MicroRNA-mediated host defense mechanisms against pathogens and herbivores in rice: balancing gains from genetic resistance with trade-offs to productivity potential.

BACKGROUND: Rice (Oryza sativa L.) is the major source of daily caloric intake for more than 30% of the human population. However, the sustained productivity of this staple food crop is continuously threatened by various pathogens and herbivores. Breeding has been successful in utilizing various mechanisms of defense by gene pyramiding in elite cultivars, but the continuous resurgence of highly resistant races of pathogens and herbivores often overcomes the inherent capacity of host plant immunity. MicroRNAs (miRNAs) are endogenous, short, single-stranded, non-coding RNA molecules that regulate gene expression by sequence-specific cleavage of target mRNA or suppressing target mRNA translation. While miRNAs function as upstream regulators of plant growth, development, and host immunity, their direct effects on growth and development in the context of balancing defenses with agronomic potential have not been extensively discussed and explored as a more viable strategy in breeding for disease and pest resistant cultivars of rice with optimal agronomic potentials. RESULTS: Using the available knowledge in rice and other model plants, this review examines the important roles of miRNAs in regulating host responses to various fungal, bacterial, and viral pathogens, and insect pests, in the context of gains and trade-offs to crop yield. Gains from R-gene-mediated resistance deployed in modern rice cultivars are often undermined by the rapid breakdown of resistance, negative pleiotropic effects, and linkage drags with undesirable traits. In stark contrast, several classes of miRNAs are known to efficiently balance the positive gains from host immunity without significant costs in terms of losses in agronomic potentials (i.e., yield penalty) in rice. Defense-related miRNAs such as Osa-miR156, Osa-miR159, Osa-miR162, Osa-miR396, Osa-530, Osa-miR1432, Osa-miR1871, and Osa-miR1873 are critical in fine-tuning and integrating immune responses with physiological processes that are necessary to the maintenance of grain yield. Recent research has shown that many defense-related miRNAs regulate complex and agronomically important traits. CONCLUSIONS: Identification of novel immune-responsive miRNAs that orchestrate physiological processes critical to the full expression of agronomic potential will facilitate the stacking of optimal combinations of miRNA-encoding genes to develop high-yielding cultivars with durable resistance to disease and insect pests with minimal penalties to yield.

Novel cis-acting regulatory elements in wild Oryza species impart improved rice bran quality by lowering the expression of phospholipase D alpha1 enzyme (OsPLDα1).

Rice bran oil is good quality edible oil, rich in antioxidants and comprised typically of oleic-linoleic type fatty acids. However, presence of a highly lipolytic enzyme Phospholipase D alpha1 (OsPLDα1) increases free fatty acid content in the oil which further leads to stale flavor and rancidity of the oil, making it unfit for human consumption. In this study, we compared the upstream regions of OsPLDα1 orthologs across 34 accessions representing 5 wild Oryza species and 8 cultivars, to uncover sequence variations and identify cis-elements involved in differential transcription of orthologs. Alignment of the upstream sequences to the Nipponbare OsPLDα1 reference sequence revealed the presence of 39 SNPs. Phylogenetic analysis showed that all the selected cultivars and wild species accessions are closely related to the reference except for three accessions of O. rufipogon (IRGC89224, IRGC104425, and IRGC105902). Furthermore, using exon-specific qRT-PCR, OsPLDα1 expression patterns in immature grains indicated significant differences in transcript abundance between the wild species accessions. In comparison to the control, lowest gene expression was observed in IRGC89224 accession (0.20-fold) followed by IRGC105902 (0.26-fold) and IRGC104425 (0.41-fold) accessions. In-silico analysis of the OsPLDα1 promoter revealed that the copy number variations of CGCGBOXAT, GT1CONSENSUS, IBOXCORE, NODCON2GM, OSE2ROOTNODULE, SURECOREATSULTR11, and SORLIP1AT cis-elements play an important role in the transcriptional activities of orthologous genes. Owing to the presence of ARFAT and SEBF elements only in the IRGC89224 accession, which had the lowest gene expression as well, these putative upstream regulatory sequences have been identified as novel cis-elements which may act as repressors in regulating the OsPLDα1 gene expression. The accessions identified with low OsPLDα1 expressions could be further deployed as potential donors of ideal OsPLDα1 allele for transfer of the desired trait into elite rice cultivars.

Implications of miR166 and miR159 induction to the basal response mechanisms of an andigena potato (Solanum tuberosum subsp. andigena) to salinity stress, predicted from network models in Arabidopsis.

MicroRNA (miRNA) mediated changes in gene expression by post-transcriptional modulation of major regulatory transcription factors is a potent mechanism for integrating growth and stress-related responses. Exotic plants including many traditional varieties of Andean potatoes (Solanum tuberosum subsp. andigena) are known for better adaptation to marginal environments. Stress physiological studies confirmed earlier reports on the salinity tolerance potentials of certain andigena cultivars. Guided by the hypothesis that certain miRNAs play important roles in growth modulation under suboptimal conditions, we identified and characterized salinity stress-responsive miRNA-target gene pairs in the andigena cultivar Sullu by parallel analysis of noncoding and coding RNA transcriptomes. Inverse relationships were established by the reverse co-expression between two salinity stress-regulated miRNAs (miR166, miR159) and their target transcriptional regulators HD-ZIP-Phabulosa/Phavulota and Myb101, respectively. Based on heterologous models in Arabidopsis, the miR166-HD-ZIP-Phabulosa/Phavulota network appears to be involved in modulating growth perhaps by mediating vegetative dormancy, with linkages to defense-related pathways. The miR159-Myb101 network may be important for the modulation of vegetative growth while also controlling stress-induced premature transition to reproductive phase. We postulate that the induction of miR166 and miR159 under salinity stress represents important network hubs for balancing gene expression required for basal growth adjustments.

Metagenomic investigation reveals bacteriophage-mediated horizontal transfer of antibiotic resistance genes in microbial communities of an organic agricultural ecosystem.

Agricultural microbiomes are major reservoirs of antibiotic resistance genes (ARGs), posing continuous risks to human health. To understand the role of bacteriophages as vehicles for the horizontal transfer of ARGs in the agricultural microbiome, we investigated the diversity of bacterial and viral microbiota from fecal and environmental samples on an organic farm. The profiles of the microbiome indicated the highest abundance of Bacteroidetes, Firmicutes, and Proteobacteria phyla in animal feces, with varying Actinobacteria and Spirochaetes abundance across farm animals. The most predominant composition in environmental samples was the phylum Proteobacteria. Compared to the microbiome profiles, the trends in virome indicated much broader diversity with more specific signatures between the fecal and environmental samples. Overall, viruses belonging to the order Caudovirales were the most prevalent across the agricultural samples. Additionally, the similarities within and between fecal and environmental components of the agricultural environment based on ARG-associated bacteria alone were much lower than those of total microbiome composition. However, there were significant similarities in the profiles of ARG-associated viruses across the fecal and environmental components. Moreover, the predictive models of phage-bacterial interactions on bipartite ARG transfer networks indicated that phages belonging to the order Caudovirales, particularly in the Siphoviridae family, contained diverse ARG types in different samples. Their interaction with various bacterial hosts further implied the important role of bacteriophages in ARG transmission across bacterial populations. Our findings provided a novel insight into the potential mechanisms of phage-mediated ARG transmission and their correlation with resistome evolution in natural agricultural environments. IMPORTANCE Antibiotic resistance has become a serious health concern worldwide. The potential impact of viruses, bacteriophages in particular, on spreading antibiotic resistance genes is still controversial due to the complexity of bacteriophage-bacterial interactions within diverse environments. In this study, we determined the microbiome profiles and the potential antibiotic resistance gene (ARG) transfer between bacterial and viral populations in different agricultural samples using a high-resolution analysis of the metagenomes. The results of this study provide compelling genetic evidence for ARG transfer through bacteriophage-bacteria interactions, revealing the inherent risks associated with bacteriophage-mediated ARG transfer across the agricultural microbiome.

Novel genes and alleles of the BTB/POZ protein family in Oryza rufipogon.

The BTB/POZ family of proteins is widespread in plants and animals, playing important roles in development, growth, metabolism, and environmental responses. Although members of the expanded BTB/POZ gene family (OsBTB) have been identified in cultivated rice (Oryza sativa), their conservation, novelty, and potential applications for allele mining in O. rufipogon, the direct progenitor of O. sativa ssp. japonica and potential wide-introgression donor, are yet to be explored. This study describes an analysis of 110 BTB/POZ encoding gene loci (OrBTB) across the genome of O. rufipogon as outcomes of tandem duplication events. Phylogenetic grouping of duplicated OrBTB genes was supported by the analysis of gene sequences and protein domain architecture, shedding some light on their evolution and functional divergence. The O. rufipogon genome encodes nine novel BTB/POZ genes with orthologs in its distant cousins in the family Poaceae (Sorghum bicolor, Brachypodium distachyon), but such orthologs appeared to have been lost in its domesticated descendant, O. sativa ssp. japonica. Comparative sequence analysis and structure comparisons of novel OrBTB genes revealed that diverged upstream regulatory sequences and regulon restructuring are the key features of the evolution of this large gene family. Novel genes from the wild progenitor serve as a reservoir of potential new alleles that can bring novel functions to cultivars when introgressed by wide hybridization. This study establishes a foundation for hypothesis-driven functional genomic studies and their applications for widening the genetic base of rice cultivars through the introgression of novel genes or alleles from the exotic gene pool.

Cis-regulatory signatures of orthologous stress-associated bZIP transcription factors from rice, sorghum and Arabidopsis based on phylogenetic footprints.

BACKGROUND: The potential contribution of upstream sequence variation to the unique features of orthologous genes is just beginning to be unraveled. A core subset of stress-associated bZIP transcription factors from rice (Oryza sativa) formed ten clusters of orthologous groups (COG) with genes from the monocot sorghum (Sorghum bicolor) and dicot Arabidopsis (Arabidopsis thaliana). The total cis-regulatory information content of each stress-associated COG was examined by phylogenetic footprinting to reveal ortholog-specific, lineage-specific and species-specific conservation patterns. RESULTS: The most apparent pattern observed was the occurrence of spatially conserved 'core modules' among the COGs but not among paralogs. These core modules are comprised of various combinations of two to four putative transcription factor binding site (TFBS) classes associated with either developmental or stress-related functions. Outside the core modules are specific stress (ABA, oxidative, abiotic, biotic) or organ-associated signals, which may be functioning as 'regulatory fine-tuners' and further define lineage-specific and species-specific cis-regulatory signatures. Orthologous monocot and dicot promoters have distinct TFBS classes involved in disease and oxidative-regulated expression, while the orthologous rice and sorghum promoters have distinct combinations of root-specific signals, a pattern that is not particularly conserved in Arabidopsis. CONCLUSIONS: Patterns of cis-regulatory conservation imply that each ortholog has distinct signatures, further suggesting that they are potentially unique in a regulatory context despite the presumed conservation of broad biological function during speciation. Based on the observed patterns of conservation, we postulate that core modules are likely primary determinants of basal developmental programming, which may be integrated with and further elaborated by additional intrinsic or extrinsic signals in conjunction with lineage-specific or species-specific regulatory fine-tuners. This synergy may be critical for finer-scale spatio-temporal regulation, hence unique expression profiles of homologous transcription factors from different species with distinct zones of ecological adaptation such as rice, sorghum and Arabidopsis. The patterns revealed from these comparisons set the stage for further empirical validation by functional genomics.

Applicability of hyperspectral imaging during salinity stress in rice for tracking Na+ and K+ levels in planta.

The ratio of Na+ and K+ is an important determinant of the magnitude of Na+ toxicity and osmotic stress in plant cells. Traditional analytical approaches involve destructive tissue sampling and chemical analysis, where real-time observation of spatio-temporal experiments across genetic or breeding populations is unrealistic. Such an approach can also be very inaccurate and prone to erroneous biological interpretation. Analysis by Hyperspectral Imaging (HSI) is an emerging non-destructive alternative for tracking plant nutrient status in a time-course with higher accuracy and reduced cost for chemical analysis. In this study, the feasibility and predictive power of HSI-based approach for spatio-temporal tracking of Na+ and K+ levels in tissue samples was explored using a panel recombinant inbred line (RIL) of rice (Oryza sativa L.; salt-sensitive IR29 x salt-tolerant Pokkali) with differential activities of the Na+ exclusion mechanism conferred by the SalTol QTL. In this panel of RILs the spectrum of salinity tolerance was represented by FL499 (super-sensitive), FL454 (sensitive), FL478 (tolerant), and FL510 (super-tolerant). Whole-plant image processing pipeline was optimized to generate HSI spectra during salinity stress at EC = 9 dS m-1. Spectral data was used to create models for Na+ and K+ prediction by partial least squares regression (PLSR). Three datasets, i.e., mean image pixel spectra, smoothened version of mean image pixel spectra, and wavelength bands, with wide differences in intensity between control and salinity facilitated the prediction models with high R2. The smoothened and filtered datasets showed significant improvements over the mean image pixel dataset. However, model prediction was not fully consistent with the empirical data. While the outcome of modeling-based prediction showed a great potential for improving the throughput capacity for salinity stress phenotyping, additional technical refinements including tissue-specific measurements is necessary to maximize the accuracy of prediction models.

DECUSSATE network with flowering genes explains the variable effects of qDTY12.1 to rice yield under drought across genetic backgrounds.

The impact of qDTY12.1 in maintaining yield under drought has not been consistent across genetic backgrounds. We hypothesized that synergism or antagonism with additive-effect peripheral genes across the background genome either enhances or undermines its full potential. By modeling the transcriptional networks across sibling qDTY12.1-introgression lines with contrasting yield under drought (LPB = low-yield penalty; HPB = high-yield penalty), the qDTY12.1-encoded DECUSSATE gene (OsDEC) was revealed as the core of a synergy with other genes in the genetic background. OsDEC is expressed in flag leaves and induced by progressive drought at booting stage in LPB but not in HPB. The unique OsDEC signature in LPB is coordinated with 35 upstream and downstream peripheral genes involved in floral development through the cytokinin signaling pathway. Results support the differential network rewiring effects through genetic coupling-uncoupling between qDTY12.1 and other upstream and downstream peripheral genes across the distinct genetic backgrounds of LPB and HPB. The functional DEC-network in LPB defines a mechanism for early flowering as a means for avoiding the drought-induced depletion of photosynthate needed for reproductive growth. Its impact is likely through the timely establishment of stronger source-sink dynamics that sustains a robust reproductive transition under drought.

Responses of Escherichia coli and Listeria monocytogenes to ozone treatment on non-host tomato: Efficacy of intervention and evidence of induced acclimation.

Because of the continuous rise of foodborne illnesses caused by the consumption of raw fruits and vegetables, effective post-harvest anti-microbial strategies are necessary. The aim of this study was to evaluate the anti-microbial efficacy of ozone (O3) against two common causes of fresh produce contamination, the Gram-negative Escherichia coli O157:H7 and Gram-positive Listeria monocytogenes, and to relate its effects to potential mechanisms of xenobiosis by transcriptional network modeling. The study on non-host tomato environment correlated the dose × time aspects of xenobiosis by examining the correlation between bacterial survival in terms of log-reduction and defense responses at the level of gene expression. In E. coli, low (1 µg O3/g of fruit) and moderate (2 µg O3/g of fruit) doses caused insignificant reduction in survival, while high dose (3 µg/g of fruit) caused significant reduction in survival in a time-dependent manner. In L. monocytogenes, moderate dose caused significant reduction even with short-duration exposure. Distinct responses to O3 xenobiosis between E. coli and L. monocytogenes are likely related to differences in membrane and cytoplasmic structure and components. Transcriptome profiling by RNA-Seq showed that primary defenses in E. coli were attenuated after exposure to a low dose, while the responses at moderate dose were characterized by massive upregulation of pathogenesis and stress-related genes, which implied the activation of defense responses. More genes were downregulated during the first hour at high dose, with a large number of such genes getting significantly upregulated after 2 hr and 3 hr. This trend suggests that prolonged exposure led to potential adaptation. In contrast, massive downregulation of genes was observed in L. monocytogenes regardless of dose and exposure duration, implying a mechanism of defense distinct from that of E. coli. The nature of bacterial responses revealed by this study should guide the selection of xenobiotic agents for eliminating bacterial contamination on fresh produce without overlooking the potential risks of adaptation.

Novel and Transgressive Salinity Tolerance in Recombinant Inbred Lines of Rice Created by Physiological Coupling-Uncoupling and Network Rewiring Effects.

The phenomenon of transgressive segregation, where a small minority of recombinants are outliers relative to the range of parental phenotypes, is commonly observed in plant breeding populations. While this phenomenon has been attributed to complementation and epistatic effects, the physiological and developmental synergism involved have not been fully illuminated by the QTL mapping approach alone, especially for stress-adaptive traits involving highly complex interactions. By systems-level profiling of the IR29 × Pokkali recombinant inbred population of rice, we addressed the hypothesis that novel salinity tolerance phenotypes are created by reconfigured physiological networks due to positive or negative coupling-uncoupling of developmental and physiological attributes of each parent. Real-time growth and hyperspectral profiling distinguished the transgressive individuals in terms of stress penalty to growth. Non-parental network signatures that led to either optimal or non-optimal integration of developmental with stress-related mechanisms were evident at the macro-physiological, biochemical, metabolic, and transcriptomic levels. Large positive net gain in super-tolerant progeny was due to ideal complementation of beneficial traits while shedding antagonistic traits. Super-sensitivity was explained by the stacking of multiple antagonistic traits and loss of major beneficial traits. The synergism uncovered by the phenomics approach in this study supports the modern views of the Omnigenic Theory, emphasizing the synergy or lack thereof between core and peripheral components. This study also supports a breeding paradigm rooted on genomic modeling from multi-dimensional genetic, physiological, and phenotypic profiles to create novel adaptive traits for new crop varieties of the 21st century.

Transcriptional regulatory network triggered by oxidative signals configures the early response mechanisms of japonica rice to chilling stress.

BACKGROUND: The transcriptional regulatory network involved in low temperature response leading to acclimation has been established in Arabidopsis. In japonica rice, which can only withstand transient exposure to milder cold stress (10 degrees C), an oxidative-mediated network has been proposed to play a key role in configuring early responses and short-term defenses. The components, hierarchical organization and physiological consequences of this network were further dissected by a systems-level approach. RESULTS: Regulatory clusters responding directly to oxidative signals were prominent during the initial 6 to 12 hours at 10 degrees C. Early events mirrored a typical oxidative response based on striking similarities of the transcriptome to disease, elicitor and wounding induced processes. Targets of oxidative-mediated mechanisms are likely regulated by several classes of bZIP factors acting on as1/ocs/TGA-like element enriched clusters, ERF factors acting on GCC-box/JAre-like element enriched clusters and R2R3-MYB factors acting on MYB2-like element enriched clusters.Temporal induction of several H2O2-induced bZIP, ERF and MYB genes coincided with the transient H2O2 spikes within the initial 6 to 12 hours. Oxidative-independent responses involve DREB/CBF, RAP2 and RAV1 factors acting on DRE/CRT/rav1-like enriched clusters and bZIP factors acting on ABRE-like enriched clusters. Oxidative-mediated clusters were activated earlier than ABA-mediated clusters. CONCLUSION: Genome-wide, physiological and whole-plant level analyses established a holistic view of chilling stress response mechanism of japonica rice. Early response regulatory network triggered by oxidative signals is critical for prolonged survival under sub-optimal temperature. Integration of stress and developmental responses leads to modulated growth and vigor maintenance contributing to a delay of plastic injuries.

Supra-optimal expression of the cold-regulated OsMyb4 transcription factor in transgenic rice changes the complexity of transcriptional network with major effects on stress tolerance and panicle development.

The R2R3-type OsMyb4 transcription factor of rice has been shown to play a role in the regulation of osmotic adjustment in heterologous overexpression studies. However, the exact composition and organization of its underlying transcriptional network has not been established to be a robust tool for stress tolerance enhancement by regulon engineering. OsMyb4 network was dissected based on commonalities between the global chilling stress transcriptome and the transcriptome configured by OsMyb4 overexpression. OsMyb4 controls a hierarchical network comprised of several regulatory sub-clusters associated with cellular defense and rescue, metabolism and development. It regulates target genes either directly or indirectly through intermediary MYB, ERF, bZIP, NAC, ARF and CCAAT-HAP transcription factors. Regulatory sub-clusters have different combinations of MYB-like, GCC-box-like, ERD1-box-like, ABRE-like, G-box-like, as1/ocs/TGA-like, AuxRE-like, gibberellic acid response element (GARE)-like and JAre-like cis-elements. Cold-dependent network activity enhanced cellular antioxidant capacity through radical scavenging mechanisms and increased activities of phenylpropanoid and isoprenoid metabolic processes involving various abscisic acid (ABA), jasmonic acid (JA), salicylic acid (SA), ethylene and reactive oxygen species (ROS) responsive genes. OsMyb4 network is independent of drought response element binding protein/C-repeat binding factor (DREB/CBF) and its sub-regulons operate with possible co-regulators including nuclear factor-Y. Because of its upstream position in the network hierarchy, OsMyb4 functions quantitatively and pleiotrophically. Supra-optimal expression causes misexpression of alternative targets with costly trade-offs to panicle development.

Networks of Physiological Adjustments and Defenses, and Their Synergy With Sodium (Na+) Homeostasis Explain the Hidden Variation for Salinity Tolerance Across the Cultivated Gossypium hirsutum Germplasm.

The abilities to mobilize and/or sequester excess ions within and outside the plant cell are important components of salt-tolerance mechanisms. Mobilization and sequestration of Na+ involves three transport systems facilitated by the plasma membrane H+/Na+ antiporter (SOS1), vacuolar H+/Na+ antiporter (NHX1), and Na+/K+ transporter in vascular tissues (HKT1). Many of these mechanisms are conserved across the plant kingdom. While Gossypium hirsutum (upland cotton) is significantly more salt-tolerant relative to other crops, the critical factors contributing to the phenotypic variation hidden across the germplasm have not been fully unraveled. In this study, the spatio-temporal patterns of Na+ accumulation along with other physiological and biochemical interactions were investigated at different severities of salinity across a meaningful genetic diversity panel across cultivated upland Gossypium. The aim was to define the importance of holistic or integrated effects relative to the direct effects of Na+ homeostasis mechanisms mediated by GhHKT1, GhSOS1, and GhNHX1. Multi-dimensional physio-morphometric attributes were investigated in a systems-level context using univariate and multivariate statistics, randomForest, and path analysis. Results showed that mobilized or sequestered Na+ contributes significantly to the baseline tolerance mechanisms. However, the observed variance in overall tolerance potential across a meaningful diversity panel were more significantly attributed to antioxidant capacity, maintenance of stomatal conductance, chlorophyll content, and divalent cation (Mg2+) contents other than Ca2+ through a complex interaction with Na+ homeostasis. The multi-tier macro-physiological, biochemical and molecular data generated in this study, and the networks of interactions uncovered strongly suggest that a complex physiological and biochemical synergy beyond the first-line-of defense (Na+ sequestration and mobilization) accounts for the total phenotypic variance across the primary germplasm of Gossypium hirsutum. These findings are consistent with the recently proposed Omnigenic Theory for quantitative traits and should contribute to a modern look at phenotypic selection for salt tolerance in cotton breeding.

Contributions of Adaptive Plant Architecture to Transgressive Salinity Tolerance in Recombinant Inbred Lines of Rice: Molecular Mechanisms Based on Transcriptional Networks.

Genetic novelties are important nucleators of adaptive speciation. Transgressive segregation is a major mechanism that creates genetic novelties with morphological and developmental attributes that confer adaptive advantages in certain environments. This study examined the morpho-developmental and physiological profiles of recombinant inbred lines (RILs) from the salt-sensitive IR29 and salt-tolerant Pokkali rice, representing the total range of salt tolerance including the outliers at both ends of the spectrum. Morpho-developmental and physiological profiles were integrated with a hypothesis-driven interrogation of mRNA and miRNA transcriptomes to uncover the critical genetic networks that have been rewired for novel adaptive architecture. The transgressive super-tolerant FL510 had a characteristic small tiller angle and wider, more erect, sturdier, and darker green leaves. This unique morphology resulted in lower transpiration rate, which also conferred a special ability to retain water more efficiently for osmotic avoidance. The unique ability for water retention conferred by such adaptive morphology appeared to enhance the efficacy of defenses mediated by Na+ exclusion mechanism (SalTol-effects) inherited from Pokkali. The super-tolerant FL510 and super-sensitive FL499 had the smallest proportions of differentially expressed genes with little overlaps. Genes that were steadily upregulated in FL510 comprised a putative cytokinin-regulated genetic network that appeared to maintain robust growth under salt stress through well-orchestrated cell wall biogenesis and cell expansion, likely through major regulatory (OsRR23, OsHK5) and biosynthetic (OsIPT9) genes in the cytokinin signaling pathway. Meanwhile, a constitutively expressed cluster in FL510 prominently featured two transcription factors (OsIBH1, TAC3) that control tiller angle and growth habit through the brassinosteroid signaling pathway. Both the putative cytokinin-mediated and brassinosteroid-mediated clusters appeared to function as highly coordinated network synergies in FL510. In contrast, both networks appeared to be sub-optimal and inferior in the other RILs and parents as they were disjointed and highly fragmented. Transgressively expressed miRNAs (miR169, miR397, miR827) were also identified as prominent signatures of FL510, with functional implications to mechanisms that support robust growth, homeostasis, and osmotic stress avoidance. Results of this study demonstrate how genetic recombination creates novel morphology that complements inducible defenses hence transgressive adaptive phenotypes.

Novel allelic variation in the Phospholipase D alpha1 gene (OsPLDα1) of wild Oryza species implies to its low expression in rice bran.

Rice bran, a by-product after milling, is a rich source of phytonutrients like oryzanols, tocopherols, tocotrienols, phytosterols, and dietary fibers. Moreover, exceptional properties of the rice bran oil make it unparalleled to other vegetable oils. However, a lipolytic enzyme Phospholipase D alpha1 (OsPLDα1) causes rancidity and 'stale flavor' in the oil, and thus limits the rice bran usage for human consumption. To improve the rice bran quality, sequence based allele mining at OsPLDα1 locus (3.6 Kb) was performed across 48 accessions representing 11 wild Oryza species, 8 accessions of African cultivated rice, and 7 Oryza sativa cultivars. From comparative sequence analysis, 216 SNPs and 30 InDels were detected at the OsPLDα1 locus. Phylogenetic analysis revealed 20 OsPLDα1 cDNA variants which further translated into 12 protein variants. The O. officinalis protein variant, when compared to Nipponbare, showed maximum variability comprising 22 amino acid substitutions and absence of two peptides and two ß-sheets. Further, expression profiling indicated significant differences in transcript abundance within as well as between the OsPLDα1 variants. Also, a new OsPLDα1 transcript variant having third exon missing in it, Os01t0172400-06, has been revealed. An O. officinalis accession (IRGC101152) had lowest gene expression which suggests the presence of novel allele, named as OsPLDα1-1a (GenBank accession no. MF966931). The identified novel allele could be further deployed in the breeding programs to overcome rice bran rancidity in elite cultivars.

Xenobiotic Effects of Chlorine Dioxide to Escherichia coli O157:H7 on Non-host Tomato Environment Revealed by Transcriptional Network Modeling: Implications to Adaptation and Selection.

Escherichia coli serotype O157:H7 is one of the major agents of pathogen outbreaks associated with fresh fruits and vegetables. Gaseous chlorine dioxide (ClO2) has been reported to be an effective intervention to eliminate bacterial contamination on fresh produce. Although remarkable positive effects of low doses of ClO2 have been reported, the genetic regulatory machinery coordinating the mechanisms of xenobiotic effects and the potential bacterial adaptation remained unclear. This study examined the temporal transcriptome profiles of E. coli O157:H7 during exposure to different doses of ClO2 in order to elucidate the genetic mechanisms underlying bacterial survival under such harsh conditions. Dosages of 1 µg, 5 µg, and 10 µg ClO2 per gram of tomato fruits cause different effects with dose-by-time dynamics. The first hour of exposure to 1 µg and 5 µg ClO2 caused only partial killing with significant growth reduction starting at the second hour, and without further significant reduction at the third hour. However, 10 µg ClO2 exposure led to massive bacterial cell death at 1 h with further increase in cell death at 2 and 3 h. The first hour exposure to 1 µg ClO2 caused activation of primary defense and survival mechanisms. However, the defense response was attenuated during the second and third hours. Upon treatment with 5 µg ClO2, the transcriptional networks showed massive downregulation of pathogenesis and stress response genes at the first hour of exposure, with decreasing number of differentially expressed genes at the second and third hours. In contrast, more genes were further downregulated with exposure to 10 µg ClO2 at the first hour, with the number of both upregulated and downregulated genes significantly decreasing at the second hour. A total of 810 genes were uniquely upregulated at the third hour at 10 µg ClO2, suggesting that the potency of xenobiotic effects had led to potential adaptation. This study provides important knowledge on the possible selection of target molecules for eliminating bacterial contamination on fresh produce without overlooking potential risks of adaptation.

Reverse engineering module networks by PSO-RNN hybrid modeling.

BACKGROUND: Inferring a gene regulatory network (GRN) from high throughput biological data is often an under-determined problem and is a challenging task due to the following reasons: (1) thousands of genes are involved in one living cell; (2) complex dynamic and nonlinear relationships exist among genes; (3) a substantial amount of noise is involved in the data, and (4) the typical small sample size is very small compared to the number of genes. We hypothesize we can enhance our understanding of gene interactions in important biological processes (differentiation, cell cycle, and development, etc) and improve the inference accuracy of a GRN by (1) incorporating prior biological knowledge into the inference scheme, (2) integrating multiple biological data sources, and (3) decomposing the inference problem into smaller network modules. RESULTS: This study presents a novel GRN inference method by integrating gene expression data and gene functional category information. The inference is based on module network model that consists of two parts: the module selection part and the network inference part. The former determines the optimal modules through fuzzy c-mean (FCM) clustering and by incorporating gene functional category information, while the latter uses a hybrid of particle swarm optimization and recurrent neural network (PSO-RNN) methods to infer the underlying network between modules. Our method is tested on real data from two studies: the development of rat central nervous system (CNS) and the yeast cell cycle process. The results are evaluated by comparing them to previously published results and gene ontology annotation information. CONCLUSION: The reverse engineering of GRNs in time course gene expression data is a major obstacle in system biology due to the limited number of time points. Our experiments demonstrate that the proposed method can address this challenge by: (1) preprocessing gene expression data (e.g. normalization and missing value imputation) to reduce the data noise; (2) clustering genes based on gene expression data and gene functional category information to identify biologically meaningful modules, thereby reducing the dimensionality of the data; (3) modeling GRNs with the PSO-RNN method between the modules to capture their nonlinear and dynamic relationships. The method is shown to lead to biologically meaningful modules and networks among the modules.

Application of cranberry concentrate (Vaccinium macrocarpon) to control Escherichia coli O157:H7 in ground beef and its antimicrobial mechanism related to the downregulated slp, hdeA and cfa.

The possible use of cranberry concentrate (CC) as a natural food preservative was studied by examining its antimicrobial effect on the growth of Escherichia coli O157:H7 inoculated in ground beef, its organoleptical effect on beef patties, and its antimicrobial mechanism on the gene regulation level. Inoculated ground beef was added with CC and stored at 4 degrees C for 5 days. Bacteria were detected on day 0, 1, 3, and 5. Cranberry concentrate (2.5%, 5%, and 7.5% w/w) reduced total aerobic bacteria 1.5 log, 2.1 log, and 2.7 log CFU/g and E. coli O157:H7 0.4 log, 0.7 log, and 2.4 log CFU/g, respectively, when compared to the control on day 5. Fifty panelists evaluated the burgers supplemented with CC. No differences in appearance, flavor, and taste were found among burgers with 0%, 2.5%, and 5% CC. The expression of E. coli O157:H7 cyclopropane fatty acyl phospholipid synthase (cfa), hypothetical protein (hdeA), outer membrane porin protein C (ompC), hyperosmotically inducible periplasmic protein (osmY), and outer membrane protein induced after carbon starvation (slp) genes with or without CC (2.5% v/v) treatment was investigated by quantitative real-time PCR. Compared to the control, slp, hdeA, and cfa were markedly downregulated, ompC was slightly downregulated, while osmY was slightly affected.

Genomic and epigenomic bases of transgressive segregation - New breeding paradigm for novel plant phenotypes.

For a holistic approach in developing the stress-resilient crops of the 21st century, modern genomic biology will need to re-envision the underappreciated phenomena in classical genetics, and incorporate them into the new plant breeding paradigm. Advances in evolutionary genomics support a theory that genetic recombination under genome shock during hybridization of widely divergent parents is an important driver of adaptive speciation, by virtue of the novelties of rare hybrids and recombinants. The enormous potential of genetic network rewiring to generate developmental or physiological novelties with adaptive advantage to special ecological niches has been appreciated. Developmental and physiological reconfiguration through network rewiring involves intricate molecular synergies controlled both at the genetic and epigenetic levels, as typified by the phenomenon of transgressive segregation, observed in both natural and breeding populations. This paper presents modern views on the possible molecular underpinnings of transgressive phenotypes as they are created in plant breeding, expanded from classical explanations through the Omnigenic Theory for quantitative traits and modern paradigms of epigenetics. Perspectives on how genomic biology can fully exploit this phenomenon to create novel phenotypes beyond what could be achieved through the more reductionist approach of functional genomics are presented in context of genomic modeling.

Simultaneous Colorimetric Detection of a Variety of Salmonella spp. in Food and Environmental Samples by Optical Biosensing Using Oligonucleotide-Gold Nanoparticles.

Optical biosensors for rapid detection of significant foodborne pathogens are steadily gaining popularity due to its simplicity and sensitivity. While nanomaterials such as gold nanoparticles (AuNPs) are commonly used as signal amplifiers for optical biosensors, AuNPs can also be utilized as a robust biosensing platform. Many reported optical biosensors were designed for individual pathogen detection in a single assay and have high detection limit (DL). Salmonella spp. is one of the major causative agents of foodborne sickness, hospitalization and deaths. Unfortunately, there are around 2,000 serotypes of Salmonella worldwide, and rapid and simultaneous detection of multiple strains in a single assay is lacking. In this study, a comprehensive and highly sensitive simultaneous colorimetric detection of nineteen (19) environmental and outbreak Salmonella spp. strains was achieved by a novel optical biosensing platform using oligonucleotide-functionalized AuNPs. A pair of newly designed single stranded oligonucleotides (30-mer) was displayed onto the surface of AuNPs (13 nm) as detection probes to hybridize with a conserved genomic region (192-bases) of ttrRSBCA found on a broad range of Salmonella spp. strains. The sandwich hybridization (30 min, 55°C) resulted in a structural formation of highly stable oligonucleotide/AuNPs-DNA complexes which remained undisturbed even after subjecting to an increased salt concentration (2 M, final), thus allowing a direct discrimination via color change of target (red color) from non-target (purplish-blue color) reaction mixtures by direct observation using the naked eye. In food matrices (blueberries and chicken meat), nineteen different Salmonella spp. strains were concentrated using immunomagnetic separation and then simultaneously detected in a 96-well microplate by oligonucleotide-functionalized AuNPs after DNA preparation. Successful oligonucleotide/AuNPs-DNA hybridization was confirmed by gel electrophoresis while AuNPs aggregation in non-target and control reaction mixtures was verified by both spectrophotometric analysis and TEM images. Results showed that the optical AuNP biosensing platform can simultaneously screen nineteen (19) viable Salmonella spp. strains tested with 100% specificity and a superior detection limit of <10 CFU/mL or g for both pure culture and complex matrices setups. The highly sensitive colorimetric detection system can significantly improve the screening and detection of viable Salmonella spp. strains present in complex food and environmental matrices, therefore reducing the risks of contamination and incidence of foodborne diseases.