RAD51C-RAD51D interplays with MSH5 and regulates crossover maturation in rice meiosis.

Meiotic crossovers ensure accurate chromosome segregation and increase genetic diversity. RAD51C and RAD51D play an early role in facilitating RAD51 during homologous recombination. However, their later function in meiosis is largely unknown in plants. Here, through targeted disruption of RAD51C and RAD51D, we generated three new mutants and revealed their later meiotic role in crossover maturation. The rad51c-3 and rad51d-4 mutants showed a mixture of bivalents and univalents and no chromosomal entanglements, whereas rad51d-5 exhibited an intermediate phenotype with reduced chromosomal entanglements and increased bivalent formation compared with knockout alleles. Comparisons of RAD51 loadings and chromosomal entanglements in these single mutants, rad51c-3 rad51d-4, rad51c-3 dmc1a dmc1b, and rad51d-4 dmc1a dmc1b suggest that the retained level of RAD51 in mutants is required for uncovering their function in crossover formation. Reductions in chiasma frequency and later HEI10 foci in these mutants support that crossover maturation requires RAD51C and RAD51D. Moreover, interaction between RAD51D and MSH5 indicates that RAD51 paralogs may cooperate with MSH5 to ensure accurate Holliday junction processing into crossover products. This finding of the role of RAD51 paralogs in crossover control may be conserved from mammals to plants and advances our current understanding of these proteins.

MUS81 is required for atypical recombination intermediate resolution but not crossover designation in rice.

The endonuclease methyl methanesulfonate and UV-sensitive protein 81 (MUS81) has been reported to participate in DNA repair during mitosis and meiosis. However, the exact meiotic function of MUS81 in rice remains unclear. Here, we use a combination of physiological, cytological, and genetic approaches to provide evidence that MUS81 functions in atypical recombination intermediate resolution rather than crossover designation in rice. Cytological and genetic analysis revealed that the total chiasma numbers in mus81 mutants were indistinguishable from wild-type. The numbers of HEI10 foci (the sites of interference-sensitive crossovers) in mus81 were also similar to that of wild-type. Moreover, disruption of MUS81 in msh5 or msh4 msh5 background did not further decrease chiasmata frequency, suggesting that rice MUS81 did not function in crossover designation. Mutation of FANCM and ZEP1 could enhance recombination frequency. Unexpectedly, chromosome fragments and bridges were frequently observed in mus81 zep1 and mus81 fancm, illustrating that MUS81 may resolve atypical recombination intermediates. Taken together, our data suggest that MUS81 contributes little to crossover designation but plays a crucial role in the resolution of atypical meiotic intermediates by working together with other anti-crossover factors.

Microfluidic Investigation of the Ion-Specific Effect on Bubble Coalescence in Salt Solutions.

A microfluidic method was developed to study the ion-specific effect on bubble coalescence in salt solutions. Compared with other reported methods, microfluidics provides a more direct and accurate means of measuring bubble coalescence in salt solutions. We analyzed the coalescence time and approach velocity between bubbles and used simulation to investigate the pressure evolution during the coalescence process. The coalescence time of the three salt solutions decreased initially and then increased as the concentration of the salt solution was increased. The concentration with the shortest coalescence time is considered as the transition concentration (TC) and exhibits ion-specific. At the TC, the change in coalescence time indicates a shift in the effect of salt on bubble coalescence from facilitation to initial inhibition. Meanwhile, it can be seen that the sodium halide solutions significantly inhibit the bubble coalescence and the inhibition capability follows the order NaCl > NaBr > NaI. The results of the approach velocity show that the coalescence time decreases with increasing approach velocity, as well as the approach velocity was strongly influenced by concentration. The approach velocity undergoes a significant change at the TC. Furthermore, simulations of bubble coalescence in the microchannel indicate that the vertical pressure gradient at the center point of the bubble pairs increases as bubbles approach, driving liquid film drainage until bubble coalescence. The pressure at the center of the bubble pair reaches the maximum when the bubbles have first coalesced. It was further revealed that the concentration of the salt solution has a significant impact on the maximum pressure, as evidenced by the observed trend of decreasing pressure values with increasing concentrations.

Rapid detection of goose astrovirus genotypes 2 using real-time reverse transcription recombinase polymerase amplification.

BACKGROUND: Goose astrovirus (GoAstV) is an important pathogen that causes joint and visceral gout in goslings. It has been circulating in many provinces of China since 2017. Goose astrovirus genotypes 2 (GoAstV-2) is the main epidemic strain, and its high morbidity and mortality have caused huge economic losses to the goose industry. An accurate point-of-care detection for GoAstV-2 is of great significance. In this study, we developed a real-time reverse transcription recombinase polymerase amplification (RT-RPA) method for the on-site detection of GoAstV-2 infection. RESULTS: The real-time RT-RPA reaction was carried out at a constant temperature of 39 °C, and the entire detection time from nucleic acid preparation to the end of amplification was only 25 min using the portable device. The results of a specificity analysis showed that no cross-reaction was observed with other related pathogens. The detection limit of the assay was 100 RNA copies/µL. The low coefficient of variation value indicated excellent repeatability. We used 270 clinical samples to evaluate the performance of our established method, the positive concordance rates with RT-qPCR were 99.6%, and the linear regression analysis revealed a strong correlation. CONCLUSIONS: The established real-time RT-RPA assay showed high rapidity, specificity and sensitivity, which can be widely applied in the laboratory, field and especially in the resource-limited settings for GoAstV-2 point-of-care diagnosis.

Transcriptome Screening of Long Noncoding RNAs and Their Target Protein-Coding Genes Unmasks a Dynamic Portrait of Seed Coat Coloration Associated with Anthocyanins in Tibetan Hulless Barley.

Many plants have the capability to accumulate anthocyanins for coloration, and anthocyanins are advantageous to human health. In the case of hulless barley (Hordeum vulgare L. var. nudum), investigation into the mechanism of anthocyanin formation is limited to the level of protein-coding genes (PCGs). Here, we conducted a comprehensive bioinformatics analysis to identify a total of 9414 long noncoding RNAs (lncRNAs) in the seed coats of purple and white hulless barley along a developmental gradient. Transcriptome-wide profiles of lncRNAs documented several properties, including GC content fluctuation, uneven length, a diverse range of exon numbers, and a wide variety of transcript classifications. We found that certain lncRNAs in hulless barley possess detectable sequence conservation with Hordeum vulgare and other monocots. Furthermore, both differentially expressed lncRNAs (DElncRNAs) and PCGs (DEPCGs) were concentrated in the later seed development stages. On the one hand, DElncRNAs could potentially cis-regulate DEPCGs associated with multiple metabolic pathways, including flavonoid and anthocyanin biosynthesis in the late milk and soft dough stages. On the other hand, there was an opportunity for trans-regulated lncRNAs in the color-forming module to affect seed coat color by upregulating PCGs in the anthocyanin pathway. In addition, the interweaving of hulless barley lncRNAs and diverse TFs may function in seed coat coloration. Notably, we depicted a dynamic portrait of the anthocyanin synthesis pathway containing hulless barley lncRNAs. Therefore, this work provides valuable gene resources and more insights into the molecular mechanisms underlying anthocyanin accumulation in hulless barley from the perspective of lncRNAs, which facilitate the development of molecular design breeding in crops.

Replication protein A large subunit (RPA1a) limits chiasma formation during rice meiosis.

Replication protein A (RPA), a single-stranded DNA-binding protein, plays essential role in homologous recombination. However, because deletion of RPA causes embryonic lethality in mammals, the exact function of RPA in meiosis remains unclear. In this study, we generated an rpa1a mutant using CRISPR/Cas9 technology and explored its function in rice (Oryza sativa) meiosis. In rpa1a, 12 bivalents were formed at metaphase I, just like in wild-type, but chromosome fragmentations were consistently observed at anaphase I. Fluorescence in situ hybridization assays indicated that these fragmentations were due to the failure of the recombination intermediates to resolve. Importantly, the mutant had a highly elevated chiasma number, and loss of RPA1a could completely restore the 12 bivalent formations in the zmm (for ZIP1-4, MSH4/5, and MER3) mutant background. Protein-protein interaction assays showed that RPA1a formed a complex with the methyl methansulfonate and UV sensitive 81 (and the Fanconi anemia complementation group M-Bloom syndrome protein homologs (RECQ4A)-Topoisomerase3α-RecQ-mediated genome instability 1 complex to regulate chiasma formation and processing of the recombination intermediates. Thus, our data establish a pivotal role for RPA1a in promoting the accurate resolution of recombination intermediates and in limiting redundant chiasma formation during rice meiosis.

Influence of Air Solubility on the Flotation Performance of Low-Rank Coal.

Low-rank coal (LRC) contains large amounts of harmful impurities that must be processed before utilization. Flotation is an effective means for separating fine particles, which can be influenced by air solubility in water. In this work, deaerated water (DW), ordinary water (OW), and pressurized water (PW) were prepared to research the underlying mechanism of the effect of air solubility on the flotation performance of LRC. The results show that PW dissolves the greatest amount of air in the three kinds of water (DW, OW, and PW). The flotation performance of LRC in different water types is directly proportional to air solubility in aqueous solutions. In addition, the induction time of LRC in PW (600 ms) is significantly shorter than those in OW (1200 ms) and DW (4000 ms). Atomic force microscopy (AFM) studies reveal that typical interfacial nanobubbles (NBs) only form on a highly oriented pyrolytic graphite (HOPG) surface in PW due to the supersaturated air in water. Furthermore, the interaction between LRC particles and HOPG in PW is significantly stronger than those in both OW and DW, which is attributed to the capillary force of rgw nanobubble bridge formed between particles. The hydrophobic interaction enhanced by NBs is critically important for the attachment of LRC particles to macrobubbles in flotation. Overall, air solubility in water has a great effect on LRC flotation performance, and PW flotation technology can be extended to LRC purification.

Applying resonance Rayleigh scattering and spectrofluorimetric techniques for the selective determination of neomycin sulfate using Congo red as a probe: Applications for wastewater analysis.

Two simple, sensitive, and low-cost fluorescence spectroscopy methods for neomycin (NEO) detection were developed. Both methods were based on the interaction between NEO and Congo red (CR) in acidic buffer medium to form an ion-association complex. The quenching effect of the formed ion-association complex on the fluorescence of CR at 421 nm is a basic principle of fluorescence analysis, whilst the resonance Rayleigh scattering (RRS) method was used to enhance the resonance Rayleigh scattering spectrum at 384 nm by adding NEO. Experimental conditions such as pH, temperature, reaction time, CR concentration, and the ionic strength of the two methods were investigated and optimized. In addition, the effect of common coexisting substances on the method was tested and the results showed good selectivity. The composition of ion-association complexes, the reaction mechanism, and reasons for the enhanced intensity of RRS are discussed. Under optimum conditions, the responses of the fluorescence spectrophotometry and RRS methods showed good linearity with NEO concentrations in the range 0.2-3.0 µg ml-1 and 0.1-3.0 µg ml-1 , respectively. The detection limits of fluorescence spectrophotometry and RRS spectroscopy techniques were 0.02 µg ml-1 and 0.01 µg ml-1 , respectively. Finally, the two methods were applied to the analysis of wastewater and the results were satisfactory.

Divergent Symptoms Caused by Geminivirus-Encoded C4 Proteins Correlate with Their Ability To Bind NbSKη.

Geminiviruses induce severe developmental abnormalities in plants. The C4/AC4 protein encoded by geminiviruses, especially those not associated with betasatellites, functions as a symptom determinant by hijacking a shaggy-related protein kinase (SKη) and interfering with its functions. Here, we report that the symptom determinant capabilities of C4 proteins encoded by different geminiviruses are divergent and tightly correlated with their abilities to interact with SKη from Nicotiana benthamiana (NbSKη). Swap of the minidomain of tomato leaf curl Yunnan virus (TLCYnV) C4 critical for the interaction with NbSKη increases the capacities of the C4 proteins encoded by tomato yellow leaf curl China virus (TYLCCNV) or tobacco curly shoot virus (TbCSV) to induce symptoms. The severity of symptoms induced by recombinant TYLCCNV C4 or TbCSV C4 correlates with the amount of NbSKη tethered to the plasma membrane by the viral protein. Moreover, a recombinant TYLCCNV harboring the minidomain of TLCYnV C4 induces more-severe symptoms than wild-type TYLCCNV. Thus, this study provides new insights into the mechanism by which different geminivirus-encoded C4 proteins possess divergent symptom determinant capabilities.IMPORTANCE Geminiviruses constitute the largest group of known plant viruses and cause devastating diseases in many economically important crops worldwide. Geminivirus-encoded C4 protein is a multifunctional protein. In this study, we found that the C4 proteins from different geminiviruses showed differential abilities to interact with NbSKη, which correlated with their symptom determinant capabilities. Moreover, a minidomain of tomato leaf curl Yunnan virus (TLCYnV) C4 that is indispensable for interacting with NbSKη and tethering it to the plasma membrane, thus leading to symptom induction, was determined. Supporting these findings, a recombinant geminivirus carrying the minidomain of TLCYnV C4 induced more-severe symptoms than the wild type. Therefore, these findings expand the scope of the interaction of NbSKη and C4-mediated symptom induction and thus contribute to further understanding of the multiple roles of C4.

PRD1, a homologous recombination initiation factor, is involved in spindle assembly in rice meiosis.

The bipolar spindle structure in meiosis is essential for faithful chromosome segregation. PUTATIVE RECOMBINATION INITIATION DEFECT 1 (PRD1) previously has been shown to participate in the formation of DNA double strand breaks (DSBs). However, the role of PRD1 in meiotic spindle assembly has not been elucidated. Here, we reveal by both genetic analysis and immunostaining technology that PRD1 is involved in spindle assembly in rice (Oryza sativa) meiosis. We show that DSB formation and bipolar spindle assembly are disturbed in prd1 meiocytes. PRD1 signals display a dynamic pattern of localization from covering entire chromosomes at leptotene to congregating at the centromere region after leptotene. Centromeric localization of PRD1 signals depends on the organization of leptotene chromosomes, but not on DSB formation and axis establishment. PRD1 exhibits interaction and co-localization with several kinetochore components. We also find that bi-orientation of sister kinetochores within a univalent induced by mutation of REC8 can restore bipolarity in prd1. Furthermore, PRD1 directly interacts with REC8 and SGO1, suggesting that PRD1 may play a role in regulating the orientation of sister kinetochores. Taken together, we speculate that PRD1 promotes bipolar spindle assembly, presumably by modulating the orientation of sister kinetochores in rice meiosis.

OsATM Safeguards Accurate Repair of Meiotic Double-Strand Breaks in Rice.

ATAXIA TELANGIECTASIA-MUTATED (ATM) protein has been well studied for its roles in the DNA damage response. However, its role in meiosis has not been fully explored. Here, we characterized the functions of the rice (Oryza sativa) ATM homolog during meiosis. Aberrant chromosome associations and DNA fragmentations were observed after the completion of homologous pairing and synapsis in Osatm pollen mother cells (PMCs). Aberrant chromosome associations disappeared in Osspo11-1 Osatm-1 double mutants and more severe defects were observed in Osdmc1 Osatm, suggesting that OsATM functions downstream of OsSPO11-1-catalyzed double-strand break formation and in parallel with OsDMC1-mediated homologous recombination. We further demonstrated that phosphorylation of H2AX in PMCs did not depend on OsATM, in contrast to the situation in somatic cells. Moreover, the removal of OsDMC1 from chromosomes in Osatm PMCs was delayed and the number of HEI10 foci (markers of interference-sensitive crossover intermediates) decreased. Together, these findings suggest that OsATM plays important roles in the accurate repair of meiotic double-strand breaks in rice.

The SUN Domain Proteins OsSUN1 and OsSUN2 Play Critical but Partially Redundant Roles in Meiosis.

During meiosis, Sad1/UNC-84 (SUN) domain proteins play conserved roles in promoting telomere bouquet formation and homologous pairing across species. Arabidopsis (Arabidopsis thaliana) AtSUN1 and AtSUN2 have been shown to have overlapping functions in meiosis. However, the role of SUN proteins in rice (Oryza sativa) meiosis and the extent of functional redundancy between them remain elusive. Here, we generated single and double mutants of OsSUN1 and OsSUN2 in rice using genome editing. The Ossun1 Ossun2 double mutant showed severe defects in telomere clustering, homologous pairing, and crossover formation, suggesting that OsSUN1 and OsSUN2 are essential for rice meiosis. When introducing a mutant allele of O. sativa SPORULATION11-1 (OsSPO11-1), which encodes a topoisomerase initiating homologous recombination, into the Ossun1 Ossun2 mutant, we observed a combined Osspo11-1- and Ossun1 Ossun2-like phenotype, demonstrating that OsSUN1 and OsSUN2 promote bouquet formation independent of OsSPO11-1 but regulate pairing and crossover formation downstream of OsSPO11-1. Importantly, the Ossun1 single mutant had a normal phenotype, but meiosis was disrupted in the Ossun2 mutant, indicating that OsSUN1 and OsSUN2 are not completely redundant in rice. Further analyses revealed a genetic dosage-dependent effect and an evolutionary differentiation between OsSUN1 and OsSUN2 These results suggested that OsSUN2 plays a more critical role than OsSUN1 in rice meiosis. Taken together, this work reveals the essential but partially redundant roles of OsSUN1 and OsSUN2 in rice meiosis and demonstrates that functional divergence of SUN proteins has taken place during evolution.

HEIP1 regulates crossover formation during meiosis in rice.

During meiosis, the number of double-strand breaks (DSBs) far exceeds the final number of crossovers (COs). Therefore, to identify proteins involved in determining which of these DSBs repaired into COs is critical in understanding the mechanism of CO control. Across species, HEI10-related proteins play important roles in CO formation. Here, through screening for HEI10-interacting proteins via a yeast two-hybrid system, we identify a CO protein HEI10 Interaction Protein 1 (HEIP1) in rice. HEIP1 colocalizes with HEI10 in a dynamic fashion along the meiotic chromosomes and specially localizes onto crossover sites from late pachytene to diplotene. Between these two proteins, HEI10 is required for the loading of HEIP1, but not vice versa. Moreover, mutations of the HEIP1 gene cause the severe reduction of chiasma frequency, whereas early homologous recombination processes are not disturbed and synapsis proceeds normally. HEIP1 interacts directly with ZIP4 and MSH5. In addition, the loading of HEIP1 depends on ZIP4, but not on MER3, MSH4, or MSH5. Together, our results suggest that HEIP1 may be a member of the ZMM group and acts as a key element regulating CO formation.

PLEKHA5 regulates tumor growth in metastatic melanoma.

BACKGROUND: PLEKHA5 has previously been identified as a novel molecule implicated in melanoma brain metastasis, a disease that continues to portend a poor prognosis. The aim of this study was to further investigate the functional role of PLEKHA5 in disseminated melanoma. METHODS: The impact of PLEKHA5 on proliferation and tumor growth was examined in vitro and in melanoma xenograft models, including brain-tropic melanomas (melanomas tending to disseminate to the brain). In vitro loss- and gain-of-function studies were used to explore the underlying mechanisms of PLEKHA5-mediated tumor growth and the crosstalk between PLEKHA5 and PI3K/AKT/mTOR or MAPK/ERK signaling. The clinical relevance of PLEKHA5 dysregulation was further investigated in a cohort of matched cranial and extracranial melanoma metastases. RESULTS: PLEKHA5 stable knockdown negatively regulated cell proliferation by inhibiting the G1 -to-S cell cycle transition, which coincided with upregulation of the cell cycle regulator PDCD4. Conversely, ectopic PLEKHA5 expression exhibited the inverse effect. PLEKHA5 knockdown significantly inhibited tumor growth, whereas its overexpression upregulated the growth of tumors, which was induced by cranial and subcutaneous inoculation of cells in nude mice. PLEKHA5 modulation affected PDCD4 protein stability and was coupled with changes in PI3K/AKT/mTOR pathway signaling. High PDCD4 expression in cerebral specimens was associated with better overall survival. CONCLUSIONS: This study further supports the role of PLEKHA5 as a regulator of melanoma growth at distant sites, including the brain. Furthermore, the results highlight the significance of PDCD4 dysregulation in disseminated melanoma and implicate PDCD4 as a possible causal link between PLEKHA5 and cell proliferation and growth.

OsRAD51D promotes homologous pairing and recombination by preventing nonhomologous interactions in rice meiosis.

Homologous recombination is carefully orchestrated to maintain genome integrity. RAD51D has been previously shown to be essential for double-strand break repair in mammalian somatic cells. However, the function of RAD51D during meiosis is largely unknown. Here, through detailed analyses of Osrad51d single and double mutants, we pinpoint the specific function of OsRAD51D in coordinating homologous pairing and recombination by preventing nonhomologous interactions during meiosis. OsRAD51D is associated with telomeres in both meiocytes and somatic cells. Loss of OsRAD51D leads to significant induction of nonhomologous pairing and chromosome entanglements, suggesting its role in suppressing nonhomologous interactions. The failed localization of OsRAD51 and OsDMC1 in Osrad51d, together with the genetic analysis of Osrad51d Osdmc1a Osdmc1b, indicates that OsRAD51D acts at a very early stage of homologous recombination. Observations from the Osrad51d pair1 and Osrad51d ku70 double mutants further demonstrate that nonhomologous interactions require double-strand break formation but do not depend on the KU70-mediated repair pathway. Moreover, the interplay between OsRAD51D and OsRAD51C indicates both conservation and divergence of their functions in meiosis. Altogether, this work reveals that OsRAD51D plays an essential role in the inhibition of nonhomologous connections, thus guaranteeing faithful pairing and recombination during meiosis.

The F-Box Protein ZYGO1 Mediates Bouquet Formation to Promote Homologous Pairing, Synapsis, and Recombination in Rice Meiosis.

Telomere bouquet formation, a highly conserved meiotic event, plays an important role in homologous pairing and therefore progression of meiosis; however, the underlying molecular mechanism remains largely unknown. Here, we identified ZYGOTENE1 (ZYGO1), a novel F-box protein in rice (Oryza sativa), and verified its essential role in bouquet formation during early meiosis. In zygo1 mutants, zygotene chromosome aggregation and telomere clustering failed to occur. The suppressed telomere clustering in homologous pairing aberration in rice meiosis1 (pair1) zygo1 and rice completion of meiotic recombination (Oscom1) zygo1 double mutants, together with the altered localization of OsSAD1 (a SUN protein associated with the nuclear envelope) in zygo1, showed that ZYGO1 has a significant function in bouquet formation. In addition, the interaction between ZYGO1 and rice SKP1-like protein 1 suggested that ZYGO1 might modulate bouquet formation as a component of the SKP1-Cullin1-F-box complex. Although double-strand break formation and early recombination element installation occurred normally, zygo1 mutants showed defects in full-length pairing and synaptonemal complex assembly. Furthermore, crossover (CO) formation was disturbed, and foci of Human enhancer of invasion 10 were restricted to the partially synapsed chromosome regions, indicating that CO reduction might be caused by the failure of full-length chromosome alignment in zygo1 Therefore, we propose that ZYGO1 mediates bouquet formation to efficiently promote homolog pairing, synapsis, and CO formation in rice meiosis.

OsHOP2 regulates the maturation of crossovers by promoting homologous pairing and synapsis in rice meiosis.

Meiotic recombination is closely linked with homologous pairing and synapsis. Previous studies have shown that HOMOLOGOUS PAIRING PROTEIN2 (HOP2), plays an essential role in homologous pairing and synapsis. However, the mechanism by which HOP2 regulates crossover (CO) formation has not been elucidated. Here, we show that OsHOP2 mediates the maturation of COs by promoting homologous pairing and synapsis in rice (Oryza sativa) meiosis. We used a combination of genetic analysis, immunolocalization and super-resolution imaging to analyze the function of OsHOP2 in rice meiosis. We showed that full-length pairing, synapsis and CO formation are disturbed in Oshop2 meiocytes. Moreover, structured illumination microscopy showed that OsHOP2 localized to chromatin and displayed considerable co-localization with axial elements (AEs) and central elements (CEs). Importantly, the interaction between OsHOP2 and a transverse filament protein of synaptonemal complex (ZEP1), provided further evidence that OsHOP2 was involved in assembly or stabilization of the structure of the synaptonemal complex (SC). Although the initiation of recombination and CO designation occur normally in Oshop2 mutants, mature COs were severely reduced, and human enhancer of invasion 10 (HEI10)10 foci were only present on the synapsed region. Putting the data together, we speculate that OsHOP2 may serve as a global regulator to coordinate homologous pairing, synapsis and meiotic recombination in rice meiosis.

Detection and genetic characterization of porcine pegivirus from pigs in China.

Porcine pegiviruses (PPgV) have been first discovered in serum samples from domestic pigs in Germany in 2016 and then in the USA in 2018. To date, there is no documentation with respect to the presence of PPgVs in domestic pigs in China. Herein, we attempted to determine the presence and prevalence of PPgV in China and its genetic characterization. In this study, 469 sera were tested and 34 (7.25%) were positive for PPgV. An ascending trend of the positive rate for PPgV was observed from suckling piglets (1.61%) to nursing piglets (1.85%), finishing pigs (6.56%), and sows (11.34%). The complete genome sequence of a representative strain of PPgV, PPgV_GDCH2017, and the complete E2 gene of 17 PPgV isolates discovered in this study was determined. Sequence analysis indicated that PPgV_GDCH2017 was highly related to other PPgVs with nucleotide and amino acid identities ranging from 87.3 to 97.4% and 94.6-99.3%, respectively, in the complete coding region. Phylogenetic analyses demonstrated that the PPgV_GDCH2017 discovered in this study was closely related to the PPgVs from the USA and clustered in the same genus with pegiviruses from other hosts. The topology of the phylogenetic tree based on the complete E2 gene was consistent with that based on the complete genome of PPgV. Further studies on pathogenicity and pathogenesis of PPgVs are needed.

Prevalence and phylogenetic analysis of porcine diarrhea associated viruses in southern China from 2012 to 2018.

BACKGROUND: In China, large-scale outbreaks of severe diarrhea caused by viruses have occurred in pigs since late 2010. To investigate the prevalence and genetic evolution of diarrhea-associated viruses responsible for the outbreaks, a total of 2987 field diarrheal samples collected from 168 pig farms in five provinces in Southern China during 2012-2018 were tested. RESULTS: Porcine epidemic diarrhea virus (PEDV) was most frequently detected virus with prevalence rates between 50.21 and 62.10% in samples, and 96.43% (162/168) in premises, respectively. Porcine deltacoronavirus (PDCoV) was the second prevalent virus with prevalence rates ranging from 19.62 to 29.19% in samples, and 70.24% (118/168) in premises, respectively. Both transmissible gastroenteritis virus (TGEV) and porcine rotavirus (PoRV) were detected at low prevalence rates of < 3% in samples and 10.12% in premises. In this study, we identified a newly emerged swine acute diarrhea syndrome coronavirus (SADS-CoV) in diarrheal samples of piglets from Fujian province in Southern China, and the prevalence rate of SADS-CoV was 10.29% (7/68). Co-infections of these diarrhea-associated viruses were common. The most frequent co-infection was PEDV with PDCoV, with an average detection rate of 12.72% (380/2987, ranging from 8.26-17.33%). Phylogenetic analysis revealed that PEDVs circulating in Southern China during the last 7 years were clustered with the variant strains of PEDV in genotype IIa. The most frequent mutations were present in the collagenase equivalent (COE) and epitope regions of the spike gene of the PEDVs currently circulating in the field. Genetic relationships of PDCoVs were closely related with Chinese strains, other than those present in the USA, South Korea, Thailand and Lao's public. CONCLUSIONS: The findings of this study indicated that variant PEDV, PDCoV, and SADS-CoV were leading etiologic agents of porcine diarrhea, and either mono-infections or co-infections of pathogenic enteric CoVs were common in pigs in Southern China during 2012-2018. Thus, significant attention should be paid in order to effectively prevent and control porcine viral diarrhea.

OsSPL regulates meiotic fate acquisition in rice.

In angiosperms, the key step in sexual reproduction is successful acquisition of meiotic fate. However, the molecular mechanism determining meiotic fate remains largely unknown. Here, we report that OsSPOROCYTELESS (OsSPL) is critical for meiotic entry in rice (Oryza sativa). We performed a large-scale genetic screen of rice sterile mutants aimed to identify genes regulating meiotic entry and identified OsSPL using map-based cloning. We showed that meiosis-specific callose deposition, chromatin organization, and centromere-specific histone H3 loading were altered in the cells corresponding to pollen mother cells in Osspl anthers. Global transcriptome analysis showed that the enriched differentially expressed genes in Osspl were mainly related to redox status, meiotic process, and parietal cell development. OsSPL might form homodimers and interact with TEOSINTE BRANCHED1/CYCLOIDEA/PCF (TCP) transcription factor OsTCP5 via the SPL dimerization and TCP interaction domain. OsSPL also interacts with TPL (TOPLESS) corepressors, OsTPL2 and OsTPL3, via the EAR motif. Our results suggest that the OsSPL-mediated signaling pathway plays a crucial role in rice meiotic entry, which appears to be a conserved regulatory mechanism for meiotic fate acquisition in angiosperms.