Oxidase Gene Research Articles

Characterization and Identification of Drought-Responsive ABA-Aldehyde Oxidase (AAO) Genes in Potato (Solanum tuberosum L.).

Abscisic acid (ABA) is an important stress hormone that affects plants' tolerance to stress. Changes in the content of abscisic can have an impact on plant responses to abiotic stress. The abscisic acid aldehyde oxidase (AAO) plays a crucial role in the final step in the synthesis of abscisic acid; therefore, understanding the function of the AAO gene family is of great significance for insight into plants' response to abiotic stresses. In this study, Solanum tuberosum AAO (StAAO) members were exhaustively explored using genome databases, and nine StAAOs were identified. Chromosomal location analysis indicated that StAAO genes mapped to 4 of the 14 potato chromosomes. Further analyses of gene structure and motif composition showed that members of the specific StAAO subfamily showed relatively conserved characteristics. Phylogenetic relationship analysis indicated that StAAOs proteins were divided into three major clades. Promoter analysis showed that most StAAO promoters contained cis-elements related to abiotic stress response and plant hormones. The results of tissue-specific expression analysis indicated that StAAO4 was predominantly expressed in the roots. Analysis of transcriptome data revealed that StAAO2/4/6 genes responded significantly to drought treatments. Moreover, further qRT-PCR analysis results indicated that StAAO2/4/6 not only significantly responded to drought stress but also to various phytohormone (ABA, SA, and MeJA) and abiotic stresses (salt and low temperature), albeit with different expression patterns. In summary, our study provides comprehensive insights into the sequence characteristics, structural properties, evolutionary relationships, and expression patterns of the StAAO gene family. These findings lay the foundation for a deeper understanding of the StAAO gene family and offer a potential genetic resource for breeding drought-resistant potato varieties.

Manure trial evaluation of Bacillus velezensis PHP1601 as a viable biocontrol candidate against nuisance fly species at an operational piggery

Fly infestations threaten farming and livestock operations, contributing to animal irritation and the spread of serious diseases. In this study, the application of Bacillus velezensis PHP1601 (Bacillales: Bacillaceae) treatments (105 and 1010 endospore g−1) to pig manure slurries was evaluated as a means of controlling fly reproductive cycles. Two cycles of fly emergence were evaluated over a 33 days period for each replicated trial. For both treatments, the first emergence event resulted in a significant reduction in the percentage of flies emerging compared to an un-treated control. Extended monitoring of the biocontrol containers revealed that fly emergence was completely eliminated by the time that a second round of fly emergence was observed for the control. A B. velezensis-specific real-time PCR method was developed and used to assess the population dynamics of the applied biocontrol agent over the course of the experiment. Strain PHP1601 remained viable in the manure and cell numbers increased by several orders of magnitude. REP-PCR fingerprinting was used to confirm the clonality of endospores recovered from the manure. Fly species recovered from the trials were identified by cytochrome oxidase gene barcode sequencing. Several species of veterinary and medically significant flies were identified. They were all deemed to be susceptible to treatments with PHP1601 and constituted part of the strain's host range. The study demonstrated the effectiveness of B. velezensis PHP1601 as a promising biocontrol agent for controlling fly infestations under conditions similar to its intended use.

Identification of key overlapping DEGs and molecular pathways under multiple stressors in the liver of Nile tilapia (Oreochromis niloticus)

The identification of key genes and molecular pathways that are involved in the response to stressors is crucial for controlling stress in fish and sustainable aquaculture. Environmental stressors can induce stress responses in aquatic animals, resulting in compromised immune function, inhibited growth, and increased mortality rates. mRNA-seq analysis provides a powerful tool to identify key genes and pathways associated with stress response. In the present study, mRNA-seq analysis was employed to identify key overlapping differentially expressed genes (DEGs) and molecular pathways under salinity, nitrite, copper, and pH stress in the liver of Nile tilapia (Oreochromis niloticus). The pathways associated with the immune response, oxygen transport, homeostasis, and oxidative stress were enriched across all stressors. The top KEGG pathways were complement and coagulation cascades, PPAR signaling pathway, and cardiac muscle contraction. The top GO enrichment terms were oxidoreductase activity, aerobic respiration, endopeptidase inhibitor activity, endopeptidase regulator activity, heme binding, and iron ion binding. The complement genes (C3, C4, C5, factor B, and factor H), alpha-2-macroglobulin (A2M), hemoglobin subunit epsilon (HBE), hemoglobin subunit alpha (HBA), coagulation factor genes (XI and X) and the cytochrome c oxidase (COX) gene family (cox1, cox2, cox3, cytochrome P450) were identified as key shared genes across multiple stressors. The discovery of these genes and molecular pathways provided a better understanding of the molecular mechanism underlying the stress response in Nile tilapia. The results of the present study can facilitate the development of stress management strategies in Nile tilapia.

Photomodulatory effects in the hypothalamus of sleep-deprived young and aged rats

Insufficient sleep is associated with impaired hypothalamic activity and declined attentional performance. In this study, alterations in the hypothalamus of REM sleep-deprived (SD) young and aged rats, and the modulatory effect of near-infrared (NIR) laser were investigated. Forty-eight male Wistar rats (24 young at 2 months and 24 senile at 14 months) were divided into three groups: the control, the SD group subjected to 72 hr of sleep deprivation, and the transcranial-NIR laser-treated (TLT) group subjected to SD for 72 hr and irradiated with 830 nm laser. The hypothalamic levels of oxidative stress, inflammatory biomarkers, antioxidant enzymes, mitochondrial cytochrome C oxidase (CCO), apoptotic markers (BAX, BCL-2), and neuronal survival-associated genes (BDNF, GLP-1) were evaluated. Furthermore, the hypothalamic tissue alterations were analyzed via histological examination. The results revealed that TLT treatment has enhanced the antioxidant status, prevented oxidative insults, suppressed neuroinflammation, regulated CCO activity, reduced apoptotic markers, and tuned the survival genes (BDNF & GLP-1) in hypothalamic tissue of SD young and aged rats. Microscopically, TLT treatment has ameliorated the SD-induced alterations and restored the normal histological features of hypothalamus tissue. Moreover, the obtained data showed that SD and NIR laser therapy are age-dependent. Altogether, our findings emphasize the age-dependent adverse effects of SD on the hypothalamus and suggest the use of low-laser NIR radiation as a potential non-invasive and therapeutic approach against SD-induced adverse effects in young and aged animals.

Gallionellaceae pangenomic analysis reveals insight into phylogeny, metabolic flexibility, and iron oxidation mechanisms.

Neutrophilic iron-oxidizing bacteria (FeOB) produce copious iron (oxyhydr)oxides that can profoundly influence biogeochemical cycles, notably the fate of carbon and many metals. To fully understand environmental microbial iron oxidation, we need a thorough accounting of iron oxidation mechanisms. In this study, we show the Gallionellaceae FeOB genomes encode both characterized iron oxidases as well as uncharacterized multiheme cytochromes (MHCs). MHCs are predicted to transfer electrons from extracellular substrates and likely confer metabolic capabilities that help Gallionellaceae occupy a range of different iron- and mineral-rich niches. Gallionellaceae appear to specialize in iron oxidation, so it would be advantageous for them to have multiple mechanisms to oxidize various forms of iron, given the many iron minerals on Earth, as well as the physiological and kinetic challenges faced by FeOB. The multiple iron/mineral oxidation mechanisms may help drive the widespread ecological success of Gallionellaceae.

Roles of ROS metabolism and phenylpropanoid pathway in quality maintenance of postharvest Pleurotus eryngii under hyperoxia stress

Hyperoxia application has been proven to be effective in mushrooms preservation, however, the possible induction mechanisms of hyperoxia stress on P. eryngii quality maintenance are incompletely understood. In this study, Pleurotus eryngii treated with hyperoxia (80% O2 +20% N2) and the induced resistance responses of fruiting bodies were mainly investigated through ROS metabolism and phenylpropanoid pathway. The application of hyperoxia could significantly alleviate the quality deterioration of P. eryngii during storage, as observed by higher maintenance of firmness, nutrient substance and membrane integrity. Moreover, hyperoxia application induced the production of hydrogen peroxide (H2O2) in the early stage, as well as the enzymatic antioxidant defense system. The NADPH oxidase genes (PeNoxR) and MAPK signaling pathway were also up-regulated during hyperoxia stress. Besides, hyperoxia stress also promoted the accumulation of major secondary metabolites and up-regulated the transcription levels of the key genes involved in phenylpropanoid pathway. Taken together, this study provided substantial evidences that hyperoxia could induce stress resistance in P. eryngii due to its positive regulation of ROS metabolism-modifying enzyme activities and genes, as well as possible activated phenylpropanoid pathway, therefore delaying the quality deterioration of P. eryngii during postharvest period.

Usefulness of current sgRNA design guidelines and in vitro cleavage assays for plant CRISPR/Cas genome editing: a case targeting the polyphenol oxidase gene family in eggplant (Solanum melongena L.).

The advent of genome editing platforms such as the CRISPR/Cas9 system ushers an unprecedented speed in the development of new crop varieties that can withstand the agricultural challenges of the 21st century. The CRISPR/Cas9 system depends on the specificity of engineered single guide RNAs (sgRNAs). However, sgRNA design in plants can be challenging due to the multitude of design tools to choose from, many of which use guidelines that are based on animal experiments yet allow the use of plant genomes. Upon choosing sgRNAs, it is also unclear whether an in vitro assay is needed to validate the targeting efficiency of a particular sgRNA before in vivo delivery of the CRISPR/Cas9 system. Here, we demonstrate the in vitro and in vivo activity of four different sgRNAs that we selected based on their ability to target multiple members of the eggplant polyphenol oxidase gene family. Some sgRNAs that have high in vitro cleavage activity did not produce edits in vivo, suggesting that an in vitro assay may not be a reliable basis to predict sgRNAs with highly efficient in vivo cleavage activity. Further analysis of our sgRNAs using other design algorithms suggest that plant-validated criteria such as the presence of necessary secondary structures and appropriate base-pairing may be the reason for the discrepancy between our observed in vitro and in vivo cleavage efficiencies. However, recent reports and our data suggests that there is no guaranteed way to ensure the in vivo cleavage of chosen sgRNAs.

Argas persicus and Carios vespertilionis Ticks Infesting Ducks, Domestic Fowls and Bats in Pakistan: First Report on Molecular Survey and Phylogenetic Position of Borrelia anserina.

Argasid ticks have the vectorial potential for transmitting disease-causing pathogens to avian hosts, resulting in economic losses that may not be fully estimated. Borrelia species are the responsible agents of borreliosis in poultry, animals and humans. Our previous studies have reported a high prevalence of Argas persicus infesting domestic fowls in Khyber Pakhtunkhwa (KP), Pakistan. However, molecular screening and genetic characterization of Borrelia spp. in A. persicus have been neglected in Pakistan. In this study, we focused on the molecular epidemiology and genetic characterization of Borrelia spp. associated with A. persicus ticks infesting domestic fowls and ducks, and Carios vespertilionis infesting bats in selected districts of KP. Overall, 1818 ticks, including females (415; 23%), males (345; 19%), nymphs (475; 26%) and larvae (583; 32%), were collected from 27 locations in nine districts (Peshawar, Mardan, Swabi, Charsadda, Chitral, Lakki Marwat, Bannu, Bajaur and Hangu) from domestic fowls, ducks and their shelters, and bats. A subset of 197 ticks was selected for DNA extraction and PCR to amplify fragments of the cytochrome c oxidase (cox) gene for ticks and flagellin B (flaB) for the detection and genetic characterization of associated Borrelia spp. Among these, only Borrelia anserina DNA was detected in 40 ticks (27.2%) of different life stages, where highest prevalence was found in female ticks (18; 45%), followed by nymphs (12; 30%), larvae (7; 17.5%) and males (3; 7.5%). Tick infestation in shelters (1081; 77%) was higher than on hosts (323; 23%). The resultant cox amplicons of A. persicus showed 100% identity with the same species reported from Pakistan, China, Iran, Kenya, Kazakhstan, Algeria and Egypt and C. vespertilionis show 100% identity with the species reported from Pakistan, China, Japan, Kenya, Vietnam, Spain, Netherlands, the United Kingdom and Hungry, and clustered with the aforementioned species in the phylogenetic tree. The obtained Borrelia sequences showed 100% identity with B. anserina and revealed a close resemblance to the relapsing fever group and clustered in a monophyletic clade with B. anserina from India, Iran and Brazil in a phylogenetic tree. These results establish the first molecular characterization of B. anserina in A. persicus infesting domestic fowls and ducks in the region, as well as their shelters. To effectively control zoonotic consequences, country-wide surveillance research should be encouraged to screen soft ticks infesting various birds for associated pathogens.

Association of LOX gene G473A polymorphism with the occurrence of allergic rhinitis and efficacy of montelukast sodium in children

ABSTRACT Allergic rhinitis (AR) is very common in adolescents, and current treatment options are complex and unsatisfactory. The objective of this study was to analyze the association of lysyl oxidase (LOX) gene G473A polymorphism with susceptibility to AR in children. In addition, we analyzed the therapeutic effect of montelukast sodium on AR. Forty-five children with AR (research group, 8.16±2.88 years old) and 51 healthy children (control group, 8.22±3.87 years old) during the same period were selected. The LOX gene G473A polymorphism was detected with polymerase chain reaction (PCR)-restriction fragment length polymorphism method. The effect of G473A polymorphism in the occurrence of AR was assessed by logistic regression analysis. In addition, the levels of C-reactive protein (CRP), Interleukin (IL-6), and IL-8 were measured to observe the relationship between G473A polymorphism and inflammatory factors. Finally, montelukast sodium was given to children with AR to investigate the effect of G473A polymorphism on clinical outcomes. The number of G473A polymorphisms in the research group was not significantly different from the control group for GA-type (P = 0.521). However, the number of GG-type polymorphisms was less while the number of type AA was more than the control group (P = 0.044 and 0.046). Children carrying the AA gene had an approximately 4-fold increased risk of AR, while those carrying the GG gene had a decreased risk (P < 0.001). Moreover, children carrying the GG gene had lower levels of CRP, IL-6, and IL-8 and better clinical outcomes, while those carrying the AA gene had higher levels of inflammatory factors and worse outcomes (P<0.05). LOX gene G473A polymorphism is closely associated with AR pathogenesis and may have an important research value in antagonizing the therapeutic effect of montelukast sodium.

Unveiling the molecular identity of the diminutive cyprinid, Horadandia brittani (Teleostei: Cyprinidae), a species endemic to Southern India.

Horadandia brittani is a small cyprinid fish species initially discovered in the coastal floodplains of southern India. For almost 50 years, the genus Horadandia was monotypic with a single species confined to Sri Lanka. In 1992, a new species H. brittani was described from south-western India. Despite being described as a separate species, H. brittani was later considered a synonym of H. atukorali, but in 2013, researchers recognized it as a distinct species based on morphological differences. Despite this clarification, there was still a need to validate the identity of H. brittani and determine its evolutionary relationship with its closely related species using DNA sequences. To address the uncertainties surrounding the identity of H. brittani, the present study utilized molecular techniques to generate DNA sequences. Sample collection involved obtaining specimens of H. brittani from their natural habitats. Subsequently, DNA was extracted from the collected samples, and the mitochondrial cytochrome C oxidase (COI) gene was amplified using appropriate methods. The analysis of DNA sequences obtained from the COI gene revealed significant genetic distinctions between H. brittani and H. atukorali. The genetic distance values between these two species ranged from 3.21 to 3.63%, clearly indicating that these two species are genetically separate entities. The study successfully established the phylogenetic relationships between H. brittani and H. atukorali based on the COI gene sequences, further confirming the validity of H. brittani as a distinct and separate species. The findings of this study conclusively demonstrate that H. brittani is a valid and separate species, distinct from H. atukorali. The genetic analysis based on mitochondrial COI gene sequences provided strong evidence for the differentiation between these two species. The molecular data generated in this research can be used to identify H. brittani quickly and accurately in the future.

Growth Responses and Adventitious Root Formation of Cucumber Hybrid Lines in a Waterlogged Condition

Cucumber (Cucumis sativus L.) F1 hybrids are grown commercially in open-field or greenhouse conditions. Hybrids are well adapted to these settings due to directed breeding. In protected cultivation systems, a small rhizosphere volume and intensive, continuous fertigation predispose the roots to waterlogging (WL) conditions and potentially to hypoxia. However, high productivity is expected and achieved under these conditions. The aim of this study was to identify traits that play a role in this surprising behavior. Initial observations revealed the presence of a significantly higher number of adventitious roots (ARs) in three greenhouse (7–14) vs. three open-field cultivars (less than two) grown under normal conditions. Further on, two contrasting representative hybrids typically grown in open-field and in greenhouse conditions were subjected to WL stress. Declining oxygen levels in the media and increased alcohol dehydrogenase activity (ADH) in the roots were experienced during the WL treatment in both hybrids, with anaerobic metabolism triggered less intensively (~4-fold less ADH activity) in the greenhouse-type ‘Oitol’. The induction characteristics of cysteine oxidase (CysOx) genes, key components of the hypoxia sensing pathway, were used to confirm the hypoxic stress experienced by the roots. The lower extent of upregulation in CysOx genes expression agreed with the milder level of hypoxic stress in the roots of ‘Oitol’ than in ‘Joker’. The more efficient induction of AR formation with a ~50% increase upon waterlogging stress was found to be a prominent trait in ‘Oitol’, apparently helping root internal aeration and mitigating hypoxia. The shoot growth of neither hybrid was set back by hypoxic root conditions. ‘Joker’ plants maintained the same growth rate as that of the control, while the growth of ‘Oitol’ accelerated when its root system was flooded with nutrient solution. Acclimation processes to hypoxia were proposed to explain the lack of growth retardation in both varieties. This corresponded well with a general abundance of AR development in greenhouse-type (slicing) cucumbers that are typically cultivated in soilless systems.

Drought Stress and High Temperature Affect the Antioxidant Metabolism of Cotton (Gossypium hirsutum L.) Anthers and Reduce Pollen Fertility

Both drought and high temperature can influence the antioxidant metabolism of crop reproductive organs in different ways, affecting the fertility of reproductive organs and yield formation. However, the combined effects of drought stress and high temperature on the crop reproductive physiology have not yet been widely considered. In order to broaden our understanding of this mechanism of influence, a pond experiment was conducted using a cotton variety Yuzaomian 9110 divided into four treatment groups: control (CK), drought stress (DS), high temperature (HT), and drought stress coupled with high temperature (DS+HT). Results showed a significant negative correlation between pollen viability and superoxide anion (O2−) content, as well as hydrogen peroxide (H2O2). Compared with CK, DS did not alter O2− content in anthers, but HT treatment resulted in higher anther O2−. Compared with single-stress groups, DS+HT further promoted the formation of O2− in anthers, leading to more malondialdehyde in anthers. Moreover, a higher H2O2 content in anthers was found in DS and HT than in CK. DS+HT did not show altered H2O2 content relative to HT treatment, although its H2O2 was higher than in DS. Further analyses of the antioxidant enzyme system showed that DS had no significant effect on superoxide dismutase gene (GhCu/ZnSOD) expression, but HT and DS+HT significantly downregulated its expression. The expression of GhCu/ZnSOD was lower under DS+HT than HT, which might be why O2− content was not altered under DS treatment compared with CK and was higher in DS+HT than HT. DS and HT significantly downregulated the expression of the peroxidase gene (GhPOD) and catalase gene (GhCAT), which should be the main reason for the larger accumulation of H2O2 under drought stress and high-temperature conditions. Compared with single-stress groups, DS+HT had lower expression of GhCAT, resulting in a larger H2O2 content. Regarding the ascorbic acid–glutathione (AsA–GSH) cycle, DS and HT significantly downregulated the expression of monodehydroascorbate reductase gene (GhMDHAR) to hinder the production of AsA and upregulated the expression of ascorbate oxidase gene (GhAAO) to promote the oxidation of AsA, which was theoretically detrimental to AsA accumulation. However, HT downregulated the expression of the ascorbate peroxidase gene (GhAPX), hindering the reduction of H2O2 by AsA, which was the reason for AsA and H2O2 accumulation. Moreover, DS also significantly upregulated the expression of the dehydroascorbate reductase gene (GhDHAR2) to enhance the reduction of dehydroascorbate to form AsA, leading to a higher content of AsA under DS than HT. The combined stress significantly downregulated the expression of GhAAO to inhibit the oxidation of AsA but significantly upregulated the expression of GhMDHAR and GhDHAR2, promoting the AsA production, and downregulated the expression of GhAPX, hindering the reduction of H2O2 by AsA. All these resulted in increased AsA content under combined stresses. In addition, HT significantly downregulated the glutathione reductase gene (GhGR) expression, hindering the reduction of oxidized glutathione (GSSG), which led to the reduction of GSH. However, DS and DS+HT significantly downregulated the glutathione peroxidase gene (GhGPX) expression, resulting in the accumulation of GSH. Overall, compared with single-stress treatments, the effects of DS+HT on cotton pollen fertility and peroxide accumulation were more significant. The effects of DS+HT on the antioxidant enzyme system were mainly caused by high temperature, while the mechanism of abnormal accumulation of AsA and GSH caused by DS+HT was different from those of single-stress groups.

Impact of varying light spectral compositions on photosynthesis, morphology, chloroplast ultrastructure, and expression of light-responsive genes in Marchantia polymorpha

Marchantia polymorpha is a convenient model for studying light of different spectral compositions on various physiological and biochemical processes because its photoreceptor system is vastly simplified. The influence of red light (RL, 660 nm), far-red light (FRL, 730 nm), blue light (BL, 450 nm), and green light (GL, 525 nm) compared to white light (high-pressure sodium light (HPSL), white LEDs (WL 450 + 580 nm) and white fluorescent light (WFL) on photosynthetic and transpiration rates, photosystem II (PSII) activity, photomorphogenesis, and the expression of light and hormonal signaling genes was studied. The ultrastructure of the chloroplasts in different tissues of the gametophyte M. polymorpha was examined. FRL led to the formation of agranal chloroplasts (in the epidermis and the chlorenchyma) with a high starch content (in the parenchyma), which led to a reduced intensity of photosynthesis. BL increased the transcription of genes for the biosynthesis of secondary metabolites - chalcone synthase (CHS), cellulose synthase (CELL), and L-ascorbate peroxidase (APOX3), which is consistent with the increased activity of low-molecular weight antioxidants. FRL increased the expression of phytochrome apoprotein (PHY) and cytokinin oxidase (CYTox) genes, but the expression of the phytochrome interacting factor (PIF) gene decreased, which was accompanied by a significant change in gametophyte morphology. Analysis of crosstalk gene expression, and changes in morphology and photosynthetic activity was carried out.

The golden bell flower genome provides insights into its evolutionary history and reveals the potential genomic bases of its ecological divergence from weeping forsythia

The golden bell flower (GBF), a beloved flower that blooms in early spring, faces a significant challenge in genetic research and breeding due to the lack of available genomic data. Here, a genome of GBF with a genome size of 735.31 Mb and tiny gaps of 3,782 N were assembled. Our results showed GBF underwent two round of whole genome doubling (WGD) events. The more recent WGD events, along with subsequent structural variations in GBF and weeping forsythia, might lead to the divergence of GBF and weeping forsythia. Analysis of enriched KEGG pathways and Gene Ontology categories of the expanded and contracted genes indicated an increase in the GBF abilities of faster growth and stronger cold and drought tolerance since the separation of the two species. Our results showed that the effective population size (Ne) of the GBF species has been decreasing, declining from 0.352 Mya to 0.018 Mya. However, the stronger cold tolerance of GBF likely helped to alleviate the reduction of Ne during the Pleistocene glacial periods. The expansion of peroxidase (POD) and l-gulonolactone oxidase (GULO) genes and more POD and GULO genes with anaerobic and ABA-induced cis elements in GBF genome might be responsible for the strong waterlogging tolerance of GBF. Our study characterized the evolutionary history of GBF and the genomic bases for the ecological divergenece between GBF and weeping forsythia. Our genome provides genetic resources for future genetic and breeding studies on GBF and its related species.

Elucidating the effects on polyphenol oxidase activity and allelic variation of polyphenol oxidase genes on dough and whole wheat-derived product color parameters

ABSTRACT Polyphenol oxidase (PPO) activity is a primary cause of the development of unattractive dark brown discoloration of wheat-based end products. The present study aims to evaluate a set of 41 diverse wheat genotypes grown at three different locations in India for grain phenol color reaction, PPO activity and molecular marker-based characterization of alleles of PPO genes. Relationships among these parameters were analyzed along with the effects of grain PPO activity on dough and chapati color at different time intervals. The mean PPO activity ranged from 7.42 to 27.57 min−1 g−1 10−3 among the genotypes and it showed a significant negative correlation with color brightness (L*) of dough rested for 0 min (r = -0.406), 15 min (r = -0.406), 2 h (r = -0.502) and 4 h (r = -0.551) and whole wheat flour-derived chapati rested for 2 h (r = -0.267) and 4 h (r = -0.424). The overall quality color score was negatively correlated with PPO activity (r = -0.863) and showed a positive correlation with both dough and chapati visual color measured at different time intervals. PPO activity in the genotypes carrying different alleles was found to be Ppo-A1a>Ppo-A1b; Ppo-B2d>Ppo-B2a; and Ppo-D1b>Ppo-D1a. The allelic constitution Ppo-A1bPpo-B2aPpo-D1a and Ppo-A1bPpo-B2dPpo-D1 was found to produce the lowest PPO activity, and thus these alleles are recommended to be used in marker assisted breeding for low PPO activity genotypes to minimize the discoloration of wheat-based end-products.

Improving the diagnosis of Streptococcus iniae using a novel probe-based qPCR assay combined with an enrichment step.

Streptococcus iniae is a bacterial pathogen that causes streptococcosis, leading to significant losses in fish aquaculture globally. This study reported a newly developed probe-based quantitative polymerase chain reaction (qPCR) method for the detection of S. iniae. The primers and probes were designed to target the lactate oxidase gene. The optimized method demonstrated a detection limit of 20 copies per reaction and was specific to S. iniae, as evidenced by no cross-reactivity when assayed against genetic materials extracted from 23 known aquatic animal pathogens, and fish samples infected with Streptococcus agalactiae or Streptococcus dysgalactiae. To validate the newly developed qPCR protocol with field samples, fish specimens were systematically investigated following the Food and Agriculture Organization of the United Nations & Network of Aquaculture Centres in Asia-Pacific three diagnostic levels approach, which integrated basic and advanced techniques for disease diagnosis, including observation of gross signs (level I), bacterial isolation (level II), qPCR and 16S rDNA sequencing (level III). The result showed that 7/7 affected farms (three Asian seabass farms and four tilapia farms) experiencing clinical signs of streptococcosis were diagnosed positive for S. iniae. qPCR assays using DNA extracted directly from fish tissue detected S. iniae in 11 out of 36 fish samples (30.6%), while 24 out of 36 samples (66.7%) tested positive after an enrichment step, including apparently healthy fish from affected farms. Bacterial isolation of S. iniae was only successful in a proportion of clinically diseased fish but not in healthy-looking fish from the same farm. Overall, the newly developed qPCR protocol combined with enrichment would be a useful tool for the diagnosis and surveillance of S. iniae infections in fish populations, thereby aiding in the disease control and prevention.

Heterologous codA Gene Expression Leads to Mitigation of Salt Stress Effects and Modulates Developmental Processes.

Transgenic tobacco plants overexpressing the choline oxidase gene from A. globiformis showed an increase in resistance at the level of primary and secondary biosynthesis of metabolites, removing the damage characteristic of salinity and stabilizing the condition of plants. We used 200 mM NaCl, which inhibits the growth of tobacco plants at all stages of development. Leaves of transgenic and wild-type (WT) plants Nicotiána tabácum were used for biochemical, cytological and molecular biological analysis. However, for transgenic lines cultivated under normal conditions (without salinity), we noted juvenile characteristics, delay in flowering, and slowing down of development, including the photosynthetic apparatus. This caused changes in the amount of chlorophyll, a delay in the plastid grana development with the preservation of prolamellar bodies. It also caused changes in the amount of sugars and indirectly downstream processes. A significant change in the activity of antioxidant enzymes and a change in metabolism is probably compensated by the regulation of a number of genes, the expression level of which was also changed. Thus, the tolerance of transgenic tobacco plants to salinity, which manifested itself as a result of the constitutive expression of codA, demonstrates an advantage over WT plants, but in the absence of salinity, transgenic plants did not have such advantages due to juvenilization.

Wheat gibberellin oxidase genes and their functions in regulating tillering.

Multiple genetic factors control tillering, a key agronomy trait for wheat (Triticum aestivum L.) yield. Previously, we reported a dwarf-monoculm mutant (dmc) derived from wheat cultivar Guomai 301, and found that the contents of gibberellic acid 3 (GA3) in the tiller primordia of dmc were significantly higher. Transcriptome analysis indicated that some wheat gibberellin oxidase (TaGAox) genes TaGA20ox-A2, TaGA20ox-B2, TaGA3ox-A2, TaGA20ox-A4, TaGA2ox-A10 and TaGA2ox-B10 were differentially expressed in dmc. Therefore, this study systematically analyzed the roles of gibberellin oxidase genes during wheat tillering. A total of 63 TaGAox genes were identified by whole genome analysis. The TaGAoxs were clustered to four subfamilies, GA20oxs, GA2oxs, GA3oxs and GA7oxs, including seven subgroups based on their protein structures. The promoter regions of TaGAox genes contain a large number of cis-acting elements closely related to hormone, plant growth and development, light, and abiotic stress responses. Segmental duplication events played a major role in TaGAoxs expansion. Compared to Arabidopsis, the gene collinearity degrees of the GAoxs were significantly higher among wheat, rice and maize. TaGAox genes showed tissue-specific expression patterns. The expressions of TaGAox genes (TaGA20ox-B2, TaGA7ox-A1, TaGA2ox10 and TaGA3ox-A2) were significantly affected by exogenous GA3 applications, which also significantly promoted tillering of Guomai 301, but didn't promote dmc. TaGA7ox-A1 overexpression transgenic wheat lines were obtained by Agrobacterium mediated transformation. Genomic PCR and first-generation sequencing demonstrated that the gene was integrated into the wheat genome. Association analysis of TaGA7ox-A1 expression level and tiller number per plant demonstrated that the tillering capacities of some TaGA7ox-A1 transgenic lines were increased. These data demonstrated that some TaGAoxs as well as GA signaling were involved in regulating wheat tillering, but the GA signaling pathway was disturbed in dmc. This study provided valuable clues for functional characterization of GAox genes in wheat.

Protein Kinase A Controls the Melanization of Candida auris through the Alteration of Cell Wall Components.

Candida auris, a multidrug-resistant fungal pathogen, significantly threatens global public health. Recent studies have identified melanin production, a key virulence factor in many pathogenic fungi that protects against external threats like reactive oxygen species, in C. auris. However, the melanin regulation mechanism remains elusive. This study explores the role of the Ras/cAMP/PKA signaling pathway in C. auris melanization. It reveals that the catalytic subunits Tpk1 and Tpk2 of protein kinase A (PKA) are essential, whereas Ras1, Gpr1, Gpa2, and Cyr1 are not. Under melanin-promoting conditions, the tpk1Δ tpk2Δ strain formed melanin granules in the supernatant akin to the wild-type strain but failed to adhere them properly to the cell wall. This discrepancy is likely due to a decreased expression of chitin-synthesis-related genes. Our findings also show that Tpk1 primarily drives melanization, with Tpk2 having a lesser impact. To corroborate this, we found that C. auris must deploy Tpk1-dependent melanin deposition as a defensive mechanism against antioxidant exposure. Moreover, we confirmed that deletion mutants of multicopper oxidase and ferroxidase genes, previously assumed to influence C. auris melanization, do not directly contribute to the process. Overall, this study sheds light on the role of PKA in C. auris melanization and enhances our understanding of the pathogenicity mechanisms of this emerging fungal pathogen.

Metagenomic and culture-dependent approaches unveil active microbial community and novel functional genes involved in arsenic mobilization and detoxification in groundwater

BackgroundArsenic (As) and its species are major pollutants in ecological bodied including groundwater in Bangladesh rendering serious public health concern. Bacteria with arsenotrophic genes have been found in the aquifer, converting toxic arsenite [As (III)] to less toxic arsenate [As (V)] that is easily removed using chemical and biological trappers. In this study, genomic and metagenomic approaches parallel to culture-based assay (Graphical abstract) have made it possible to decipher phylogenetic diversity of groundwater arsenotrophic microbiomes along with elucidation of their genetic determinants.ResultsSeventy-two isolates were retrieved from six As-contaminated (average As concentration of 0.23 mg/L) groundwater samples from Munshiganj and Chandpur districts of Bangladesh. Twenty-three isolates harbored arsenite efflux pump (arsB) gene with high abundance, and ten isolates possessing arsenite oxidase (aioA) gene, with a wide range of minimum inhibitory concentration, MICAs (2 to 32 mM), confirming their role in arsenite metabolism. There was considerable heterogeneity in species richness and microbial community structure. Microbial taxa from Proteobacteria, Firmicutes and Acidobacteria dominated these diversities. Through these combinatorial approaches, we have identified potential candidates such as, Pseudomonas, Acinetobacter, Stenotrophomonas, Achromobacter, Paraburkholderia, Comamonas and Klebsiella and associated functional genes (arsB, acr3, arsD, arsH, arsR) that could significantly contribute to arsenite detoxification, accumulation, and immobilization.ConclusionsCulture-dependent and -independent shotgun metagenomic investigation elucidated arsenotrophic microbiomes and their functions in As biogeochemical transformation. These findings laid a foundation for further large-scale researches on the arsenotrophic microbiomes and their concurrent functions in As biogeochemical transformation in As-contaminated areas of Bangladesh and beyond.