The complete chloroplast genome of Brassica rapa var. purpuraria (L.H.Bailey) Kitam 1950 and its phylogenetic analysis.

Zicaitai (Brassica rapa var. purpuraria (L.H.Bailey) Kitam 1950) is a vegetable crop that boasts a high nutritional value and unique flavor. It originated from Central China and was formed after long-term cultivation and domestication. In this study, we obtained the complete sequence of the chloroplast genome of zicaitai, a circular molecule of 153,483 bp in length. This chloroplast genome consists of a large single-copy (LSC) region (83,282 bp), a small single-copy (SSC) region (17,775 bp), and a pair of inverted repeats (IRs) (26,213 bp). By sequence annotation, 132 genes, including 87 protein-coding genes, 37 tRNA genes, and eight rRNA genes were identified in the zicaitai chloroplast. A total of 315 simple sequence repeats (SSRs) were found located in LSC (197), SSC (72), and IR (46), respectively. Phylogenetic analysis based on chloroplast genomes indicated the relationship of zicaitai and the Brassicaceae family, which supports zicaitai as a variety of B. rapa in taxonomy. The results obtained in this study provide insight into further research on Brassica chloroplasts and their phylogeny.

Chromosome-level genome assembly and annotation of Zicaitai (Brassica rapa var. purpuraria).

Zicaitai is a seasonal vegetable known for its high anthocyanin content in both stalks and leaves, yet its reference genome has not been published to date. Here, we generated the first chromosome-level genome assembly of Zicaitai using a combination of PacBio long-reads, Illumina short-reads, and Hi-C sequencing techniques. The final genome length is 474.12 Mb with a scaffold N50 length of 43.82 Mb, a BUSCO score of 99.30% and the LAI score of 10.14. Repetitive elements accounted for 60.89% (288.72 Mb) of the genome, and Hi-C data enabled the allocation of 430.87 Mb of genome sequences to ten pseudochromosomes. A total of 42,051 protein-coding genes were successfully predicted using multiple methods, of which 99.74% were functionally annotated. Notably, comparing the genome of Zicaitai with seven other species in the Cruciferae family revealed strong conservation in terms of gene numbers and structures. Overall, the high-quality genome assembly provides a critical resource for studying the genetic basis of important agronomic traits in Zicaitai.

A New Fast Control Strategy of Terminal Sliding Mode with Nonlinear Extended State Observer for Voltage Source Inverter.

To overcome the sensitivity of voltage source inverters (VSIs) to parameter perturbations and their susceptibility to load variations, a fast terminal sliding mode control (FTSMC) method is proposed as the core and combined with an improved nonlinear extended state observer (NLESO) to resist aggregate system perturbations. Firstly, a mathematical model of the dynamics of a single-phase voltage type inverter is constructed using a state-space averaging approach. Secondly, an NLESO is designed to estimate the lumped uncertainty using the saturation properties of hyperbolic tangent functions. Finally, a sliding mode control method with a fast terminal attractor is proposed to improve the dynamic tracking of the system. It is shown that the NLESO guarantees convergence of the estimation error and effectively preserves the initial derivative peak. The FTSMC enables the output voltage with high tracking accuracy and low total harmonic distortion and enhances the anti-disturbance ability.

An automated microfluidic system with one-dimensional beads array for multiplexed torch detection at point-of-care testing.

An automated microfluidic system with functionalized beads has been developed for multiplexed TORCH detection at point-of-care testing. A concise microfluidic chip consisting of a one-dimensional beads array is developed to simultaneously detect TOX, RUB, CMV, HSV-I and HSV-II respectively with five functionalized beads. A compact liquid handling module has been developed to automate the sandwiched chemiluminescence immunoassay within the one-dimensional beads array of the microfluidic chip. A precise ram pump is adopted to not only add reagent into the microfluidic chip from outside, but also facilitate elaborate fluid control inside the microfluidic chip for improved performance. A large-size waste chamber with a liquid-absorbing sponge holds the waste reagent within the microfluidic chip to prevent backflow. The one-dimensional beads array is heated from double-sides at 37 ℃ for sensitive detection with reduced time. A sensitive CMOS camera is adopted to take chemiluminescence image from the one-dimensional beads array, and a custom processing algorithm is adopted to analyze the image. For each serum sample, five different infections can be simultaneously detected with the automated microfluidic system. Experimental results show that efficient, sensitive, and accurate multiplexed TORCH detection can be conveniently achieved with the integrated microfluidic system.

A microfluidic system for rapid nucleic acid analysis based on real-time convective PCR at point-of-care testing.

A microfluidic system for rapid nucleic acid analysis based on real-time convective PCR is developed. To perform 'sample-in, answer-out' nucleic acid analysis, a microfluidic chip is developed to efficiently extract nucleic acid, and meanwhile convective PCR (CPCR) is applied for rapid nucleic acid amplification. With an integrated microfluidic chip consisting of reagent pre-storage chambers, a lysis & wash chamber, an elution chamber and a waste chamber, nucleic acid extraction based on magnetic beads can be automatically performed for a large size of test sample within a limited time. Based on an easy-to-operate strategy, different pre-stored reagents can be conveniently released for consecutive reaction at different steps. To achieve efficient mixing, a portable companion device is developed to introduce properly controlled 3-D actuation to magnetic beads in nucleic acid extraction. In CPCR amplification, PCR reagent can be spontaneously and repeatedly circulated between hot and cool zones of the reactor for space-domain thermal cycling based on pseudo-isothermal heating. A handheld real-time CPCR device is developed to perform nucleic acid amplification and in-situ detection. To extend the detection throughput, multiple handheld real-time CPCR devices can be grouped together by a common control system. It is demonstrated that influenza A (H1N1) viruses with the reasonable concentration down to 1.0 TCID50/ml can be successfully detected with the microfluidic system.

Development and internal validation of a Wasp Sting Severity Score to assess severity and indicate blood purification in persons with Asian wasp stings.

BACKGROUND: In recent years, the incidence of wasp sting has increased annually in China. Organ damage and high mortality due to mass wasp envenomation remain major challenges. Timely and appropriate medical intervention can improve survival. However, there are currently no normalized tools for early assessment of severity. METHODS: The clinical data of wasp sting patients hospitalized from 2011 to 2019 were used as a training set. Logistic regression was used to explore major risk factors for the development of a severe case of wasp sting (SC). The Wasp Sting Severity Score (WSS) was determined considering these risk factors to identify SCs and was tested in a validation dataset that was prospectively collected in 2020. RESULTS: The data of 1131 wasp sting patients from 2011 to 2019 were included in the training set. Logistic regression analysis showed that tea-colored urine, number of stings, and lactate dehydrogenase and total bilirubin levels were risk factors for developing an SC. The WSS was developed considering these four risk factors, and the total possible WSS was 20 points. The WSS was tested using the validation dataset, comprising the data of 153 patients, in 2020, and we found that a WSS ≥3 points was an important indication for blood purification, with a sensitivity of 71.9%, specificity of 92.6% and an area under the curve of 0.918 (95% confidence interval 0.873-0.962). Among patients with more than 30 stings, mortality in those who underwent plasma exchange (PE) within 24 h after admission was significantly lower than that in those who did not receive PE treatment (14.3% versus 46.9%, P = 0.003). However, continuous venovenous hemofiltration (CVVH) (P = 0.317) and hemoperfusion (HP) (P = 0.869) did not significantly reduce mortality. CONCLUSIONS: Patients with WSS scores ≥3 should be considered for blood purification as early as possible in addition to routine treatment. In addition, PE is better than CVVH and HP at reducing mortality in patients suffering from severe wasp stings.

Actively implementing an evidence-based feeding guideline for critically ill patients (NEED): a multicenter, cluster-randomized, controlled trial.

BACKGROUND: Previous cluster-randomized controlled trials evaluating the impact of implementing evidence-based guidelines for nutrition therapy in critical illness do not consistently demonstrate patient benefits. A large-scale, sufficiently powered study is therefore warranted to ascertain the effects of guideline implementation on patient-centered outcomes. METHODS: We conducted a multicenter, cluster-randomized, parallel-controlled trial in intensive care units (ICUs) across China. We developed an evidence-based feeding guideline. ICUs randomly allocated to the guideline group formed a local "intervention team", which actively implemented the guideline using standardized educational materials, a graphical feeding protocol, and live online education outreach meetings conducted by members of the study management committee. ICUs assigned to the control group remained unaware of the guideline content. All ICUs enrolled patients who were expected to stay in the ICU longer than seven days. The primary outcome was all-cause mortality within 28 days of enrollment. RESULTS: Forty-eight ICUs were randomized to the guideline group and 49 to the control group. From March 2018 to July 2019, the guideline ICUs enrolled 1399 patients, and the control ICUs enrolled 1373 patients. Implementation of the guideline resulted in significantly earlier EN initiation (1.20 vs. 1.55 mean days to initiation of EN; difference - 0.40 [95% CI - 0.71 to - 0.09]; P = 0.01) and delayed PN initiation (1.29 vs. 0.80 mean days to start of PN; difference 1.06 [95% CI 0.44 to 1.67]; P = 0.001). There was no significant difference in 28-day mortality (14.2% vs. 15.2%; difference - 1.6% [95% CI - 4.3% to 1.2%]; P = 0.42) between groups. CONCLUSIONS: In this large-scale, multicenter trial, active implementation of an evidence-based feeding guideline reduced the time to commencement of EN and overall PN use but did not translate to a reduction in mortality from critical illness. TRIAL REGISTRATION: ISRCTN, ISRCTN12233792 . Registered November 20th, 2017.

[Case control study on T-plate combined with suture anchors for the treatment of Neer â ¡b clavicle fractures].

OBJECTIVE: To compare the clinical efficacy of distal radius T-plate combined with suture anchor and distal clavicle anatomical locking plate combined with suture anchor in the treatment of Neer Ⅱb distal clavicle fracture. METHODS: From June 2014 to June 2018, 42 patients with Neer Ⅱb distal clavicle fractures were retrospectively analyzed. According to different surgical methods, they were divided into the observation group (T-shaped plate combined with suture anchor) and the control group (anatomical locking plate combined with suture anchor). There were 22 patients in the observation group and 20 patients in the control group. In the observation group, there were 13 males and 9 females, aged from 22 to 70 (45.78± 14.44) years old, 12 cases on the left side and 10 cases on the right side, 8 cases of traffic accident injury and 14 cases of fall. In the control group, there were 12 males and 8 females, aged from 24 to 66 (44.17±15.58) years, 13 cases on the left side and 7 cases on the right side, 6 cases of traffic accident injuryand 14 cases of fall. The operation time, intraoperative blood loss and fracture healing time were compared between the two groups, and Constant Murley score was used to evaluate shoulder joint function. RESULTS: The patients in both groups were followed up for 18 to 24 (20.96±2.02) months. The incisions of both groups were healed at stageⅠ. The fracture ends of both groups were bony healed at the last follow up. There was no significant difference in operation time, intraoperative blood loss and fracture healing time between two groups (P>0.05);there was no significant difference in shoulder joint function between two groups at 3 months after operation (P>0.05). CONCLUSION: The two methods can obtain satisfactory results in the treatment of Neer Ⅱb distal clavicle fractures, especially suitable for patients with comminuted distal clavicle fractures or osteoporosis; the clinical effect of the treatment of NeerⅡb distal clavicle fractures with T type distal radius plate combined with suture anchor is satisfactory, which provides another feasible treatment scheme for clinic.

First Report of Watermelon mosaic virus causing a Mosaic Disease on Cucumis metuliferus in China.

Cucumis metuliferus, also called horned cucumber or jelly melon, is considered as a wild species in the Cucumis genus and a potential material for nematodes- or viruses-resistant breeding (Provvidenti, et al. 1977; Sigüenza et al. 2005; Chen et al. 2020). This species, originating from Africa, has been cultivated as a fruit in China in recent years. In July 2020, a mosaic disease was observed on C. metuliferus growing in five fields (approximately 0.7 hectare) in Urumqi, Xijiang, China, where more than 85~100% of the field plants exhibited moderate to severe viral disease-like leaf mosaic and/or deformation symptoms. Delayed flowering and small and/or deformed fruits on the affected plants could result in yield loss of about 50%. To identify the causal pathogen, the symptomatic leaf samples were collected from the five fields (five plants/points for each field) and their total RNAs were extracted using a commercial RNA extraction kit. The universal potyviral primers (Ha et al. 2008) and specific primers for a number of frequently-occurring, cucurbit crop-infecting viruses including Papaya ringspot virus (PRSV) (Lin et al. 2013), Cucumber mosaic virus (CMV) and Watermelon mosaic virus (WMV) were designed and used for detection by RT-PCR. The result showed that only the WMV primers (forward: 5'-AAGTGTGACCAAGCTTGGACTGCA-3' and reverse: 5'-CTCACCCATTGTGCCAAAGAACGT-3') could amplify the corresponding target fragment from the total RNA templates, and direct sequencing of the RT-PCR products and GenBank BLAST confirmed the presence of WMV (genus Potyvirus) in the collected C. metuliferus samples. To complete Koch's postulates, the infected C. metuliferus leaves were ground in the sodium phosphate buffer (0.01 M, pH 7.0) and the sap was mechanically inoculated onto 30 four-leaf-stage C. metuliferus seedlings (two leaves for each seedling were inoculated) kept in an insect-proof, temperature-controlled greenhouse at 25~28℃. Twenty-five of the inoculated plants were observed to have apparent leaf mosaic similar to the field symptoms two weeks after inoculation, and positive result was obtained in RT-PCR detection for the symptomatic leaves of inoculated plants using the WMV primers aforementioned, confirming the virus as the pathogen of C. metuliferus in Urumqi. To our knowledge, this is the first report of WMV naturally infecting C. metuliferus in China. We obtained the full-length sequence of the WMV Urumqi isolation (WMV-Urumqi) by sequencing the RT-PCR amplicons from seven pairs of primers spanning the viral genome and the 5'RACE and 3'RACE products. The complete sequence of WMV-Urumqi (GenBank accession no. MW345911) is 10046 nucleotides (nt) long and contains an open reading frame that encodes a polyprotein of 3220 amino acids (aa). WMV-Urumqi shares the highest nt identity (95.9%) and aa identity (98.0%) with the Cucurbita pepo-infecting isolation (KX664483) from Shanxi province, China. Our findings provide a better understanding of the host range and genetic diversity of WMV, and a useful reference for virus-resistant breeding involving C. metuliferus.

The wide distribution and horizontal transfers of beta satellite DNA in eukaryotes.

Beta satellite DNA (satDNA), also known as Sau3A sequences, are repetitive DNA sequences reported in human and primate genomes. It is previously thought that beta satDNAs originated in old world monkeys and bursted in great apes. In this study, we searched 7821 genome assemblies of 3767 eukaryotic species and found that beta satDNAs are widely distributed across eukaryotes. The four major branches of eukaryotes, animals, fungi, plants and Harosa/SAR, all have multiple clades containing beta satDNAs. These results were also confirmed by searching whole genome sequencing data (SRA) and PCR assay. Beta satDNA sequences were found in all the primate clades, as well as in Dermoptera and Scandentia, indicating that the beta satDNAs in primates might originate in the common ancestor of Primatomorpha or Euarchonta. In contrast, the widely patchy distribution of beta satDNAs across eukaryotes presents a typical scenario of multiple horizontal transfers.

Enteral nutrition feeding in Chinese intensive care units: a cross-sectional study involving 116 hospitals.

BACKGROUND: There is a lack of large-scale epidemiological data on the clinical practice of enteral nutrition (EN) feeding in China. This study aimed to provide such data on Chinese hospitals and to investigate factors associated with EN delivery. METHODS: This cross-sectional study was launched in 118 intensive care units (ICUs) of 116 mainland hospitals and conducted on April 26, 2017. At 00:00 on April 26, all patients in these ICUs were included. Demographic and clinical variables of patients on April 25 were obtained. The dates of hospitalization, ICU admission and nutrition initiation were reviewed. The outcome status 28 days after the day of investigation was obtained. RESULTS: A total of 1953 patients were included for analysis, including 1483 survivors and 312 nonsurvivors. The median study day was day 7 (IQR 2-19 days) after ICU entry. The proportions of subjects starting EN within 24, 48 and 72 h after ICU entry was 24.8% (84/352), 32.7% (150/459) and 40.0% (200/541), respectively. The proportion of subjects receiving > 80% estimated energy target within 24, 48, 72 h and 7 days after ICU entry was 10.5% (37/352), 10.9% (50/459), 11.8% (64/541) and 17.8% (162/910), respectively. Using acute gastrointestinal injury (AGI) 1 as the reference in a Cox model, patients with AGI 2-3 were associated with reduced likelihood of EN initiation (HR 0.46, 95% CI 0.353-0.599; p < 0.001). AGI 4 was significantly associated with lower hazard of EN administration (HR 0.056; 95% CI 0.008-0.398; p = 0.004). In a linear regression model, greater Sequential Organ Failure Assessment scores (coefficient - 0.002, 95% CI - 0.008 to - 0.001; p = 0.024) and male gender (coefficient - 0.144, 95% CI - 0.203 to - 0.085; p < 0.001) were found to be associated with lower EN proportion. As compared with AGI 1, AGI 2-3 was associated with lower EN proportion (coefficient - 0.206, 95% CI - 0.273 to - 0.139; p < 0.001). CONCLUSIONS: The study showed that EN delivery was suboptimal in Chinese ICUs. More attention should be paid to EN use in the early days after ICU admission.

Characteristics of siRNAs derived from Southern rice black-streaked dwarf virus in infected rice and their potential role in host gene regulation.

BACKGROUND: Virus-derived siRNAs (vsiRNAs)-mediated RNA silencing plays important roles in interaction between plant viruses and their hosts. Southern rice black-streaked dwarf virus (SRBSDV) is a newly emerged devastating rice reovirus with ten dsRNA genomic segments. The characteristics of SRBSDV-derived siRNAs and their biological implications in SRBSDV-rice interaction remain unexplored. METHODS: VsiRNAs profiling from SRBSDV-infected rice samples was done via small RNA deep sequencing. The putative rice targets of abundantly expressed vsiRNAs were bioinformatically predicted and subjected to functional annotation. Differential expression analysis of rice targets and RNA silencing components between infected and healthy samples was done using RT-qPCR. RESULTS: The vsiRNA was barely detectable at 14 days post infection (dpi) but abundantly present along with elevated expression level of the viral genome at 28 dpi. From the 28-dpi sample, 70,878 reads of 18 ~ 30-nt vsiRNAs were recognized (which mostly were 21-nt and 22-nt), covering 75 ~ 91% of the length of the ten genomic segments respectively. 86% of the vsiRNAs had a <50% GC content and 79% of them were 5'-uridylated or adenylated. The production of vsiRNAs had no strand polarity but varied among segment origins. Each segment had a few hotspot regions where vsiRNAs of high abundance were produced. 151 most abundant vsiRNAs were predicted to target 844 rice genes, including several types of host resistance or pathogenesis related genes encoding F-box/LRR proteins, receptor-like protein kinases, universal stress proteins, tobamovirus multiplication proteins, and RNA silencing components OsDCL2a and OsAGO17 respectively, some of which showed down regulation in infected plants in RT-qPCR. GO and KEGG classification showed that a majority of the predicted targets were related to cell parts and cellular processes and involved in carbohydrate metabolism, translation, and signal transduction. The silencing component genes OsDCL2a, OsDCL2b, OsDCL4, and OsAGO18 were down regulated, while OsAGO1d, OsAGO2, OsRDR1 and OsRDR6 were up regulated, significantly, upon SRBSDV infection. CONCLUSIONS: SRBSDV can regulate the expression of rice RNA silencing pathway components and the virus might compromise host defense and influence host pathogenesis via siRNA pathways.

Tolerance and responsive gene expression of Sogatella furcifera under extreme temperature stresses are altered by its vectored plant virus.

Southern rice black-streaked dwarf virus (SRBSDV), a newly emerged fijivirus causing great loss to rice production in eastern and southeastern Asian countries in recent years, is efficiently transmitted by a rice pest, white-backed planthopper (WBPH, Sogatella furcifera) in a persistent, circulative propagative manner and can be considered as an insect virus. In this study, SRBSDV infection in WBPH was found to increase the vector's death rate under extreme cold stress but improve its survival rate under extreme heat stress. Digital gene expression profiling based on RNA-Seq revealed different gene regulation patterns in WBPH under viral and/or temperature stress. Under cold stress, the virus infection upregulated 1540 genes and downregulated 131 genes in the insect, most of which were related to membrane properties and biological processes of actin and cytoskeleton; whereas under heat stress, it upregulated 363 genes and downregulated 548 genes, most of which were associated to metabolism and intracellular organelles. Several types of stress-responsive genes involving intestinal mucin, cuticle protein, ubiquitin protease, immune response, RNA interference and heat shock response, were largely upregulated under cold stress, but largely downregulated under heat stress, by SRBSDV infection. Our results suggest two distinct mechanisms of virus-altered vector insect tolerance to temperature stress.

MicroRNAs responding to southern rice black-streaked dwarf virus infection and their target genes associated with symptom development in rice.

Southern rice black-streaked dwarf virus (SRBSDV) is a recently emerged rice virus that has spread across Asia. This devastating virus causes rice plants to produce a variety of symptoms during different growth stages. MicroRNAs (miRNAs) comprise a large group of 21-24-nt RNA molecules that are important regulators of plant development processes and stress responses. In this study, we used microarray profiling to investigate rice miRNAs responding to SRBSDV infection at 3, 9, 15, and 20 days post-inoculation (dpi). Expression levels of 56 miRNAs were altered in SRBSDV-infected rice plants, with these changes classified into eight different regulation patterns according to their temporal expression dynamics. Fourteen miRNAs belonging to six families (miR164, R396, R530, R1846, R1858, and R2097) were significantly regulated at 20 dpi. We used RT-qPCR to search for expression level correlations between members of these families and their putative targets at 3, 9, and 15 dpi. Some members of the miR164, R396, R530, and R1846 families were found to be positively or negatively correlated with their respective targets during 3-15 days after SRBSDV infection, whereas in more cases the rice miRNAs were not in correlation with their targets along the post-inoculation period, suggesting that some additional factors may be involved in rice miRNA-target interactions. The reported functions of rice genes targeted by the miR164, R396, R530, R1846, and R1858 families indicated that these genes are associated with symptom development. These results provide insights into miRNA-mediated SRBSDV-rice interactions.

Southern rice black-streaked dwarf virus alters insect vectors' host orientation preferences to enhance spread and increase rice ragged stunt virus co-infection.

In recent years, Southern rice black-streaked dwarf virus (SRBSDV), a tentative species in the genus Fijivirus (family Reoviridae), has spread rapidly and caused serious rice losses in eastern and southeastern Asia. With this virus spread, Rice ragged stunt virus (RRSV, genus Oryzavirus, family Reoviridae) became more common in southern China, usually in co-infection with the former. SRBSDV and RRSV are transmitted by two different species of planthoppers, white-backed planthopper (WBPH, Sogatella furcifera) and brown planthopper (BPH, Nilaparvata lugens), respectively, in a persistent, circulative, propagative manner. In this study, using a Y-shape olfactometer-based device, we tested the host preference of three types of macropterous WBPH adults for healthy or SRBSDV-infected rice plants. The results showed that virus-free WBPHs significantly preferred infected rice plants to healthy plants, whereas both the viruliferous and nonviruliferous WBPHs preferred healthy plants to infected plants. In additional tests, we found that the BPHs significantly preferred healthy plants when they were virus free, whereas RRSV-carrying BPHs preferred SRBSDV-infected rice plants. From these findings, we propose that plant viruses may alter host selection preference of vectors to enhance their spread and that of insects vectoring another virus to result in co-infection with more than one virus.

Southern rice black-streaked dwarf virus: a white-backed planthopper-transmitted fijivirus threatening rice production in Asia.

Southern rice black-streaked dwarf virus (SRBSDV), a non-enveloped icosahedral virus with a genome of 10 double-stranded RNA segments, is a novel species in the genus Fijivirus (family Reoviridae) first recognized in 2008. Rice plants infected with this virus exhibit symptoms similar to those caused by Rice black-streaked dwarf virus. Since 2009, the virus has rapidly spread and caused serious rice losses in East and Southeast Asia. Significant progress has been made in recent years in understanding this disease, especially about the functions of the viral genes, rice-virus-insect interactions, and epidemiology and control measures. The virus can be efficiently transmitted by the white-backed planthopper (WBPH, Sogatella furcifera) in a persistent circulative propagative manner but cannot be transmitted by the brown planthopper (Nilaparvata lugens) and small brown planthopper (Laodelphax striatellus). Rice, maize, Chinese sorghum (Coix lacryma-jobi) and other grass weeds can be infected via WBPH. However, only rice plays a major role in the virus infection cycle because of the vector's preference. In Southeast Asia, WBPH is a long-distance migratory rice pest. The disease cycle can be described as follows: SRBSDV and its WBPH vector overwinter in warm tropical or sub-tropical areas; viruliferous WBPH adults carry the virus from south to north via long-distance migration in early spring, transmit the virus to rice seedlings in the newly colonized areas, and lay eggs on the infected seedlings; the next generation of WBPHs propagate on infected seedlings, become viruliferous, disperse, and cause new disease outbreaks. Several molecular and serological methods have been developed to detect SRBSDV in plant tissues and individual insects. Control measures based on protection from WBPH, including seedbed coverage, chemical seed treatments, and chemical spraying of seedlings, have proven effective in China.

Evaluation of three RT-qPCR-based miRNA detection methods using seven rice miRNAs.

Three frequently-used reverse transcription-quantitative polymerase chain reaction (RT-qPCR)-based miRNA detection methods, stem-loop RT-qPCR, poly(A)-tailing RT-qPCR, and miQPCR, were evaluated using seven selected rice miRNAs. The results revealed that miRNA abundance and sequence characteristics can affect capability of detection. The stem-loop amplification technique detected highly and moderately abundant miRNAs. The poly(A)-tailing method detected both highly abundant and sparsely present miRNAs, but failed to detect miRNAs with a hairpin structure. Only a few miRNAs were detectable by the miQPCR method. We suggest that a combination of methods should be used for reliable quantitative investigation of miRNAs.

Effects of southern rice black-streaked dwarf virus on the development and fecundity of its vector, Sogatella furcifera.

BACKGROUND: Southern rice black-streaked dwarf virus (SRBSDV) threatens rice production in China and Vietnam. The virus is vectored by the migrating white-backed planthopper (WBPH, Sogatella furcifera) in a circulative, propagative, and persistent manner. A persistently-transmitted plant virus might affect its vector's development and fecundity directly by infecting the vector itself and/or indirectly altering the host plant. This study evaluated the direct and indirect effects of SRBSDV on WBPH performance to better understand the virus-vector-host plant relationship in terms of its effects on the biological parameters of the vector. METHODS: Three experimental WBPH populations were established. Viruliferous and non-viruliferous populations were fed on SRBSDV-infected rice seedlings for 48 h as first-instar nymphs; infection status was confirmed by RT-PCR after they died. The control population was fed on healthy rice. Each insect was individually transferred to a healthy rice plant grown in a glass tube at 20°C, 25°C, or 28°C. Life parameters, including nymphal duration, survival rate, adult sex ratio, macropterous proportion, longevity, and oviposition amounts, of each population were measured at each temperature. RESULTS: The life parameter data indicated that SRBSDV and infected rice plants adversely influenced WBPH; the effects were temperature dependent. Compared with the control population, viruliferous populations showed significant changes, including prolonged nymphal stages and reduced survival rates at 20°C, while the non-viruliferous population had higher survival rates at 20°C and lower rates at 28°C compared with the control. Both populations had significantly shorter adult life spans at 25°C and lower oviposition amounts at 28°C relative to the control. CONCLUSIONS: Both SRBSDV-infection and feeding on infected rice plants affected vector performance. Although a longer nymphal period benefits viral acquisition and transmission by nymphs and might increase rice infection rate, in general, SRBSDV infection of the vectors and host plants was unfavorable to WBPH population expansion.

Simultaneous detection and differentiation of four closely related sweet potato potyviruses by a multiplex one-step RT-PCR.

Four closely related potyviruses, Sweet potato feathery mottle virus (SPFMV), Sweet potato virus C (SPVC), Sweet potato virus G (SPVG) and/or Sweet potato virus 2 (SPV2), are involved in sweet potato virus disease complexes worldwide. Identification and detection of these viruses are complicated by high similarity among their genomic sequences, frequent occurrence as mixed infections and low titer in many sweet potato cultivars. A one-tube multiplex reverse transcription-PCR (mRT-PCR) assay was developed for simultaneous detection and differentiation of SPFMV, SPVC, SPVG and SPV2. Four specific forward primers unique to each virus and one reverse primer based on the region conserved in all four viruses were selected and used in the assay. The mRT-PCR assay was optimized for primer concentration and cycling conditions. It was tested using sweet potato plants infected naturally with one to four target viruses and then evaluated using field samples collected from southwestern China. The mRT-PCR assay is reliable and sensitive as a simple, rapid and cost-effective method to detect these pathogens in sweet potato. This assay will be useful to quarantine and certification programs and virus surveys when large numbers of samples are tested.