Alkaline Stress Research Articles

Alkaline stress inhibits the growth of Staphylococcus epidermidis by inducing TCA cycle-triggered ROS production

Many species of bacteria interact on the human skin to form a certain microbiome. Delftia acidovorans, a bacterium detected from human skin, inhibits the growth of S. epidermidis, a dominant bacterium of the human skin microbiota. Here, we show that ammonia secreted by D. acidovorans inhibits the growth of S. epidermidis by increasing the pH value of the medium. The pH value of D. acidovorans culture supernatant (CS) was higher than that of the medium without culture. The inhibitory activity of the D. acidovorans CS against the growth of S. epidermidis was decreased by neutralization with hydrochloric acid. Genes encoding enzymes related to ammonia production were found in the D. acidovorans genome. Moreover, the D. acidovorans CS contained a high concentration of ammonia. The addition of ammonia to S. epidermidis culture led to an increase in the reactive oxygen species (ROS) production and inhibited S. epidermidis growth. The addition of sodium hydroxide also led to an increase in the ROS production and inhibited S. epidermidis growth. The inhibitory activity of ammonia and sodium hydroxide against S. epidermidis growth was suppressed by malonic acid, an inhibitor of succinate dehydrogenase in the tricarboxylic acid (TCA) cycle, and N-acetyl-l-cysteine, a free radical scavenger. These findings suggest that D. acidovorans secretes ammonia and alkaline stress inhibits the growth of S. epidermidis by inducing TCA cycle-triggered ROS production.

The Chlamydomonas bZIP transcription factor BLZ8 confers oxidative stress tolerance by inducing the carbon-concentrating mechanism.

Photosynthetic organisms are exposed to various environmental sources of oxidative stress. Land plants have diverse mechanisms to withstand oxidative stress, but how microalgae do so remains unclear. Here, we characterized the Chlamydomonas reinhardtii basic leucine zipper (bZIP) transcription factor BLZ8, which is highly induced by oxidative stress. Oxidative stress tolerance increased with increasing BLZ8 expression levels. BLZ8 regulated the expression of genes likely involved in the carbon-concentrating mechanism (CCM): HIGH-LIGHT ACTIVATED 3 (HLA3), CARBONIC ANHYDRASE 7 (CAH7), and CARBONIC ANHYDRASE 8 (CAH8). BLZ8 expression increased the photosynthetic affinity for inorganic carbon under alkaline stress conditions, suggesting that BLZ8 induces the CCM. BLZ8 expression also increased the photosynthetic linear electron transfer rate, reducing the excitation pressure of the photosynthetic electron transport chain and in turn suppressing reactive oxygen species (ROS) production under oxidative stress conditions. A carbonic anhydrase inhibitor, ethoxzolamide, abolished the enhanced tolerance to alkaline stress conferred by BLZ8 overexpression. BLZ8 directly regulated the expression of the three target genes and required bZIP2 as a dimerization partner in activating CAH8 and HLA3. Our results suggest that a CCM-mediated increase in the CO2 supply for photosynthesis is critical to minimize oxidative damage in microalgae, since slow gas diffusion in aqueous environments limits CO2 availability for photosynthesis, which can trigger ROS formation.

Untargeted metabolomics of the alkaliphilic cyanobacterium Plectonema terebrans elucidated novel stress-responsive metabolic modulations

Alkaliphilic cyanobacteria are suitable candidates to study the effect of alkaline wastewater cultivation on molecular metabolic responses. In the present study, the impact of wastewater, alkalinity, and alkaline wastewater cultivation was studied on the biomass production, biochemical composition, and the alkalinity responsive molecular mechanism through metabolomics. The results suggested a 1.29 to 1.44-fold higher biomass production along with improved lipid, carbohydrate, and pigment production under alkaline wastewater cultivation. The metabolomics analysis showed 1.2-fold and 5.54-fold increase in the indole-acetic acid and phytoene biosynthesis which contributed to overall enhanced cell differentiation and photo-protectiveness. Furthermore, lower levels of Ribulose-1,5-bisphosphate (RuBP), and higher levels of 2-phosphoglycerate and 3-phosphoglycerate suggested the efficient fixation of CO2 into biomass, and storage compounds including polysaccharides, lipids, and sterols. Interestingly, except L-histidine and L-phenylalanine, all the metabolites related to protein biosynthesis were downregulated in response to wastewater and alkaline wastewater cultivation. The cells protected themselves from alkalinity and nutrient stress by improving the biosynthesis of sterols, non-toxic antioxidants, and osmo-protectants. Alkaline wastewater cultivation regulated the activation of carbon concentration mechanism (CCM), glycolysis, fatty-acid biosynthesis, and shikimate pathway. The data revealed the importance of alkaline wastewater cultivation for improved CO2 fixation, wastewater treatment, and producing valuable bioproducts including phytoene, Lyso PC 18:0, and sterols. These metabolic pathways could be future targets of metabolic engineering for improving biomass and metabolite production. SignificanceAlkalinity is an imperative factor, responsible for the contamination control and biochemical regulation in cyanobactera, especially during the wastewater cultivation. Currently, understanding of alkaline wastewater responsive molecular mechanism is lacking and most of the studies are focused on transcriptomics of model organisms for this purpose. In this study, untargeted metabolomics was employed to analyze the impact of wastewater and alkaline wastewater on the growth, CO2 assimilation, nutrient uptake, and associated metabolic modulations of the alkaliphilic cyanobacterium Plectonema terebrans BERC10. Results unveiled that alkaline wastewater cultivation regulated the activation of carbon concentration mechanism (CCM), glycolysis, fatty-acid biosynthesis, and shikimate pathway. It indicated the feasibility of alkaline wastewater as promising low-cost media for cyanobacterium cultivation. The identified stress-responsive pathways could be future genetic targets for strain improvement.

Validation of Analytical Methods for Tacrolimus Determination in Poly(ε-caprolactone) Nanocapsules and Identification of Drug Degradation Products.

The aim of this paper was to use chromatographic tools for validating an analytical method for the tacrolimus (TAC) determination in polymeric nanocapsules and for identifying the drug degradation products after alkaline stress. A rapid Ultra-High-Performance Liquid Chromatography coupled with photo-diode array (UHPLC-PDA) method was successfully performed using the following chromatographic conditions: the Shimadzu Shim-pack XR-ODS III C18 column (100 mm×2.00 mm, 2.2 μm), the mobile phase consisting of methanol and acidified ultrapure water (89:11 v/v), the flow rate of 0.55 mL·min-1, and the ultraviolet (UV) detection at 235 nm. This method was validated as per International Council for Harmonisation (ICH) guidelines. In addition, a TAC forced degradation assay was carried out after alkaline stress and its degradation products were investigated using Liquid Chromatography coupled tandem mass spectroscopy (LC-MS/MS). The calibration curve was linear in the range of 100.0-300.0 μg·mL-1 (r >0.9999). Accuracy was confirmed by the TAC recovery of 96.55 to 98.19%. Precision (intraday and interday) were demonstrated by relative standard deviation lower than 0.89% and 3.25%, respectively. Selectivity and robustness were also proved. The method developed it was successfully applied to quantify TAC from polymeric nanocapsules, showing a high loading efficiency rate (>96.47%). The main drug degradation product observed in a multiple reaction monitoring (MRM) experiment was m/z 844, confirming the susceptibility of TAC under alkaline conditions; this finding was first time described.

Stability Indicating Method for Known and Unknown Impurities Profiling for Vildagliptin in Vildagliptin Tablet

Background: Vildagliptin is a drug for the treatment of diabetes. DPP-IV inhibitor represents a new class of oral antihyperglycemic agents to treat patients with type 2 diabetes. Several RP-HPLC methods have been reported to determine Vildagliptin alone. However, it has been noted that there are no available stability-indicating methods in pharmacopeias (USP/BP/EP/JP) nor in the available literature to quantify known and unknown impurity patterns for vildagliptin in vildagliptin tablets. Objective: The aim of this study is to develop a new single, sensitive, robust and specific gradient RP-HPLC method to quantify known and unknown impurities and degradants of Vildagliptin in Vildagliptin tablets. Methods: Chromatographic separation has been accomplished on the Hypersil ODS column (250 x 4.6) mm, 5 μm with a mobile phase consisting of a mixture of Perchloric acid Buffer, methanol, acetonitrile and Triethylamine delivered at a flow rate of 1.0 mL minute-1 and the detection wavelength 210 nm. The developed method was validated as per ICH guidelines. Results: Vildagliptin was found degraded significantly under oxidative and alkaline stress conditions. The degradation products were well resolved from Vildagliptin and its impurities. An analytical method was found linear, accurate and precise from LOQ (Limit of Quantification) level to 150% of impurity specification limit (0.5%). Conclusion: The method found sensitive, rapid and accurate quantification of known, unknown impurities and degradants. The peak purity results confirmed that the Vildagliptin peak was homogeneous and pure in all stress samples, thus proving the stability-indicating nature of the method.

HSP90 and HSP70 Families in Lateolabrax maculatus: Genome-Wide Identification, Molecular Characterization, and Expression Profiles in Response to Various Environmental Stressors.

Heat shock proteins (HSPs) are a large class of highly conserved chaperons, which play important roles in response to elevated temperature and other environmental stressors. In the present study, 5 HSP90 genes and 17 HSP70 genes were systematically characterized in spotted seabass (Lateolabrax maculatus). The evolutionary footprint of HSP genes was revealed via the analysis of phylogeny, chromosome location, and gene copy numbers. In addition, the gene structure features and the putative distribution of heat shock elements (HSEs) and hypoxia response elements (HREs) in the promoter regions were analyzed. The protein-protein interaction (PPI) network analyses results indicated the potential transcriptional regulation between the heat shock factor 1 (HSF1) and HSPs and a wide range of interactions among HSPs. Furthermore, quantitative (q)PCR was performed to detect the expression profiles of HSP90 and HSP70 genes in gill, liver, and muscle tissues after heat stress, meanwhile, the expression patterns in gills under alkalinity and hypoxia stresses were determined by analyzing RNA-Seq datasets. Results showed that after heat stress, most of the examined HSP genes were significantly upregulated in a tissue-specific and time-dependent manners, and hsp90aa1.1, hsp90aa1.2, hsp70.1, and hsp70.2 were the most intense responsive genes in all three tissues. In response to alkalinity stress, 11 out of 13 significantly regulated HSP genes exhibited suppressed expression patterns. Alternatively, among the 12 hypoxia-responsive-expressed HSP genes, 7 genes showed induced expressions, while hsp90aa1.2, hsp70.1, and hsp70.2 had more significant upregulated changes after hypoxic challenge. Our findings provide the essential basis for further functional studies of HSP genes in response to abiotic stresses in spotted seabass.

Physiological and transcriptomic analyses reveal novel insights into the cultivar-specific response to alkaline stress in alfalfa (Medicago sativa L.)

Soil alkalization severely limits plant growth and development, however, the mechanisms of alkaline response in plants remain largely unknown. In this study, we performed physiological and transcriptomic analyses using two alfalfa cultivars (Medicago sativa L.) with different sensitivities to alkaline conditions. The chlorophyll content and shoot fresh mass drastically declined in the alkaline-sensitive cultivar Algonquin (AG) following alkaline treatment (0–25 mM Na2CO3 solution), while the alkaline-tolerant cultivar Gongnong NO.1 (GN) maintained relatively stable growth and chlorophyll content. Compared with AG, GN had higher contents of Ca2+ and Mg2+; the ratios of Ca2+ and Mg2+ to Na+, proline and soluble sugar, as well as higher enzyme activities of peroxidase (POD) and catalase (CAT) under the alkaline conditions. Furthermore, transcriptomic analysis identified three categories of alkaline-responsive differentially expressed genes (DEGs) between the two cultivars: 48 genes commonly induced in both the cultivars (CAR), 574 genes from the tolerant cultivar (TAR), and 493 genes from the sensitive cultivar (SAR). Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses showed that CAR genes were mostly involved in phenylpropanoid biosynthesis, lipid metabolism, and DNA replication and repair; TAR genes were significantly enriched in metabolic pathways, such as biosynthesis of amino acids and secondary metabolites including flavonoids, and the MAPK signaling pathway; SAR genes were specifically enriched in vitamin B6 metabolism. Taken together, the results identified candidate pathways associated with genetic variation in response to alkaline stress, providing novel insights into the mechanisms underlying alkaline tolerance in alfalfa.

Characterization of rpp5cs and Rpoat From Rumex Patientia L. In Response to Salinity and Alkalinity Stress

Rumex patientia L. can respond to abiotic stresses and withstand low temperatures. Proline accumulation plays important roles under various abiotic stresses in different plants, specially salinity. P5CS and OAT of plants are key enzymes in the proline synthesis pathway. In the present study, the RpP5cs and RpOat genes of the proline synthesis pathway in R. patientia L. were cloned with full-length cDNAs of 2178 bp and 1401 bp. The expression of RpP5cs and RpOat genes were higher in Russian Щавель Чемпион than Chinese R. patientia × R. tianschanicus cv. Rumex under salinity and alkalinity stress over 0 - 24 hrs. These results could provide insights into sequence structure and differential tolerance in many plants. Bangladesh J. Bot. 50(3): 717-727, 2021 (September) Special

The role of 14-3-3 proteins in plant growth and response to abiotic stress.

The 14-3-3 proteins widely exist in almost all plant species. They specifically recognize and interact with phosphorylated target proteins, including protein kinases, phosphatases, transcription factors and functional proteins, offering an array of opportunities for 14-3-3s to participate in the signal transduction processes. 14-3-3s are multigene families and can form homo- and heterodimers, which confer functional specificity of 14-3-3 proteins. They are widely involved in regulating biochemical and cellular processes and plant growth and development, including cell elongation and division, seed germination, vegetative and reproductive growth, and seed dormancy. They mediate plant response to environmental stresses such as salt, alkaline, osmotic, drought, cold and other abiotic stresses, partially via hormone-related signalling pathways. Although many studies have reviewed the function of 14-3-3 proteins, recent research on plant 14-3-3s has achieved significant advances. Here, we provide a comprehensive overview of the fundamental properties of 14-3-3 proteins and systematically summarize and dissect the emerging advances in understanding the roles of 14-3-3s in plant growth and development and abiotic stress responses. Some ambiguous questions about the roles of 14-3-3s under environmental stresses are reviewed. Interesting questions related to plant 14-3-3 functions that remain to be elucidated are also discussed.

Roles of Canavalia rosea metallothioneins in metal tolerance and extreme environmental adaptation to tropical coral reefs

Canavalia rosea (Sw.) DC is a perennial twining herb distributed in the semi-arid and saline–alkali areas of coastal regions and has evolved halotolerance. In this study, we present the first comprehensive survey of the metallothionein (MT) gene family in C. rosea. MT proteins belong to a family of low-molecular-weight polypeptides with a high content of cysteine residues, which have an affinity to bind with heavy metal ions. MTs also play important roles in stress responses as reactive oxygen species (ROS) scavengers. A total of six CrMTs were identified in the C. rosea genome and classified into four subgroups by phylogenetic analysis. An analysis of the cis-acting elements revealed that a series of hormone-, stress-, and development-related cis-acting elements were present in the promoter regions of CrMTs. The expression of CrMTs also showed habitat- and environmental stress-regulated patterns in C. rosea. CrMT overexpression in yeast enhanced tolerance to heavy metals and ROS, as well as high osmotic and alkalinity stress, which is consistent with their predicted roles as metal-chelating proteins and ROS scavengers. Our results indicate that the CrMT genes might contribute to the detoxification of plants to metals and provide marked tolerance against abiotic stress. The expression patterns of CrMTs in C. rosea also indicate that CrMTs play important roles in this species’ response to extreme environments on tropical islands and reefs, probably by improving the thermotolerance of C. rosea plants.

Genome-Wide Identification, Characterization and Expression Analysis of Soybean CHYR Gene Family.

The CHYR (CHY ZINC-FINGER AND RING FINGER PROTEIN) proteins have been functionally characterized in iron regulation and stress response in Arabidopsis, rice and Populus. However, their roles in soybean have not yet been systematically investigated. Here, in this study, 16 GmCHYR genes with conserved Zinc_ribbon, CHY zinc finger and Ring finger domains were obtained and divided into three groups. Moreover, additional 2–3 hemerythrin domains could be found in the N terminus of Group III. Phylogenetic and homology analysis of CHYRs in green plants indicated that three groups might originate from different ancestors. Expectedly, GmCHYR genes shared similar conserved domains/motifs distribution within the same group. Gene expression analysis uncovered their special expression patterns in different soybean tissues/organs and under various abiotic stresses. Group I and II members were mainly involved in salt and alkaline stresses. The expression of Group III members was induced/repressed by dehydration, salt and alkaline stresses, indicating their diverse roles in response to abiotic stress. In conclusion, our work will benefit for further revealing the biological roles of GmCHYRs.

Insight Between the Epigenetics and Transcription Responding of Cotton Hypocotyl Cellular Elongation Under Salt-Alkaline Stress.

Gossypium barbadense is a cultivated cotton not only known for producing superior fiber but also for its salt and alkaline resistance. Here, we used Whole Genome Bisulfite Sequencing (WGBS) technology to map the cytosine methylation of the whole genome of the G. barbadense hypocotyl at single base resolution. The methylation sequencing results showed that the mapping rates of the three samples were 75.32, 77.54, and 77.94%, respectively. In addition, the Bisulfite Sequence (BS) conversion rate was 99.78%. Approximately 71.03, 53.87, and 6.26% of the cytosine were methylated at CG, CHG, and CHH sequence contexts, respectively. A comprehensive analysis of DNA methylation and transcriptome data showed that the methylation level of the promoter region was a positive correlation in the CHH context. Saline-alkaline stress was related to the methylation changes of many genes, transcription factors (TFs) and transposable elements (TEs), respectively. We explored the regulatory mechanism of DNA methylation in response to salt and alkaline stress during cotton hypocotyl elongation. Our data shed light into the relationship of methylation regulation at the germination stage of G. barbadense hypocotyl cell elongation and salt-alkali treatment. The results of this research help understand the early growth regulation mechanism of G. barbadense in response to abiotic stress.

Nutritional responses of grafted cucumber on two types of Iranian local squash to alkalinity and salinity stresses

High bicarbonate (HCO3 -) content and associated high pH of irrigation water are detrimental to plant nutrient absorption. This experiment was arranged as a factorial in the framework of a completely randomized design with two factors, rootstock (ungrafted cucumber, Cucurbita moschata, and Cucurbita maxima) and bicarbonate (10 mM NaHCO3 or AS and 10 mM NaCl or SS). Regardless of rootstock, bicarbonate increased the pH of nutrient solution in all treatments. Under NaHCO3 treatment, the uptake and leaf concentration of K and Fe, and Mn uptake were not affected in plants grafted onto C. moschata rootstocks, whereas a significant decrease of them recorded in the control plants. Although the effect of alkalinity and rootstocks on Mg concentration was not significant, the highest Mg uptake observed in plants grafted onto C. moschata rootstocks. It is concluded that using C. moschata rootstock could reduce the deleterious effects of alkalinity stress on nutrient uptake by cucumber plants.

Genome-wide identification of IQ67 domain (IQD) gene families in Chinese jujube (Ziziphus jujuba Mill.) and expression profiles in response to cold stress

The plant-specific IQ67 DOMAIN (IQD) gene family is a possible targeted interacting protein of calmodulin that plays important roles in calcium signaling transduction and that functions in various biotic and abiotic stresses. However, no systematic analysis of the IQD gene family has been performed in Chinese jujube (Ziziphus jujuba Mill.). In the current study, 20 ZjIQDs were first identified in Chinese jujube and found to be evenly distributed on 12 chromosomes (chr) except chr 9 through a series of bioinformatic analyses. Phylogenetic analysis of 20 ZjIQDs with the IQDs in rice and Arabidopsis showed that all of the IQDs could be classified into four groups and that each group could be divided into two or three other subgroups. In addition, gene structure and motif composition analysis showed that 20 ZjIQDs had similar/respective gene structures in the same group and that the IQ calmodulin binding motif was conserved. Prediction of calmodulin binding sites revealed that all of the ZjIQDs contained at least one calmodulin binding site, which might indicate that they could be targeted by calmodulin. Syntenic analysis indicated that eight ZjIQDs showed syntenic relationships with those in Arabidopsis and four ZjIQDs showed syntenic relationships with those in rice. The expression profiles of ZjIQDs in response to cold, salt and alkaline stresses showed that most ZjIQDs were up- or downregulated, but ZjIQD15 and 16 were significantly upregulated under these stresses. In addition, ZjIQD15 was significantly regulated by oryzalin treatment (a microtubule disruption drug). Moreover, several ZjIQDs were differentially expressed between diploid and autotetraploid ‘Dongzao’ in response to cold stress, which might have important functions in the regulation of the differential cold resistance between diploid and autotetraploid ‘Dongzao’. This systematic analysis of ZjIQDs could provide important information for further functional characterization of key ZjIQDs in response to cold stress with the aim of breeding cold-resistant cultivars, especially improving the cold tolerance of autotetraploid ‘Dongzao’.

Genotyping of Listeria monocytogenes strains in Italian food-processing environments

Abstract Background In 2019, case fatality from Listeria monocytogenes (Lm) was higher than the previous years confirming Lm as a major public health problem. Food is the main source of Lm infection and cross-contamination from the food-processing environment (FPEs) is the most common route of contamination. The aim of this work was to characterize FPEs Lm strains collected from the National Reference Laboratory (NRL) for Lm in order to evaluate circulating clonal complexes (CCs) and their environmental adaptation. Methods NRL Lm database actually consists of 798 FPEs sequenced strains. In silico MLST was evaluated. Sequences related to prevalent CCs were further investigated using Pasteur's cgMLST scheme and detecting metal and detergent resistance genes and stress survival islet (SSI). Results MLST identified 25 CCs and 4 singleton. Nine isolates gave inexact match with existing alleles and were not classified. CC9 (17.4%) and CC121 (14.2%) were the prevalent clones. CC9 cgMLST analysis revealed a relatedness among strains isolated from the same establishment type (meat or dairy), even if sampled in different regions. Indeed, CC121 cgMLST clustering analysis was strictly associated to the point of sampling. Regarding stress adaptation genes, SSI1 and brcABC genes were detected in 100% and 2.9% of the CC9 strains, respectively. Meanwhile, SSI2 and transposon Tn6188 were detected, respectively, in 100% and 97.3% of CC121 strains. Conclusions Results confirmed, in Italian FPEs, prevalence of hypovirulent Lm strains, as previously observed in Italian food. Tested isolates highlighted adaptation to low pH and high salt concentration (SSI1 genes for CC9), to alkaline and oxidative stress (SSI2 genes for CC121) and benzalkonium chloride tolerance (Tn6188 in CC121) resulting in a best survival under stress conditions. Interestingly, CC9 clustering denoted allelic proximity among strains isolated in different Italian regions paying attention to clones spreading at national level. Key messages Listeria monocytogenes CC9 and CC121 prevalence in Italian establishment confirms environment is the main route of food contamination at production level. Prevalent food-processing environmental Italian Listeria monocytogenes strains denotes adaptation to environmental stress and clones spreading at national level.

Facilitation at early growth stages results in spatial associations and stable coexistence in late growth stages of two long‐lived, dominant shrubs

While models of species coexistence largely focus on how competition defines biological communities, over recent decades, a number of studies show positive plant–plant species interactions (facilitation) can also promote stable coexistence. The long‐lived, co‐dominant shrubs California buckwheatEriogonum fasciculatumand California sagebrushArtemisia californicashare a well‐documented positive association at the habitat level in their native California coastal sage scrub ecosystem, but mechanisms underlying their interactions remain unclear at finer spatial scales. Here, a hypothesis thatE. fasciculatumacidifies CSS's alkaline soils and facilitatesA. californicathrough amelioration of alkalinity stress is tested in a greenhouse experiment and association tests in the field. Greenhouse results demonstrate facilitation at early growth stages. In late growth stages, water competition is known to determine the shrubs' interactions with each other, but here, field observations of the shrubs in late growth stages show positive associations betweenA. californicaandE. fasciculatumthat have a positive linear relationship to increasing soil pH. These results highlight the importance of understanding lifecycle‐long interactions among species in evaluating facilitation's impacts on community structure.

Comparative Transcriptomic and Proteomic Analysis of Exopalaemon carinicauda in Response to Alkalinity Stress

Saline-alkaline waters are stressful environments where most aquatic animals can’t survive normally, and alkalinity is one of the key limited environmental factors. Due to strong adaptability to environment, the ridgetail white prawn Exopalaemon carinicauda is a potential good species suitable for large-scale culture in saline-alkaline waters. Exploring its alkaline adaptability mechanism will help to guide more marine crustaceans to saline-alkaline culture. In this study, an integrative analysis of the gill-specific transcriptome and proteome at 0, 12, and 36 h after alkalinity stress was performed to identify important regulators and pathways involved in alkalinity adaption of E. carinicauda. A total of 3,157 differentially expressed genes (DEGs) and 443 differentially expressed proteins (DEPs) were identified at 12 and 36 h compared with 0 h. Base on the transcriptome analysis, the Gene Ontology (GO) enriched terms were mainly related to ion transport, including “calcium-transporting ATPase activity,” “ATPase coupled ion transmembrane transporter activity,” “divalent inorganic cation transmembrane transporter activity,” etc., and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways mainly refer to the processes of endocrine system at both 12, and 36 h. Based on the proteomic analysis, KEGG pathways related to lipolysis and amino acids metabolism were significantly enriched at 12 h, and carbohydrate metabolism and immune response were significantly enriched at 36 h. There were significantly up-regulated expressions of ion transport related genes including aquaporin, carbonic anhydrase, ammonium transporter Rh type A-like, Na+/H+-exchanger, etc., as well as ion transport proteins including V-type proton ATPase 116 kDa subunit a-like isoform X1, sodium-potassium ATPase beta, vesicle associated membrane protein, etc. after alkalinity exposure, which indicating their important roles in response to alkalinity stress. The results of integrated analysis between proteome and transcriptome showed that up-regulated DEG/DEP (aldehyde dehydrogenase) was significantly enriched at 12 h and the up-regulated DEG/DEP (peptidylglycine alpha) was significantly enriched at 36 h, suggesting the two molecules may be critical in response to alkalinity change. This study reveals the first time-course, gill-specific, combined transcriptomic and proteomic profiling associated with alkalinity adaption of E. carinicauda and provides new insights into the mechanisms underlying the molecular response to alkalinity stress in shrimp.

Genomic Features and Molecular Function of a Novel Stress-Tolerant Bacillus halotolerans Strain Isolated from an Extreme Environment.

Simple SummaryThe Qinghai–Tibet Plateau is known as the “third pole of the world”. Due to the extreme geographical location, Qinghai–Tibet Plateau has unique ecosystems characterized by oxygen deficiency, low temperature, high salinity and alkalinity. We carried out the current study to explore the excellent extremophilic Bacillus strains via potential stress resistance as well as biocontrol properties in the Qinghai–Tibet Plateau. We found a Bacillus halotolerans strain with a promising ability to withstand harsh environments and which also exhibits an optimistic biocontrol activity against plant pathogens. We revealed the whole genome sequencing and its taxonomic position and elucidated its molecular functions that were responsible for enhancing stress tolerance as well as suppressing plant pathogens at the genetic level. Lastly, we identified this strain harbored the specific genes associated with stresses resistance, biocontrol function, and can be used as a biological agent in the agriculture field.Due to its topographical position and climatic conditions, the Qinghai–Tibet Plateau possesses abundant microorganism resources. The extremophilic strain KKD1 isolated from Hoh Xil possesses strong stress tolerance, enabling it to propagate under high salinity (13%) and alkalinity (pH 10.0) conditions. In addition, KKD1 exhibits promising biocontrol activity against plant pathogens. To further explore these traits at the genomic level, we performed whole-genome sequencing and analysis. The taxonomic identification according to the average nucleotide identity based on BLAST revealed that KKD1 belongs to Bacillus halotolerans. Genetic screening of KKD1 revealed that its stress resistance mechanism depends on osmotic equilibrium, membrane transportation, and the regulation of ion balance under salt and alkaline stress. The expression of genes involved in these pathways was analyzed using real-time quantitative PCR. The presence of different gene clusters encoding antimicrobial secondary metabolites indicated the various pathways by which KKD1 suppresses phytopathogenic growth. Moreover, the lipopeptides surfactin and fengycin were identified as being significant antifungal components of KKD1. Through comparative genomics analysis, we noticed that KKD1 harbored specific genes involved in stress resistance and biocontrol, thus providing a new perspective on the genomic features of the extremophilic Bacillus species.

Membrane Depolarization and Apoptosis-Like Cell Death in an Alkaline Environment in the Rice Pathogen Burkholderia glumae.

The rice pathogen Burkholderia glumae uses amino acids as a principal carbon source and thus produces ammonia in amino acid-rich culture medium such as Luria–Bertani (LB) broth. To counteract ammonia-mediated environmental alkaline toxicity, the bacterium produces a public good, oxalate, in a quorum sensing (QS)-dependent manner. QS mutants of B. glumae experience alkaline toxicity and may undergo cell death at the stationary phase when grown in LB medium. Here, we show that the cell-death processes of QS mutants due to alkaline environmental conditions are similar to the apoptosis-like cell death reported in other bacteria. Staining QS mutants with bis-(1,3-dibutylbarbituric acid)-trimethine oxonol revealed membrane depolarization. CellROX™ staining showed excessive generation of reactive oxygen species (ROS) in QS mutants. The expression of genes encoding HNH endonuclease (BGLU_1G15690), oligoribonuclease (BGLU_1G09120), ribonuclease E (BGLU_1G09400), and Hu-beta (BGLU_1G13530) was significantly elevated in QS mutants compared to that in wild-type BGR1, consistent with the degradation of cellular materials as observed under transmission electron microscopy (TEM). A homeostatic neutral pH was not attainable by QS mutants grown in LB broth or by wild-type BGR1 grown in an artificially amended alkaline environment. At an artificially adjusted alkaline pH, wild-type BGR1 underwent apoptosis-like cell death similar to that observed in QS mutants. These results show that environmental alkaline stress interferes with homeostatic neutral cellular pH, induces membrane depolarization, and causes apoptosis-like cell death in B. glumae.

Ameliorative Role of Silicon on Osmoprotectants, Antioxidant Enzymes and Growth of Maize Grown Under Alkaline Stress

PurposeMaize is a cereal crop which plays an important role in ensuring food security and social and economic stability. It is grown in different environments. Global agriculture is currently encountered with large scale alkalinity stress. Supplementing silicon, is an alternative way for overcoming the negative effects of sodicity on the plant growth and yield and is cheaper than other methods to reduce sodicity. Hence, a pot experiment was contemplated to examine the role of silicon in alleviating alkaline stress encountered by maize crop.MethodsAmeliorative role of silicon in alkaline stress soil was studied with four levels of alkaline stress (0, 25, 50, 75 mM) created through addition of sodium carbonate and three levels of silicon (0, 100 and 150 kg ha−1) applied to root using sodium meta silicate imposed on CO 8 maize as test crop. The experiment was conducted in factorial CRD with three replications.ResultsMaize crop experienced negative effect when grown in alkaline stressed soil. The growth (5 to 16 %), dry weight (28 to 59 %) and relative water content (5 to 23 %) reduced at various level of alkaline concentration. However, electrolyte leakage (6 to 49 %), proline (26 to 62 %), phenol (8 to 44 %), protein (6 to 19 %), anti-oxidant systems viz., peroxidase (30 to 52 %), SOD (4 to 16 %) and catalase activities (32 to 127 %) scaled up with levels of alkaline stress. Supplementing silicon in this soil ensured maize crop get a congenial environment to grow well and it was revealed by improved growth (5 to 10 %) and dry weight (17 to 30 %) of maize, relative water content (6 to 12 %) and antioxidant enzymes (25 to 52 %), water soluble protein (7 to10 %), phenol (10 to 18 %), while reduced electrolyte leakage (15 to 25 %) and proline (17 to 29 %).ConclusionsThe study divulged the primary role played by silicon in overhauling the hostile alkaline environment by promoting better growth and dry weight of maize through its regulatory action on osmoprotectants and anti-oxidant enzymes and maximum effect was evident with 150 kg Si/ha.