppressed photochemical efficiency (Fv/Fm), and chlorophyll fluorescence parameters (PIABS), under low-N stress. Transcriptome sequencing analysis showed a total of 2139 DEGs were identified between the two treatments. KEGG enrichment annotation analysis of DEGs revealed that 119 DEGs related to 10 pathways, including N assimilation, photosynthesis, starch, and sucrose degradation, which may respondto low-N stress in peanuts. Combined with transcriptome, small RNA, and degradome sequencing, we found that PC-3p-142756_56/A.T13EMM (CML3) and PC-5p-43940_274/A.81NSYN (YTH3) are the main modules contributing to low N stress tolerance in peanut crops. Peanut seedlings exposed to N starvation exhibited suppressed gene expression related to nitrate transport and assimilation, chlorophyll synthesis, and carbon assimilation, while also showing improved gene expression in N compensation/energy supply and carbohydrate consumption. Additionally, low N stress tolerance was strongly associated with the miRNA.

Branched oligosaccharides cause atypical starch granule initiation in Arabidopsis chloroplasts.

Plant chloroplasts store starch during the day, which acts as a source of carbohydrates and energy at night. Starch granule initiation relies on the elongation of malto-oligosaccharide primers. In Arabidopsis thaliana, PROTEIN TARGETING TO STARCH 2 (PTST2) and STARCH SYNTHASE 4 (SS4) are essential for the selective binding and elongation of malto-oligosaccharide primers, respectively, and very few granules are initiated in their absence. However, the precise origin and metabolism of the primers remain unknown. Potential origins of malto-oligosaccharide primers include de novo biosynthesis or their release from existing starch granules. For example, the endoamylase α-AMYLASE 3 (AMY3) can cleave a range of malto-oligosaccharides from the granule surface during starch degradation at night, some of which are branched. In the Arabidopsis double mutant deficient in the two debranching enzymes ISOAMYLASE 3 (ISA3) and LIMIT DEXTRINASE (LDA), branched malto-oligosaccharides accumulate in the chloroplast stroma. Here, we reveal that the isa3 lda double mutant shows a substantial increase in granule number per chloroplast, caused by these branched malto-oligosaccharides. The amy3 isa3 lda triple mutant, which lacks branched malto-oligosaccharides, has far fewer granules than isa3 lda, and its granule numbers are barely higher than in the wild type. Plants lacking both ISA3 and LDA and either PTST2 or SS4 show granule over-initiation, indicating that this process occurs independently of the recently described granule initiation pathway. Our findings provide insight into how and where starch granules are initiated. This knowledge can be used to alter granule number and morphological characteristics, traits known to affect starch properties.

The in-vitro digestibility of instant noodles: Interplay of texture, microstructure and starch structure.

Instant noodles are a worldwide food staple. However, the correlation between its production methods and nutritional characteristics remains unclear. This study aims to elucidate the effects of hydrothermal (steaming and boiling) and cooling techniques on instant noodles in-vitro digestibility. Texture of rehydrated noodles, microstructure observed by SEM and starch molecular structure measuring techniques were used to identify the underlying mechanisms. Our results demonstrate that boiling results in incomplete gelatinization of internal starch, leading to a microstructure with numerous pores and channels due to irregular starch distribution. This microstructure facilitates α-amylase diffusion and subsequent attachment to the substrate, yielding a higher starch digestion rate coefficient for boiled noodles (Nboiled) within the first 120min (K1, 5.7min-1) compared to steamed noodles (Nsteamed, K1, 4.4min-1). However, from 120 to 360min, Nboiled exhibited a lower rate coefficient (K2, 4.5min-1) due to the retarding effect of incompletely melted starch structure and greater texture rigidity, contrasting with Nsteamed (K2, 6.1min-1). Moreover, noodles subjected to boiling and subsequent cooling exhibited the lowest starch degradation by α-amylase throughout hydrolysis. This is attributed to their increased hardness, denser microstructure, and presence of ungelatinized or recrystallized starch. Therefore, the study concludes that cooking and cooling processes significantly influence noodle texture, microstructure, and starch structure, collectively impacting the nutritional value of instant noodles. Our findings provide valuable insights for the development of nutritionally enhanced instant noodles.

Upper circumpolar deep water influences microbial functional gene composition and diversity along the southern Central Indian Ridge and eastern Southwest Indian Ridge.

Deep sea microbial communities play a significant role in global biogeochemical processes. However, the depth-wise metabolic potential of microbial communities in hydrothermally influenced Central Indian Ridge (CIR) and Southwest Indian Ridge (SWIR) remains elusive. In this study, a comprehensive functional microarray-based approach was used to understand factors influencing the metabolic potential of microbial communities and depth-driven differences in microbial functional gene composition in CIR and SWIR. Stratified water column sampling at surface, mid, turbid/plume layer, and near bottom was done along with pertinent environmental variables at various locations along the ridges. The majority of genes (~38%-41%) throughout the water column in both regions encoded for C-cycling, particularly starch degradation indicating the predominance of heterotrophy. Genes encoding for nitrate reduction and arsenic and mercury resistance were enriched in the turbid and/or near-bottom waters, suggesting a localized influence of hydrothermally derived substrates on the metabolic potential of microbial communities. Indices for microbial functional gene diversity (H = 9.18) and evenness (J = 0.90) were highest for samples from turbid waters at SWIR. Potential temperature-salinity profiles showed the presence of nutrient-rich upper circumpolar deep water (UCDW) at >2,000 m in the study areas. Principal component analysis revealed that inorganic nutrient availability largely influenced functional gene diversity in deeper waters. The study signifies that rather than hydrothermal input, nutrients brought into the region through the UCDW could have a larger impact on metabolic processes mediated by autochthonous microbial communities and consequently have implications on deep-sea productivity.IMPORTANCELittle is known about depth-wise metabolic potential of microbial communities in hydrothermally influenced Central Indian Ridge (CIR) and Southwest Indian Ridge (SWIR) waters. In the present study, a comprehensive functional gene microarray approach was used to reveal the metabolic potential and depth-wise variation in microbial functional genes along the ridges. Up to 41% of microbial functional genes at both locations encoded for C-cycling. Availability of hydrothermally derived substrates in plumes detected along the ridges triggered an increase in the abundance of genes encoding for remediation of polycyclic aromatics, nitrate reduction, and arsenic and mercury resistance. Rather than hydrothermal input, the functional gene diversity at >2,000 m was largely influenced by inorganic nutrients transported by the nutrient-rich upper circumpolar deep water. Findings of this study are expanding the existing knowledge on new sites of hydrothermal activity along CIR and SWIR and gaining insights into ecosystem functioning in the deep sea.

Construction of a recombinant Bacillus subtilis strain expressing SpaA and CbpB of Erysipelothrix rhusiopathiae and evaluation of the strain immunogenicity in a mouse model

To construct a recombinant Bacillus subtilis strain expressing SpaA and CbpB of Erysipelothrix rhusiopathiae for oral administration, we constructed the recombinant plasmid pDG1730-CBJA by fusion PCR and seamless cloning. The plasmid was introduced into B. subtilis KC strain by natural transformation, and the recombinant strain KC-spaA-cbpB was screened out on the plate containing spectinomycin (sper) and confirmed by PCR and starch degradation test. The SpaA and CbpB expressed by KC-spaA-cbpB were detected by Western blotting and indirect immunofluorescence assay, and the genetic stability of the recombinant strain in mice was determined. The plasmid pMAD-∆sper with knockout of sper was constructed and transformed into KC-spaA-cbpB. The sper-deleted mutant strain KC-spaA-cbpB: : ∆sper was screened and identified, and its immunogenicity in a mouse model was evaluated by oral immunization. The results showed that the recombinant strain KC-spaA-cbpB was stable in mice, expressing SpaA on the cell surface and CbpB on the spore surface. KC-spaA-cbpB: : ∆sper expressed SpaA and CbpB. The mice vaccinated with the spores of KC-spaA-cbpB: : ∆sper had higher levels of SpaA and CbpB-specific IgG in the serum that those vaccinated with the wild-type spores 42 days after vaccination by gavage (P < 0.01). The protective rate of mice immunized with the recombinant spores was 67.5%. The results indicated that a recombinant B. subtilis strain expressing SpaA and CbpB of E. rhusiopathiae was successfully constructed, and the recombinant strain laid a foundation for the development of oral live vector vaccines for swine erysipelas.

The Effects of Mixed Inoculum Storage Time on In Vitro Rumen Fermentation Characteristics, Microbial Diversity, and Community Composition.

This study aimed to investigate the effects of different storage times of the mixed inoculum on in vitro rumen fermentation characteristics, microbial diversity, and community composition. The experiment was divided into five groups, with mixed inoculum composed of fresh rumen fluid and culture medium being stored at 39 °C for 0 h (H0), 12 h (H12), 24 h (H24), 36 h (H36), and 48 h (H48). After 48 h of in vitro fermentation, the fermentation fluid was collected to assess rumen fermentation characteristics and microbial community composition. The H24 group showed higher total gas production, ammoniacal nitrogen levels, and total volatile fatty acids, as well as higher concentrations of individual volatile fatty acids except propionate, compared to the H0 and H48 groups (p < 0.05). The Shannon and Simpson evenness indices were significantly higher in the H0, H12, and H24 groups than in the H48 group (p < 0.05). A total of nine phyla and sixteen genera involved in starch and fiber degradation were found to be more abundant in the H24 or H48 groups (p < 0.05). Moreover, nine predicted metabolic pathways were observed to be significantly enriched in either the H24 or H48 group (p < 0.05). Both principal coordinates analysis (PCoA) and non-metric multidimensional scaling (NMDS) analysis revealed distinct clustering patterns among the H0, H12, H24, H36, and H48 groups, and analysis of similarities (ANOSIM) confirmed these significant differences (R = 1.00, p < 0.05). This study demonstrates that the storage time of mixed inoculum influences rumen fermentation characteristics and microbial community composition in a time-dependent manner. It is recommended to use a mixed inoculum that has been stored within 24 h in an anaerobic environment at 39 °C for in vitro rumen fermentation tests. This study offers valuable microbial insights into the storage strategies for mixed inoculum, thereby improving the methodologies for variable control in in vitro rumen fermentation techniques.

Microbial functional genes play crucial roles in enhancing soil nutrient availability of halophyte rhizospheres in salinized grasslands.

Land degradation due to salinization threatens ecosystem health. Phytoremediation, facilitated by functional microorganisms, has gained attention for improving saline-alkali soils. However, the relationship between the functional potential of rhizosphere microbes involved in multi-element cycling and soil nutrient pools remain unclear. This study focused on the changes in functional genes related to carbon (C), nitrogen (N), and phosphorus (P) cycling in the rhizospheres of various halophytes and bulk soil in the grassland ecosystem of Chaka Salt Lake, Qinghai Province, China. Our evaluation of plant and soil characteristics revealed that halophyte growth increased soil hydrolase activity and nutrient levels, particularly available N. Significant differences were observed in foliage and root nutrients, rhizosphere soil properties, and microbial functional gene composition among plant species. Halophytes significantly altered the abundance of genes involved in C fixation (Calvin and DC/4-HB cycles), C degradation (starch, hemicellulose, cellulose, and pectin degradation), dissimilatory nitrate reduction (nirB), ammonification (ureC), organic P mineralization (phoA and ugpQ), P transport (phnE), and inorganic P dissolution (ppk1). C, N, and P cycling processes were closely related to soil N nutrients, available nutrient ratios, and C/N-cycling enzyme activities. Partial least squares path modeling (PLS-PM) analysis showed that microbial functional genes were directly associated with soil nutrient availability, with soil and plant variables indirectly affecting nutrient pools through the regulation of these genes. These findings enhance our understanding of the biochemical cycling in halophyte rhizospheres and highlight the role of microbial functional genes in saline-alkali soil restoration.

Transcriptomic analysis reveals three important carbohydrate-active enzymes contributing to starch degradation of oleaginous yeast Lipomyces starkeyi.

The oleaginous yeast Lipomyces starkeyi has a high capacity for starch assimilation, but the genes involved and specific mechanisms in starch degradation remain unclear. This study aimed to identify the critical carbohydrate-active enzyme (CAZyme) genes contributing to starch degradation in L. starkeyi. Comparative transcriptome analysis of cells cultured in glucose and soluble starch medium revealed that 55 CAZymes (including transcript IDs 3772, 1803, and 7314) were highly expressed in soluble starch medium. Protein domain structure and disruption mutant analyses revealed that 3772 encodes the sole secreted α-amylase (LsAmy1p), whereas 1803 and 7314 encode secreted α-glucosidase (LsAgd1p and LsAgd2p, respectively). Triple-gene disruption exhibited severely impaired growth in soluble starch, dextrin, and raw starch media, highlighting their critical role in degrading polysaccharides composed of glucose linked by α-1,4-glucosidic bonds. This study provided insights into the complex starch degradation mechanism in L. starkeyi.

Utilizing isotopic and elemental markers to enhance the authenticity of potatoes

AbstractGiven the economic importance of potato production, establishing the origin of unknown commercial samples declared as local, is particularly important for both producers and competent authorities. In the study presented here, stable isotopic ratios of deuterium, oxygen, carbon and nitrogen, were determined in order to develop a potato provenance methodology. Isotopic determinations were conducted in an alcohol, preceded by the enzymatic degradation of potato starch using α-amylase to yield soluble dextrins and oligosaccharides. Subsequently, the hydrolyzed potato starch underwent fermentation to produce ethanol, which was then obtained through distillation under controlled conditions. The integration of data obtained from analyzing the D/H isotopic ratios using SNIF-NMR spectroscopy in potato samples for the first time, in conjunction with measurements of the 13C/12C, 15N/14N, and 18O/16O ratios via IRMS, yielded a unique isotopic fingerprint for the potato samples under examination. Elemental analysis by ICP-OES also added important information in the dataset. The chemometric analysis by applying OPLS-DA technique, highlighted the parameters of (D/H)II, δ18O, R and Cu as the best discriminator markers. The proposed model gave 94.07% correct classification of the samples, regarding their geographical origin. It is believed that the differentiation of local potatoes is related to the unique geological and climatic conditions existing in the island. All the above-mentioned conclusions are very promising, for the protection of local potato production.

Deciphering heat wave effects on wheat grain: focusing on the starch fraction.

Wheat is an essential staple food, and its production and grain quality are affected by extreme temperature events. These effects are even more relevant considering the increasing food demand for a growing world population and the predicted augmented frequency of heat waves. This study investigated the impact of simulated heat wave (HW) conditions imposed during grain filling on starch granule characteristics, endosperm ultrastructure, and transcriptomic modulation of genes involved in starch synthesis and degradation. All these evaluations were performed with four different genotypes, two commercial wheat varieties (Antequera and Bancal), and two traditional landraces (Ardito and Magueija). Starch granule size distribution and shape were significantly altered by HW treatment, revealing an increase of A-type granules in Ardito and an opposite effect in Magueija and Bancal, while Antequera remained stable. Analysis of the largest (LD) and smallest (SD) granule diameters also revealed genotype-specific changes, with Magueija showing a shift toward more spherical A-type granules after the HW treatment. Scanning electron microscopy confirmed alterations in endosperm morphology, including increased vitreousness in Bancal and substantial increase of endosperm cavities and grain size reduction in Magueija under HW stress. The transcriptomic analysis confirmed the stability of Antequera under HW, in contrast with the other genotypes where differential gene expression related to starch metabolism was detected. These effects were particularly severe in Magueija with the downregulation of genes encoding for enzymes involved in amylopectin synthesis (both starch synthases and starch-branching enzyme) and upregulation of α-amylase-encoding genes. These findings contribute to the understanding of heat stress effects on wheat grain quality, emphasize the importance of genetic diversity in HW responses, and suggest potential avenues for breeding climate-resilient wheat varieties.

Diversity and Potential Metabolic Characteristics of Culturable Copiotrophic Bacteria That Can Grow on Low-Nutrient Medium in Zhenbei Seamount in the South China Sea

Oligotrophs are predominant in nutrient-poor environments, but copiotrophic bacteria may tolerate conditions of low energy and can also survive and thrive in these nutrient-limited conditions. In the present study, we isolated 648 strains using a dilution plating method after enrichment for low-nutrient conditions. We collected 150 seawater samples at 21 stations in different parts of the water column at the Zhenbei Seamount in the South China Sea. The 648 isolated copiotrophic strains that could grow on low-nutrient medium were in 21 genera and 42 species. A total of 99.4% (644/648) of the bacteria were in the phylum Pseudomonadota, with 73.3% (472/644) in the class Gammaproteobacteria and 26.7% (172/644) in the class Alphaproteobacteria. Among the 42 representative isolates, Pseudoalteromonas arabiensis, Roseibium aggregatum, and Vibrio neocaledonicus were present in all layers of seawater and at almost all of the stations. Almost half of these species (20/42) contained genes that performed nitrate reduction, with confirmation by nitrate reduction testing. These isolates also contained genes that functioned in sulfur metabolism, including sulfate reduction, thiosulfate oxidation, thiosulfate disproportionation, and dimethylsulfoniopropionate degradation. GH23, CBM50, GT4, GT2, and GT51 were the main carbohydrate-active enzymes (CAZymes), and these five enzymes were present in all or almost all of the isolated strains. The most abundant classes of CAZymes were those associated with the degradation of chitin, starch, and cellulose. Collectively, our study of marine copiotrophic bacteria capable of growing on low-nutrient medium demonstrated the diversity of these species and their potential metabolic characteristics.

Improvement of overall quality of black rice by stabilization combined with solid-state fermentation

The application of black rice in food industry is challenging due to its short shelf life, poor cooking and eating quality. Superheated steam (SS) and far infrared radiation (FIR) are effective ways of stabilization and might be beneficial to solid-state fermentation (SSF). Therefore, stabilization treatments (SS and FIR) combined with SSF by Aspergillus oryzae were applied to furtherly improve the overall quality of black rice in present study. Results indicated that combined treatments were more favorable to shorten cooking time, increase water absorption and solid loss, resulting improvement of cooking quality of black rice. Besides, the significant decrease in hardness and chewiness after combined treatments was also good for the texture improvement. Moreover, black rice after combined treatments showed significantly higher contents of free phenolics, flavonoids, antioxidant activities, and phenolic acids, while their contents in bound form were significantly decreased, leading to significant increase of nutritional properties. Furthermore, black rice after combined treatments had significantly lower acid value and peroxide value during accelerated storage. Particularly, far infrared radiation combined with SSF was the most valued to improve the overall quality of black rice. These improvements were mainly ascribed to the enhancement of hydrolase activities based on the synergistic effect between stabilization treatments and SSF, which led to more significant pasting viscosities decrease, crystal structure destruction, cell wall rupture and starch degradation. The above results will provide a favorable strategy to improve the overall quality of black rice and then promote its consumption and application in food industry. Industrial relevanceThe short shelf life, poor cooking and eating quality severely limit the utilities of black rice in food industry. Heat processing is one of the most efficient strategies used in food industry to improve quality and prolong shelf life. Superheated steam and far infrared radiation are typical innovative heating technologies, and they are more energy efficient and more effective than conventional heating methods. In our previous work, they were effective ways of stabilization to extend the shelf life of black rice, and the starch after above treatments was partially gelatinized, which might be beneficial to the solid-state fermentation and thereby furtherly improve the overall quality of black rice. The present study aims to provide a favorable strategy to improve the overall quality of black rice and then promote its consumption and application in food industry.

Precision Activity-Based α-Amylase Probes for Dissection and Annotation of Linear and Branched-Chain Starch-Degrading Enzymes.

α-Amylases are the workhorse enzymes of starch degradation. They are central to human health, including as targets for anti-diabetic compounds, but are also the key enzymes in the industrial processing of starch for biofuels, corn syrups, brewing and detergents. Dissection of the activity, specificity and stability of α-amylases is crucial to understanding their biology and allowing their exploitation. Yet, functional characterization lags behind DNA sequencing and genomics; and new tools are required for rapid analysis of α-amylase function. Here, we design, synthesize and apply new branched α-amylase activity-based probes. Using both α-1,6 branched and unbranched α-1,4 maltobiose activity-based probes we were able to explore the stability and substrate specificity of both a panel of human gut microbial α-amylases and a panel of industrially relevant α-amylases. We also demonstrate how we can detect and annotate the substrate specificity of α-amylases in the complex cell lysate of both a prominent gut microbe and a diverse compost sample by in-gel fluorescence and mass spectrometry. A toolbox of starch-active activity-based probes will enable rapid functional dissection of α-amylases. We envisage activity-based probes contributing to better selection and engineering of enzymes for industrial application as well as fundamental analysis of enzymes in human health.

An Integrative Analysis of the Transcriptome and Proteome of Rice Grain Chalkiness Formation Under High Temperature.

Exposure to high temperatures can impair the grain-filling process in rice (Oryza sativa L.), potentially leading to the formation of chalky endosperm, but the molecular regulation mechanism remains largely elusive. Here, we reported that high-temperature (HT) stress (day/night, 35 °C/30 °C) reduces both the grain-filling rate and grain weight of Ningjing 1 variety compared to normal temperatures (NT, day/night, 28 °C/23 °C). Grains under HT stress exhibited an opaque, milky-white appearance, alongside significant alterations in starch physicochemical properties. An integrated transcriptomic analysis of grains under HT revealed up-regulation of genes related to defense mechanisms and oxidoreductase activity, while genes involved in sucrose and starch synthesis were down-regulated, and α-amylase genes were up-regulated. Proteomic analysis of grains under HT echoed this pattern. These results demonstrate that high temperature during the grain-filling stage significantly increases rice chalkiness by down-regulating genes related to sucrose and starch synthesis, while up-regulating those involved in starch degradation.

Identification of two postharvest ripening regulatory models in kiwifruit: based on plant hormones, physiology, and transcriptome analysis

Kiwifruit (Actinidia spp.), celebrated for its unique flavor and rich nutritional content, is a globally popular fruit. This fruit requires post-harvest ripening before consumption. However, the unpredictable ripening pace significantly impacts consumer acceptance and sales, thereby hindering the commercial growth of kiwifruit. To address this, understanding the key molecular mechanisms and metabolites governing postharvest ripening and senescence could offer valuable insights for developing storage strategies and breeding techniques in yellow-fleshed kiwifruits. We constructed two models that integrated these findings with existing theories. The first model suggests that, unlike the T6P-sucrose regulatory mechanism observed in plant leaves, the separation of harvested kiwifruit from the mother plant leads to an insufficient supply of T6P, which activates the SnRK1 kinase. This, in turn, inhibits the TOR kinase signaling pathway, regulating starch metabolism. The T6P-SnRK1-TOR-starch metabolism pathway plays a regulatory role during postharvest ripening, limiting excessive starch degradation that could accelerate aging and decay in yellow-fleshed kiwifruit. The second model highlights the role of abscisic acid (ABA), cytokinins (CKs), and ethylene in regulating the process, inducing the activation of ERFs and cell wall-degrading enzymes, promoting fruit postharvest softening. These findings indicate that at least two models, the T6P-SnRK1-TOR-starch metabolism model and the ABA-CKs-ethylene-cell wall degradation model, regulate postharvest fruit ripening, offering new insights into the artificial regulation of yellow-fleshed kiwifruit ripening speed.