RPS2 Research Articles

Comparative analysis of sipeimine content, metabolome and chloroplast genome in cultivated and wild varieties of Fritillaria taipaiensis.

The wild variety Fritillaria taipaiensis E.B (EB) is known for its superior therapeutic effects, but its limited production cannot meet demand. As a result, the cultivated variety F. taipaiensis P. Y. Li (PY) has been widely grown. In this study, we conducted a comprehensive analysis comparing EB and PY in terms of external features, sipeimine content, metabolome and chloroplast genome to differentiate these two varieties. Our research revealed that the petals and pods of EB are green, while those of PY have purple markings. The bulbs of EB contain significantly higher levels of sipeimine compared to those of PY. Metabolomic analysis identified 56 differentially expressed metabolites (DMs), with 23 upregulated and 33 downregulated in EB bulbs. Particularly, 3-hydroxycinnamic acid and secoxyloganin may serve as distinctive DMs. These DMs were associated with 17 KEGG pathways, including pyrimidine metabolism, alanine, aspartate and glutamate metabolism, and galactose metabolism. Differences in the length of the chloroplast genome were primarily observed in the large single-copy (LSC) region, with the largest variation in the trnH-GUC-psbA region. The placement of the trnH gene and the rps gene in proximity to the LSC/IRb boundary differs between EB and PY. The results of this study provide valuable insights for the introduction and comprehensive development of wild F. taipaiensis from a scientific perspective. © 2024 Society of Chemical Industry.

Comparative analysis the chloroplast genomes of Celastrus (Celastraceae) species: Provide insights into molecular evolution, species identification and phylogenetic relationships

BackgroundThe genus Celastrus is an important medicinal plant resource. The similarity of morphology and the lack of complete chloroplast genome analysis have significantly impeded the exploration of species identification, molecular evolution and phylogeny of Celastrus. PurposeIn order to resolve the phylogenic controversy of Celastrus species, the chloroplast genome comparative analysis was performed to provide genetic evidence. MethodsIn this study, we collected and sequenced ten chloroplast genomes of Celastrus species from China and downloaded three chloroplast genomes from the databases. The chloroplast genomes were compared and analyzed to explore their characteristics and evolution. Furthermore, the phylogenetic relationships of Celastrus species were inferred based on the whole chloroplast genomes and protein-coding genes. ResultsAll the 13 Celastrus species chloroplast genomes showed a typical quadripartite structure with genome sizes ranging from 155,113 to 157,366 bp. The intron loss of the rps16 gene occurred in all the 13 Celastrus species. The GC content, gene sequence, repeat types and codon bias pattern were highly conserved. Ten highly variation regions were identified, which can be used as potential DNA markers in molecular identification of Celastrus species. Eight genes, including accD, atp4, ndhB, rpoC1, rbcL, rpl2, rpl20 and ycf1, were detected to experience positive selection. Phylogenetic analysis showed that Celastrus was a monophyletic group and Tripterygium was the closest sister-group. Noteworthy, C. gemmatus Loes. and C. orbiculatus Thunb. can be discriminated using the chloroplast genome as a super barcode. The comparative and phylogenetic analysis results proposed that C. tonkinensis Pitard. was the synonym of C. hindsii Benth. ConclusionThe comparative analysis of the Celastrus chloroplast genomes can provide comprehensive genetic evidence for molecular evolution, species identification and phylogenetic relationships.

Twelve newly assembled jasmine chloroplast genomes: unveiling genomic diversity, phylogenetic relationships and evolutionary patterns among Oleaceae and Jasminum species

BackgroundJasmine (Jasminum), renowned for its ornamental value and captivating fragrance, has given rise to numerous species and accessions. However, limited knowledge exists regarding the evolutionary relationships among various Jasminum species.ResultsIn the present study, we sequenced seven distinct Jasminum species, resulting in the assembly of twelve high-quality complete chloroplast (cp) genomes. Our findings revealed that the size of the 12 cp genomes ranged from 159 to 165 kb and encoded 134–135 genes, including 86–88 protein-coding genes, 38–40 tRNA genes, and 8 rRNA genes. J. nudiflorum exhibited a larger genome size compared to other species, mainly attributed to the elevated number of forward repeats (FRs). Despite the typically conservative nature of chloroplasts, variations in the presence or absence of accD have been observed within J. sambac. The calculation of nucleotide diversity (Pi) values for 19 cp genomes indicated that potential mutation hotspots were more likely to be located in LSC regions than in other regions, particularly in genes ycf2, rbcL, atpE, ndhK, and ndhC (Pi > 0.2). Ka/Ks values revealed strong selection pressure on the genes rps2, atpA, rpoA, rpoC1, and rpl33 when comparing J. sambac with the three most closely related species (J. auriculatum, J. multiflorum, and J. dichotomum). Additionally, SNP identification, along with the results of Structure, PCA, and phylogenetic tree analyses, divided the Jasminum cp genomes into six groups. Notably, J. polyanthum showed gene flow signals from both the G5 group (J. nudiflorum) and the G3 group (J. tortuosum and J. fluminense). Phylogenetic tree analysis reflected that most species from the same genus clustered together with robust support in Oleaceae, strongly supporting the monophyletic nature of cp genomes within the genus Jasminum.ConclusionOverall, this study provides comprehensive insights into the genomic composition, variation, and phylogenetic relationships among various Jasminum species. These findings enhance our understanding of the genetic diversity and evolutionary history of Jasminum.

Complete Mitochondrial Genome and Its Phylogenetic Position in Red Algae Fushitsunagia catenata from South Korea.

The mitogenome is an important tool in taxonomic and evolutionary studies. Only a few complete mitogenomes have been reported for red algae. Herein, we reported the complete mitochondrial genome sequence of Fushitsunagia catenata (Harvey) Filloramo, G.V. and Saunders, G.W. 2016, a monospecific genus. The genome was 25,889 bp in circumference and had a strongly biased AT of 70.4%. It consisted of 2 rRNAs, 23 tRNAs, and 24 protein-coding genes (PCGs). nad5 (1986 bp) was the largest and atp9 (231 bp) was the smallest PCG. All PCGs used ATG as an initiation codon and TAA as a termination codon, except TAG, which was the termination codon used in the sdh3, rps3, and rps11 genes. The general structure and gene content of the present findings were almost identical to those of other red algae genomes, particularly those of the Rhodymeniales order. The maximum likelihood analysis showed that F. catenata was closely related to Rhodymenia pseudopalmata. The mitochondrial genome data presented in this study will enhance our understanding of evolution in Rhodophyta species.

Circular RNA ath-circ032768, a competing endogenous RNA, response the drought stress by targeting miR472-RPS5 module.

CircRNAs (circular RNAs) reduce the abundance of miRNAs through ceRNA (competing endogenous RNA), to regulate many physiological processes and stress responses in plants. However, the role of circRNA in drought stress is poorly understood. Through ring identification and sequencing verification of ath-circ032768, bioinformatics analysis predicted the interaction of ath-circ032768-miR472-RPS5, and further obtained transgenic plants overexpressing ath-circ032768 and silencing STTM-miR472. The change in drought stress was analysed using biochemical and molecular biological methods. Sequencing and biological analysis confirmed that ath-circ032768, miR472 and RPS5 were responsive to drought stress, and changes in gene expression were consistent with the prediction of ceRNA. The silencing vectors ath-circ032768 and STTM-miR472 were constructed using molecular biology techniques, and stable transgenic plants with drought tolerance obtained. Further physiological and biochemical studies showed that ath-circ032768 could bind to miR472, and that miR472 could bind to the RPS5 gene, resulting in decreased expression of RPS5. Hence, ath-circ032768 can competitively inhibit degradation of RPS5 by miR472 through ceRNA. This process is accompanied by increased expression of DREB2A, RD29A and RD29B genes. Through the ath-circ032768-miR472-RPS5 pathway, the RPS5 stress resistance protein interacts with DREB2A protein to enhance expression of downstream drought resistance genes, RD29A and RD29B, and participate in the regulation mechanism of plant drought resistance, thereby improving drought tolerance of plants.

Intraspecific and Intrageneric Genomic Variation across Three Sedum Species (Crassulaceae): A Plastomic Perspective.

Sedum is the largest succulent genus in Crassulaceae. Because of predominant maternal inheritance, little recombination, and slow evolution, plastomes can serve as powerful super barcodes for inter- or intra-species phylogenetic analyses. While previous research has focused on plastomes between Sedum species, intra-species studies are scarce. Here, we sequenced plastomes from three Sedum species (Sedum alfredii, Sedum plumbizincicola, and Sedum japonicum) to understand their evolutionary relationships and plastome structural evolution. Our analyses revealed minimal size and GC content variation across species. However, gene distribution at IR boundaries, repeat structures, and codon usage patterns showed diversity at both inter-specific and intra-specific levels. Notably, an rps19 gene expansion and a bias toward A/T-ending codons were observed. Codon aversion motifs also varied, potentially serving as markers for future studies. Phylogenetic analyses confirmed the non-monophyly of Sedum and divided the Acre clade into two groups. Individuals from the same species clustered together, with strong support for the relationships between S. alfredii, S. tricarpum, and S. plumbizincicola. Additionally, S. japonicum clearly affiliates with the Acre clade. This study provides valuable insights into both intra-specific and intra-generic plastome variation in Sedum, as well as overall plastome evolution within the genus.

Exploring Plastomic Resources in Sempervivum (Crassulaceae): Implications for Phylogenetics.

The plastid organelle is vital for photosynthesis and energy production. Advances in sequencing technology have enabled the exploration of plastomic resources, offering insights into plant evolution, diversity, and conservation. As an important group of horticultural ornamentals in the Crassulaceae family, Sempervivum plants are known for their unique rosette-like structures and reproduction through offsets. Despite their popularity, the classification status of Sempervivum remains uncertain, with only a single plastome sequence currently available. Furthermore, codon usage bias (CUB) is a widespread phenomenon of the unbalanced usage of synonymous codons in the coding sequence (CDS). However, due to the limited available plastid data, there has been no research that focused on the CUB analysis among Sempervivum until now. To address these gaps, we sequenced and released the plastomes of seven species and one subspecies from Sempervivum, revealing several consistent patterns. These included a shared 110 bp extension of the rps19 gene, 14 hypervariable regions (HVRs) with distinct nucleotide diversity (π: 0.01173 to 0.02702), and evidence of selective pressures shaping codon usage. Notably, phylogenetic analysis robustly divided the monophyletic clade into two sections: Jovibarba and Sempervivum. In conclusion, this comprehensive plastomic resource provides valuable insights into Sempervivum evolution and offers potential molecular markers for DNA barcoding.

Comparative plastome analysis of the sister genera Ceratocephala and Myosurus (Ranunculaceae) reveals signals of adaptive evolution to arid and aquatic environments

BackgroundExpansion and contraction of inverted repeats can cause considerable variation of plastid genomes (plastomes) in angiosperms. However, little is known about whether structural variations of plastomes are associated with adaptation to or occupancy of new environments. Moreover, adaptive evolution of angiosperm plastid genes remains poorly understood. Here, we sequenced the complete plastomes for four species of xerophytic Ceratocephala and hydrophytic Myosurus, as well as Ficaria verna. By an integration of phylogenomic, comparative genomic, and selection pressure analyses, we investigated evolutionary patterns of plastomes in Ranunculeae and their relationships with adaptation to dry and aquatic habitats.ResultsOwing to the significant contraction of the boundary of IRA/LSC towards the IRA, plastome sizes and IR lengths of Myosurus and Ceratocephala are smaller within Ranunculeae. Compared to other Ranunculeae, the Myosurus plastome lost clpP and rps16, one copy of rpl2 and rpl23, and one intron of rpoC1 and rpl16, and the Ceratocephala plastome added an infA gene and lost one copy of rpl2 and two introns of clpP. A total of 11 plastid genes (14%) showed positive selection, two genes common to Myosurus and Ceratocephala, seven in Ceratocephala only, and two in Myosurus only. Four genes showed strong signals of episodic positive selection. The rps7 gene of Ceratocephala and the rpl32 and ycf4 genes of Myosurus showed an increase in the rate of variation close to 3.3 Ma.ConclusionsThe plastomic structure variations as well as the positive selection of two plastid genes might be related to the colonization of new environments by the common ancestor of Ceratocephala and Myosurus. The seven and two genes under positive selection might be related to the adaptation to dry and aquatic habitats in Ceratocephala and Myosurus, respectively. Moreover, intensified aridity and frequent sea-level fluctuations, as well as global cooling, might have favored an increased rate of change in some genes at about 3.3 Ma, associated with adaptation to dry and aquatic environments, respectively. These findings suggest that changing environments might have influenced structural variations of plastomes and fixed new mutations arising on some plastid genes owing to adaptation to specific habitats.

Expression of the RPM1-RIN4-RPS2 complex in two citrus species with contrasting response to Huanglongbing

Objetive/Antecedents. Persian lime (Citrus latifolia) shows a very high level of tolerance to Huanglongbing (HLB). A recent study suggests that genes from the RPM1-RIN4-RPS2 complex could be partly responsible for HLB tolerance in Persian lime, unlike other highly susceptible species such as orange (C. sinensis). The objective of this study was to compare the expression of this gene complex between orange, highly susceptible to HLB, and Persian lime, a tolerant species. Materials and Methods. Sequences of the three genes of the complex for orange and Persian lime were obtained from databases of previously published works, alignments and primer design for gene expression were performed using various bioinformatics tools. Subsequently, tissue samples from symptomatic HLB-infected orange and Persian lime were obtained and infection was confirmed. The expression of the RPM1-RIN4-RPS2 genes was compared using endpoint RT-PCR. Results. The presence of all three genes of the complex was determined in both orange and Persian lime, and it was also determined that they are highly conserved between both species. Additionally, it was observed that there is no differential expression for the RPM1 gene in symptomatic HLB tissue; however, there is a difference in the expression of the RPS2 and RIN4 genes. Conclusion. The results suggest that the contrasting response to HLB could be associated with the activity of the interaction of the RIN4 and RPS2 genes, thus, this could be of interest for citrus genetic improvement aiming at HLB control.

Identification and molecular mapping of a major gene conferring resistance to Phytophthora sansomeana in soybean 'Colfax'.

The first single dominant resistance gene contributing major resistance to the oomycete pathogen Phytophthora sansomeana was identified and mapped from soybean 'Colfax'. Phytophthora root rot (PRR) is one of the most important diseases in soybean (Glycine max). PRR is well known to be caused by Phytophthora sojae, but recent studies showed that P. sansomeana also causes extensive root rotof soybean. Depending upon the isolate, it might produce aggressive symptoms, especially in seeds and seedlings. Unlike P. sojae which can be effectively managed by Rps genes, no known majorresistance genes have yet been reported for P. sansomeana. Our previous study screened 470 soybean germplasm lines for resistance to P. sansomeana and found that soybean 'Colfax' (PI 573008) carries major resistance to the pathogen. In this study, we crossed 'Colfax' with a susceptible parent, 'Senaki', and developed three mapping populations with a total of 234 F2:3 families. Inheritance pattern analysis indicated a 1:2:1 ratio for resistant: segregating: susceptible lines among all the three populations, indicating a single dominant gene conferring the resistance in 'Colfax' (designatedas Rpsan1). Linkage analysis using extreme phenotypes anchored Rpsan1 to a 30Mb region on chromosome 3. By selecting nine polymorphic SNP markers within the region, Rpsan1 was genetically delimited into a 21.3cM region between Gm03_4487138_A_C and Gm03_5451606_A_C, which corresponds to a 1.06Mb genomic region containing nine NBS-LRR genes based on Gmax2.0 assembly. The mapping results were then validated using two breeding populations derived from 'E12076T-03' × 'Colfax' and 'E16099' × 'Colfax'. Marker-assisted resistance spectrum analyses with 9 additional isolates of P. sansomeana indicated that Rpsan1 may be effective towards a broader range of P. sansomeana isolates and has strong merit in protecting soybean to this pathogen in the future.

Extreme Reconfiguration of Plastid Genomes in Papaveraceae: Rearrangements, Gene Loss, Pseudogenization, IR Expansion, and Repeats.

The plastid genomes (plastomes) of angiosperms are typically highly conserved, with extreme reconfiguration being uncommon, although reports of such events have emerged in some lineages. In this study, we conducted a comprehensive comparison of the complete plastomes from twenty-two species, covering seventeen genera from three subfamilies (Fumarioideae, Hypecooideae, and Papaveroideae) of Papaveraceae. Our results revealed a high level of variability in the plastid genome size of Papaveraceae, ranging from 151,864 bp to 219,144 bp in length, which might be triggered by the expansion of the IR region and a large number of repeat sequences. Moreover, we detected numerous large-scale rearrangements, primarily occurring in the plastomes of Fumarioideae and Hypecooideae. Frequent gene loss or pseudogenization were also observed for ndhs, accD, clpP, infA, rpl2, rpl20, rpl32, rps16, and several tRNA genes, particularly in Fumarioideae and Hypecooideae, which might be associated with the structural variation in their plastomes. Furthermore, we found that the plastomes of Fumarioideae exhibited a higher GC content and more repeat sequences than those of Papaveroideae. Our results showed that Papaveroideae generally displayed a relatively conserved plastome, with the exception of Eomecon chionantha, while Fumarioideae and Hypecooideae typically harbored highly reconfigurable plastomes, showing high variability in the genome size, gene content, and gene order. This study provides insights into the plastome evolution of Papaveraceae and may contribute to the development of effective molecular markers.

Phylogeny and diversification of genus Sanicula L. (Apiaceae): novel insights from plastid phylogenomic analyses

BackgroundThe genus Sanicula L. is a unique perennial herb that holds important medicinal values. Although the previous studies on Sanicula provided us with a good research basis, its taxonomic system and interspecific relationships have not been satisfactorily resolved, especially for those endemic to China. Moreover, the evolutionary history of this genus also remains inadequately understood. The plastid genomes possessing highly conserved structure and limited evolutionary rate have proved to be an effective tool for studying plant phylogeny and evolution.ResultsIn the current study, we newly sequenced and assembled fifteen Sanicula complete plastomes. Combined with two previously reported plastomes, we performed comprehensively plastid phylogenomics analyses to gain novel insights into the evolutionary history of this genus. The comparative results indicated that the seventeen plastomes exhibited a high degree of conservation and similarity in terms of their structure, size, GC content, gene order, IR borders, codon bias patterns and SSRs profiles. Such as all of them displayed a typical quadripartite structure, including a large single copy region (LSC: 85,074–86,197 bp), a small single copy region (SSC: 17,047–17,132 bp) separated by a pair of inverted repeat regions (IRs: 26,176–26,334 bp). And the seventeen plastomes had similar IR boundaries and the adjacent genes were identical. The rps19 gene was located at the junction of the LSC/IRa, the IRa/SSC junction region was located between the trnN gene and ndhF gene, the ycf1 gene appeared in the SSC/IRb junction and the IRb/LSC boundary was located between rpl12 gene and trnH gene. Twelve specific mutation hotspots (atpF, cemA, accD, rpl22, rbcL, matK, ycf1, trnH-psbA, ycf4-cemA, rbcL-accD, trnE-trnT and trnG-trnR) were identified that can serve as potential DNA barcodes for species identification within the genus Sanicula. Furthermore, the plastomes data and Internal Transcribed Spacer (ITS) sequences were performed to reconstruct the phylogeny of Sanicula. Although the tree topologies of them were incongruent, both provided strong evidence supporting the monophyly of Saniculoideae and Apioideae. In addition, the sister groups between Saniculoideae and Apioideae were strongly suggested. The Sanicula species involved in this study were clustered into a clade, and the Eryngium species were also clustered together. However, it was clearly observed that the sections of Sanicula involved in the current study were not respectively recovered as monophyletic group. Molecular dating analysis explored that the origin of this genus was occurred during the late Eocene period, approximately 37.84 Ma (95% HPD: 20.33–52.21 Ma) years ago and the diversification of the genus was occurred in early Miocene 18.38 Ma (95% HPD: 10.68–25.28 Ma).ConclusionThe plastome-based tree and ITS-based tree generated incongruences, which may be attributed to the event of hybridization/introgression, incomplete lineage sorting (ILS) and chloroplast capture. Our study highlighted the power of plastome data to significantly improve the phylogenetic supports and resolutions, and to efficiently explore the evolutionary history of this genus. Molecular dating analysis explored that the diversification of the genus occurred in the early Miocene, which was largely influenced by the prevalence of the East Asian monsoon and the uplift of the Hengduan Mountains (HDM). In summary, our study provides novel insights into the plastome evolution, phylogenetic relationships, taxonomic framework and evolution of genus Sanicula.

Complete mitochondrial genome of Syzygium samarangense reveals genomic recombination, gene transfer, and RNA editing events.

Wax apple (Syzygium samarangense) is a commercial fruit that belongs to one of the most species-rich tree genera in the world. We report here the first complete S. samarangense mitogenome obtained using a hybrid assembly strategy. The mitogenome was a 530,242 bp circular molecule encoding 61 unique genes accounting for 7.99% of the full-length genome. Additionally, 167 simple sequence repeats, 19 tandem repeats, and 529 pairs of interspersed repeats were identified. Long read mapping and Sanger sequencing revealed the involvement of two forward repeats (35,843 bp and 22,925 bp) in mediating recombination. Thirteen homologous fragments in the chloroplast genome were identified, accounting for 1.53% of the mitogenome, and the longest fragment was 2,432 bp. An evolutionary analysis showed that S. samarangense underwent multiple genomic reorganization events and lost at least four protein-coding genes (PCGs) (rps2, rps7, rps11, and rps19). A total of 591 RNA editing sites were predicted in 37 PCGs, of which nad1-2, nad4L-2, and rps10-2 led to the gain of new start codons, while atp6-1156, ccmFC-1315 and rps10-331 created new stop codons. This study reveals the genetic features of the S. samarangense mitogenome and provides a scientific basis for further studies of traits with an epistatic basis and for germplasm identification.

Insights into Aquilaria phylogenetics through comparative plastomic resources.

The plastid is an essential organelle for its role in photosynthesis and energy production and its genomic information is always employed as important evolutionary markers to explore the relationship among species. Agarwood (Aquilaria), prized for its aromatic blend, finds extensive use in various cultures as incense and perfume. Despite its high economic importance, the phylogenetic status among Aquilaria based on plastomes remains ambiguous due to the lack of available plastomic resources. To bridge this knowledge gap, 22 Aquilaria plastomes were newly sequenced, similar variation patterns in this genus were determined, including a shared 16 bp extension of the rps19 gene and seven highly variable regions. The analysis highlighted the highest prevalence of the A/T motif among simple sequence repeats in these plastomes. Further phylogenetic analysis revealed Aquilaria's phylogenetic implications with an expanded dataset. This comprehensive plastomic resource not only enhances our understanding of Aquilaria evolution but also presents potential molecular markers for DNA barcoding.

Complete Chloroplast Genomes and the Phylogenetic Analysis of Three Native Species of Paeoniaceae from the Sino-Himalayan Flora Subkingdom.

Paeonia delavayi var. lutea, Paeonia delavayi var. angustiloba, and Paeonia ludlowii are Chinese endemics that belong to the Paeoniaceae family and have vital medicinal and ornamental value. It is often difficult to classify Paeoniaceae plants based on their morphological characteristics, and the limited genomic information has strongly hindered molecular evolution and phylogenetic studies of Paeoniaceae. In this study, we sequenced, assembled, and annotated the chloroplast genomes of P. delavayi var. lutea, P. delavayi var. angustiloba, and P. ludlowii. The chloroplast genomes of these strains were comparatively analyzed, and their phylogenetic relationships and divergence times were inferred. These three chloroplast genomes exhibited a typical quadripartite structure and were 152,687-152,759 bp in length. Each genome contains 126-132 genes, including 81-87 protein-coding genes, 37 transfer RNAs, and 8 ribosomal RNAs. In addition, the genomes had 61-64 SSRs, with mononucleotide repeats being the most abundant. The codon bias patterns of the three species tend to use codons ending in A/U. Six regions of high variability were identified (psbK-psbL, trnG-UCC, petN-psbM, psbC, rps8-rpl14, and ycf1) that can be used as DNA molecular markers for phylogenetic and taxonomic analysis. The Ka/Ks ratio indicates positive selection for the rps18 gene associated with self-replication. The phylogenetic analysis of 99 chloroplast genomes from Saxifragales clarified the phylogenetic relationships of Paeoniaceae and revealed that P. delavayi var. lutea, P. delavayi var. angustiloba, and P. ludlowii are monophyletic groups and sisters to P. delavayi. Divergence time estimation revealed two evolutionary divergences of Paeoniaceae species in the early Oligocene and Miocene. Afterward, they underwent rapid adaptive radiation from the Pliocene to the early Pleistocene when P. delavayi var. lutea, P. delavayi var. angustiloba, and P. ludlowii formed. The results of this study enrich the chloroplast genomic information of Paeoniaceae and reveal new insights into the phylogeny of Paeoniaceae.

In silico prediction and a systematic toxicology-based in vivo investigation uncovering the mechanism of aquatic toxicity caused by beta-lactam antibiotics

Recently, beta-lactam antibiotics have gained attention as significant contributors to public health and environmental issues due to their potential toxicity. Our study employed machine learning to develop a model for assessing the aquatic toxicity of beta-lactam antibiotics on zebrafish. Notably, aztreonam (AZT), a synthetic monobactam and a subclass of beta-lactam antibiotics, demonstrated developmental effects in zebrafish embryos comparable to cephalosporins, indicating a potential for toxicity. Using a systems toxicology-based approach, we identified apoptosis and metabolic disorders as the primary pathways affected by AZT and its impurity F exposure. During the administration of monobactams, we noted that ctsbb, nos2a, and dgat2, genes associated with apoptosis and the metabolic pathway, exhibited significant differential expression. Molecular docking studies were conducted to ascertain the binding affinity between monobactam compounds and their potential targets—Ctsbb, Nos2a, and Dgat2. Furthermore, our research revealed that monobactams influence pre-mRNA alternative splicing, resulting in disruptions in the expression of genes involved in hair cells, brain, spinal cord, and fin regeneration (e.g., krt4, krt5, krt17, cyt1). Notably, we observed a correlation between the levels of rpl3 and rps7 genes, both important ribosomal proteins, and the detected alternative splicing events. Overall, this study enhances our understanding of the toxicity of beta-lactam antibiotics in zebrafish by demonstrating the developmental effects of monobactams and uncovering the underlying mechanisms at the molecular level. It also identifies potential targets for further investigation into the mechanisms of toxicity and provides valuable insights for early assessment of biological toxicity associated with antibiotic pollutants.

A Comparative Kidney Transcriptome Analysis of Bicarbonate-Loaded insrr-Null Mice.

The maintenance of plasma pH is critical for life in all organisms. The kidney plays a critical role in acid-base regulation in vertebrates by controlling the plasma concentration of bicarbonate. The receptor tyrosine kinase IRR (insulin receptor-related receptor) is expressed in renal β-intercalated cells and is involved in alkali sensing due to its ability to autophosphorylate under alkalization of extracellular medium (pH > 7.9). In mice with a knockout of the insrr gene, which encodes for IRR, urinary bicarbonate secretion in response to alkali loading is impaired. The specific regulatory mechanisms in the kidney that are under the control of IRR remain unknown. To address this issue, we analyzed and compared the kidney transcriptomes of wild-type and insrr knockout mice under basal or bicarbonate-loaded conditions. Transcriptomic analyses revealed a differential regulation of a number of genes in the kidney. Using TaqMan real-time PCR, we confirmed different expressions of the slc26a4, rps7, slc5a2, aqp6, plcd1, gapdh, rny3, kcnk5, slc6a6 and atp6v1g3 genes in IRR knockout mice. Also, we found that the expression of the kcnk5 gene is increased in wild-type mice after bicarbonate loading but not in knockout mice. Gene set enrichment analysis between the IRR knockout and wild-type samples identified that insrr knockout causes alterations in expression of genes related mostly to the ATP metabolic and electron transport chain processes.

A Mild DBA Phenotype Associated with a Non-Canonical Splice Site Mutation in RPS26

A 37-years-old Italian female came to the observation of an adult hematology unit for the first time because of an undefined form of congenital anemia. The patient's unrelated parents and her sister were apparently healthy. Hypochromic anemia was observed for the first time at the age of 11 months and treated with one blood transfusion. Oral iron supplementation was started but was ineffective (Hb around to 7 g/dL). At that time, Fanconi's anemia was excluded by cytogenetic analysis, as also DBA The patient showed normal growth and somatopsychic development. Hematology showed moderate macrocytic anemia (MCV ranging from 115-121 fl, Hb levels ranging from 7 to 9 g/dL) with anisocytosis and hypochromia, inadequate reticulocytosis, normal bilirubin and haptoglobin, increased serum ferritin, increased HbF (7-8%), slightly increased erythropoietin, but proportional to anemia, and erythroblastic hyperplasia with variable dyserythropoiesis on bone marrow aspiration and bone marrow biopsy. From the age of 20 onwards, Hb levels ranged between 6.8 and 7.5 g/dL, without transfusion support and allowing a normal working activity. The patient underwent cycles of steroid treatment with moderate benefit. Abdomen ultrasound analysis showed mild steatosis with presence of gallstones, and no signs of focal lesions. At the age of 38 years a first spontaneous pregnancy resulted in intrauterine loss of a female fetus at 21 weeks of gestation because of umbilical cord strangulation. During pregnancy worsening of anemia (Hb 5.8 g/dL) required four blood transfusions. A second spontaneous pregnancy resulted in embryonic loss. Subsequently, she was subjected to several IVF attempts: at 47 years a pregnancy resulted in embryonic loss at 8 weeks of gestation, whereas at 49 years a further pregnancy resulted in a healthy male now 9 years old. Hb levels around 8g/dL were maintained by 20 transfusions, starting at 4 months of gestation. The patient is now 59 and has not developed any tumors, so far. To ascertain the nature of anemia, the patient underwent several investigations: RBC morphology showed anysopoikilocytosis with 10% elliptocytes and 6% spherocytes. Osmotic fragility tests, Eosin-5' maleimide (EMA)-binding test, unstable hemoglobins, SDS-PAGE electrophoresis of red cell membrane proteins as well as the assay of RBC metabolism were normal, except for a decrease of pyruvate kinase/hexokinase (PK/HK) ratio (as sometime happens in presence of dyserythropoiesis). Targeted-NGS sequencing using a panel of 48 genes associated with congenital hemolytic anemias, including RBC membrane defects, RBC metabolism and congenital dyserythropoietic anemias was performed without finding any pathogenic variant. Clinical Exome Sequencing analysis showed the presence of a heterozygous mutation in RPS26 gene: c.3+5G>C. Reverse transcription PCR on total RNA extracted from the patient's PBMC showed that the variant is associated with activation of a cryptic splice site. The aberrant transcript, which is not present in controls, lacks the last 18bp of exon 2 including the ATG (hg19). The fact that the RPS26: c. 3+5G>C mutation does not alter the canonical GT suggests that the mutation may be leaky. This would explain the patient's mild phenotype. A minigene assay is needed to ascertain this hypothesis. In conclusion, if not identified in infancy, milder forms of DBA may be under- or misdiagnosed, further underlying the role of NGS in diagnosis of atypical inherited anemias. The present case further supports the hypothesis that mutations in RPS26 may be associated to a lower cancer risk in DBA.

A rapid molecular diagnostic tool to discriminate alleles of avirulence genes and haplotypes of Phytophthora sojae using high-resolution melting analysis.

Effectors encoded by avirulence genes (Avr) interact with the Phytophthora sojae resistance gene (Rps) products to generate incompatible interactions. The virulence profile of P. sojae is rapidly evolving as a result of the large-scale deployment of Rps genes in soybean. For a successful exploitation of Rps genes, it is recommended that soybean growers use cultivars containing the Rps genes corresponding to Avr genes present in P. sojae populations present in their fields. Determination of the virulence profile of P. sojae isolates is critical for the selection of soybean cultivars. High-resolution melting curve (HRM) analysis is a powerful tool, first applied in medicine, for detecting mutations with potential applications in different biological fields. Here, we report the development of an HRM protocol, as an original approach to discriminate effectors, to differentiate P. sojae haplotypes for six Avr genes. An HRM assay was performed on 24 P. sojae isolates with different haplotypes collected from soybean fields across Canada. The results clearly confirmed that the HRM assay discriminated different virulence genotypes. Moreover, the HRM assay was able to differentiate multiple haplotypes representing small allelic variations. HRM-based prediction was validated by phenotyping assays. This HRM assay provides a unique, cost-effective and efficient tool to predict virulence pathotypes associated with six different Avr (1b, 1c, 1d, 1k, 3a and 6) genes from P. sojae, which can be applied in the deployment of appropriate Rps genes in soybean fields.

Soybean Variety Saedanbaek Confers a New Resistance Allele to Phytophthora sojae

Phytophthora root and stem rot (PRSR) disease results in substantial losses in soybean production worldwide. The occurrence of PRSR caused by Phytophthora sojae Kaufmann & Gerdemann has become increasingly important for soybean production in the Republic of Korea, but domestic soybean–P. sojae interaction has been less studied. The disease has been managed by developing varieties harboring resistance to the Phytophthora sojae (Rps) gene. The present study aimed to identify a major gene locus conferring resistance to new P. sojae isolate 2858 in the recombinant inbred line population derived from a cross between parental lines ‘Daepung’ (susceptible) and ‘Saedanbaek’ (resistant). Seventy-three recombination inbred lines (RILs) were evaluated for resistance to P. sojae isolate 2858. A resistance locus was identified in the approximate 3.3–4.3 megabase pair region on chromosome 3 using both single-marker and linkage analyses. The Rps of Saedanbaek (RpsSDB) was located on the well-known Rps gene/allele cluster region, which also partially overlapped with a locus previously identified in the Korean soybean variety, ‘Daewon’, resistant to another P. sojae isolate 2457 (RpsDW). Approximately 402 kilobase pairs of the interval region overlapped, including six nucleotide-binding site-leucine-rich repeat (NBS-LRR)-coding genes. Additional phenotypic assays revealed that Saedanbaek was susceptible to isolate 2457 and that Daewon was susceptible to isolate 2858, indicating that RpsSDB and RpsDW are different genes or alleles that confer race-specific resistance to the two P. sojae isolates. These results provide information that will be helpful for breeders developing P. sojae-resistant cultivars.