Intramolecular charge transfer enhanced optical limiting in novel hydrazone derivatives with a D1-D-Ai-π-A structure.

The present paper investigates one of the hydrazone derivatives (BTH with a D-π-A structure) based on density functional theory. With the computation results of ground state absorption (GSA), excited-state absorption (ESA) and multi-photon absorption (MPA), the optical limiting effect observed in the experiment for the BTH molecule can be well predicted and elucidated by the MPA-ESA mechanism. The analysis of the hole-electron and the electron density differences between two transition states reveal that the main transitions involved in the GSA and ESA of BTH could be recognized as local excitation. Based on these observations, four novel hydrazone derivatives based on the BTH unit with a D1-D-Ai-π-A structure were designed to promote intramolecular charge transfer (ICT). It shows that the ICT effect is well improved by adding the D1 and Ai units. Compared with the original BTH molecule, the main bands of GSA and ESA of D1-D-Ai-π-A molecules are both red-shifted. In addition, GSA, ESA and MPA probabilities are all improved because the obvious charge transfer character results in the transition dipole moment change from localized to delocalized. Accordingly, the optical limiting effect in these hydrazone derivatives is well enhanced. These observations provide guidance for designing novel optical limiting materials based on the hydrazone derivatives.

Endophytic Fungal Diversity of Mangrove Ferns Acrostichum speciosum and A. aureum in China.

Microbial communities are an important component of mangrove ecosystems. In order to reveal the diversity of endophytic fungi in the mangrove ferns Acrostichum speciosum and A. aureum in China, the internal transcribed spacer (ITS) regions of endophytic fungi in four plant tissues (leaves, petioles, roots, and rhizomes) from three locations (Zhanjiang, Haikou, and Wenchang) were sequenced. The richness, species composition, and community similarity were analyzed. The main results are as follows: the dominant fungi in A. speciosum and A. aureum belonged to the phyla Ascomycota and Basidiomycota, accounting for more than 75% of the total identified fungi; in terms of species composition at the operational taxonomic unit (OTU) level, the endophytic fungi in A. aureum were more diverse than those in A. speciosum, and the endophytic fungi in rhizomes were more diverse than in other tissues. In Zhanjiang, both A. speciosum and A. aureum showed the richest diversity of endophytic fungi, both at the OTU classification level and in terms of species composition. Conversely, the richness of endophytic fungi in the samples of A. speciosum from Wenchang and Haikou is extremely low. The regional differences in dominant fungi increase with the degrading of taxonomic levels, and there were also significant differences in the number of unique fungi among different origins, with Zhanjiang samples having a larger number of unique fungi than the other locations. There were significant differences in the dominant fungi among different tissues, with Xylariales being the dominant fungi in rhizomes of A. speciosum and Hypocreales being the dominant fungi in the petioles, roots, and rhizomes of A. aureum. Overall, the community similarity of endophytic fungi among locations is moderately dissimilar (26-50%), while the similarity between tissues is moderately similar (51-75%). The low diversity of endophytic fungi could be one of the main reasons for the endangerment of A. speciosum. The protection of the diversity of endophytic fungi in the underground parts of A. speciosum is essential for the conservation of this critically endangered mangrove fern.

Iron-based autotrophic denitrification driven by sponge iron for nitrite removal in an anaerobic bioreactor: Effect of iron and carbon source.

The study investigated the denitrification effect of the iron autotrophic denitrification process for removing nitrite under anaerobic conditions, utilizing sponge iron as the electron donor. When the C/N ratio equaled 1, defined as the ratio of chemical oxygen demand to total nitrogen (TN), and the influent nitrite nitrogen (NO2--N) was at 80 mg/L, the average steady-state TN effluent concentration of this system was 41.94 mg/L during the 79-day experiment. The TN value exhibited a significant decrease compared to both the sponge iron system (68.69 mg/L) and the carbon source system (56.50 mg/L). Sponge iron is beneficial for providing an electron donor and ensuring an anaerobic system, fostering an environment that promotes microorganism growth while effectively inhibiting the conversion of nitrite to nitrate. In addition, carbon sources play a vital role in ensuring microorganism growth and reproduction, thereby aiding in TN removal. The optimal parameters based on the effectiveness of TN removal in the iron autotrophic denitrification system were determined to be s-Fe0 dosage of 30 g/L and C/N = 1.5. These results suggest that the iron autotrophic denitrification process, driven by sponge iron, can effectively remove nitrite under anaerobic conditions.

Functions and mechanisms of sponge iron-mediated multiple metabolic processes in anaerobic ammonium oxidation.

Sponge iron (SI) is a promising material for nitrogen removal from wastewater. This study reveals the potential functions and mechanisms of SI-mediated multiple metabolic processes in the nitrogen removal of Anammox. The results showed that although the SI application prolonged the start-up time of the reactor, achieved efficient and stable nitrogen removal after a successful start-up. The total nitrogen removal efficiency of the SI-Anammox system (92.62%) was 13.30% higher than that of R0 without SI (79.32%). The increase in nitrogen removal performance was accompanied by an increase in SAA and EPS content. Further microbial analysis showed significant enrichment of functional microorganisms, such as Candidatus_Brocadia, Nitrosomonas, Ellin6067, and Nitrospira. Multi-omics evidence suggests that efficient nitrogen removal is ultimately attributable to the enhancement of the specific key Fe- and N-functional genes in Anammox.

Survival Benefit of Adjuvant Chemotherapy in Node-Positive Breast Cancer With a 21-Gene Recurrence Score of 14 to 25: A Real-World Study Based on the Inverse Probability of Treatment Weighting Method.

INTRODUCTION: The role of recurrence score in predicting the benefits of adjuvant chemotherapy for lymph-node-positive breast cancer remains uncertain. We studied chemotherapy usage in patients with 1 to 3 positive lymph nodes and a recurrence score (RS) of 25 or lower to assess changes in clinical practice based on the RxPONDER trial. METHODS: A retrospective study using the SEER database identified female patients diagnosed with ER-positive, HER2-negative breast cancer, 1 to 3 positive lymph nodes, and an RS of 25 or lower between 2010 and 2015. Patients were divided into nonchemotherapy and chemotherapy groups, with propensity score weighting to balance clinicopathologic factors. RESULTS: Among 7965 patients, 5774 (72.5%) were in the nonchemotherapy group, while 2191 (27.5%) were in the chemotherapy group. Median follow-up was 39 months. Breast cancer accounted for 67 deaths, while 128 deaths were due to other causes. The weighted 5-year overall survival (OS) rates were 95.7% for the nonchemotherapy group and 97.2% for the chemotherapy group. For high-risk patients, the weighted 5-year OS rates were 95.2% and 97.0% for the nonchemotherapy and chemotherapy groups, respectively, with a significant absolute difference of 1.8% (P = .014). Multivariate analysis showed a significant difference in weighted hazard ratios for OS between the nonchemotherapy and chemotherapy groups in high-risk patients (hazard ratio: 0.64; 95% CI: 0.48-0.86). However, there were no significant differences in weighted hazard ratios for lower-risk patients, and similar results were observed for breast cancer-specific survival (BCSS). CONCLUSION: Patients with ER-positive, HER2-negative breast cancer and 1 to 3 positive lymph nodes, assessed by a 21-gene RS of 0 to 25, exhibited heterogeneous prognosis. Adjuvant chemotherapy provided a significant survival benefit, especially for patients with RS of 14 to 25, particularly those with invasive ductal carcinoma (IDC) and 2 to 3 positive lymph nodes.

Review on risk factors, classification, and treatment of sternal wound infection.

Sternal wound infection (SWI) is the most common complication of the median sternal incision. The treatment time is long, and the reconstruction is difficult, which causes challenges for surgeons. Plastic surgeons were often involved too late in such clinical scenarios when previous empirical treatments failed and the wound damage was relatively serious. Accurate diagnosis and risk factors against sternal wound infection need to be in focus. Classification of different types of sternotomy complications post-cardiac surgery is important for specific categorization and management. Not familiar with this kind of special and complex wound, objectively increasing the difficulty of wound reconstruction. The purpose of this comprehensive review is to review the literature, introduce various SWI risk factors related to wound nonunion, various classification characteristics, advantages and disadvantages of various wound reconstruction strategies, to help clinicians understand the pathophysiological characteristics of the disease and choose a better treatment method.

Response to HIV-1 gp160-carrying recombinant virus HSV-1 and HIV-1 VLP combined vaccine in BALB/c mice.

Human immunodeficiency virus (HIV) induced AIDS causes a large number of infections and deaths worldwide every year, still no vaccines are available to prevent infection. Recombinant herpes simplex virus type 1 (HSV-1) vector-based vaccines coding the target proteins of other pathogens have been widely used for disease control. Here, a recombinant virus with HIV-1 gp160 gene integration into the internal reverse (IR) region-deleted HSV-1 vector (HSV-BAC), was obtained by bacterial artificial chromosome (BAC) technology, and its immunogenicity investigated in BALB/c mice. The result showed similar replication ability of the HSV-BAC-based recombinant virus and wild type. Furthermore, humoral and cellular immune response showed superiority of intraperitoneal (IP) administration, compared to intranasally (IN), subcutaneous (SC) and intramuscularly (IM), that evidenced by production of significant antibody and T cell responses. More importantly, in a prime-boost combination study murine model, the recombinant viruses prime followed by HIV-1 VLP boost induced stronger and broader immune responses than single virus or protein vaccination in a similar vaccination regimen. Antibody production was sufficient with huge potential for viral clearance, along with efficient T-cell activation, which were evaluated by the enzyme-linked immunosorbent assay (ELISA) and flow cytometry (FC). Overall, these findings expose the value of combining different vaccine vectors and modalities to improve immunogenicity and breadth against different HIV-1 antigens.

Effects of aggregation on the structures and excited-state absorption for zinc phthalocyanine.

In the present paper, the aggregated structures of zinc phthalocyanine (ZnPc) have been investigated by considering its dimers and trimers. Based on the density functional theory calculations, two stable conformations are obtained for the ZnPc dimer and trimer, respectively. The IGMH (independent gradient model based on the Hirshfeld partition of molecular density) analysis reveals that the π-π interaction between the ZnPc molecules causes the aggregation. Normally, stacked structures with a slight displacement are favorable for aggregation. In addition, the planar structure of the ZnPc monomer is largely maintained in the aggregated conformations. For the presently obtained structures, the first singlet excited state absorption (ESA) spectra of these aggregated conformations of ZnPc were calculated based on the linear-response time-dependent density functional theory (LR-TDDFT), which has been well applied by our group. The results of the excited state absorption spectra reveal that the aggregation causes the ESA band to blue shift compared to the ZnPc monomer. By using the conventional description of the interaction between monomer transition dipoles, this blue shift is elucidated by the side-by-side transition dipole moments in the constituted monomers. The present results for the ESA combined with the previously reported results for ground state absorption (GSA) will provide guidelines to tune the window of the optical-limiting effect for the ZnPc based materials.

Ultrafast optical limiting ability of trans-stilbene enhanced and broadened by a donor-π-acceptor structure.

It has been shown that trans-stilbene (TSB) has great potential as an ultrafast optical limiting material through the process of three-photon absorption (3PA)-induced excited state absorption (ESA). The present paper shows that the main transitions in the absorption bands of TSB are mostly local excitation. In order to improve the optical limiting performance of TSB, a series of TSB derivatives with an electron donor-π-acceptor structure are designed. The analysis of π electron localized orbital locators (LOL-π) reveals that the distribution of π electrons in the derivatives of TSB is much more continuous compared to that in the original TSB. This results in the main transitions in the ground state absorption (GSA) and ESA of the TSB derivatives showing obvious charge transfer characteristics, and the GSA, ESA and 3PA bands are largely enhanced and broadened compared to those of the original TSB molecule. These observations are well supported by the enlarged transition dipole moments of the main transitions in GSA and ESA. With these results, it is clearly shown that the TSB derivatives are promising optical limiting materials. Our observations provide clues for the development of optical limiting materials based on TSB.

The application of the Limberg flap repair technique in the surgical treatment of pilonidal sinus disease.

Pilonidal sinus disease (PNSD) challenged surgeons for decades. Limberg flap repair (LFR) is a common treatment for PNSD. The purpose of this study was to observe the effect and risk factors of LFR in PNSD. A retrospective study was conducted on the PNSD patients who visited two medical centers and four departments in the People's Liberation Army General Hospital and were taking LFR treatment between 2016 and 2022. The risk factors, the effect of the operation, and complications were observed. The effects of known risk factors on the surgical results were compared. There were 37 PNSD patients: male/female ratio of 35:2, average age: 25.1 ± 7.9 years. Average BMI: 25.2 ± 4.0 kg/m2 , average wound healing time: 15.4 ± 3.4 days. 30 patients (81.0%) healed in stage one and 7 (16.3%) had postoperative complications. Only 1 patient (2.7%) had a recurrence while others were healed after dressing-changing. There was no significant difference in age, BMI, preoperative debridement history, preoperative sinus classification, Wound area, Negative pressure drainage tube, prone time (<3d) and treatment effect. Squat defecate and premature defecation were associated with treatment effect, and they were independent predictors of treatment effect in the multivariate analysis. LFR has a stable therapeutic outcome. Compared with other skin flaps, the therapeutic effect of this flap is not significantly different, but the design is simple and is not affected by the known risk factors before operation. However, it is necessary to avoid the influence of two independent risk factors, squatting defecation and premature defecation, on the therapeutic effect.

Effects of sponge iron dosage on nitrogen removal performance and microbial community structure in sequencing batch reactors.

The application of sponge iron (SI) carriers can improve the biochemical treatment performance of sequencing batch reactors (SBR) during wastewater treatment. This study used SBR reactors to explore the effects of SI dosage on the nitrogen removal performance and reactor stability and microbial community structure under low temperature and ultra-low load. In contrast to conventional SBR, the average removal rate of total nitrogen (TN) in the biological sponge iron system (BSIS) was increased by 5.38 % for 45 g/L, 18.93 % for 90 g/L, and 13.52 % for 135 g/L, respectively. The nitrogen removal performance and reactor stability showed the best performance under the SI dosage of 90 g/L. The addition of SI formed the anaerobic-anoxic-aerobic microenvironments, which facilitate the propagation of denitrifying bacteria (Saccharimonadales, Hydrogenophaga) and iron bacteria (Rhodoferax and Acinetobacter) in the BSIS. This study provides a new insight on the application of SI in the wastewater treatment.

Three-photon-induced singlet excited-state absorption for tunable ultrafast optical-limiting in distyrylbenzene: a first-principles study.

The ground and first singlet excited state absorption in distyrylbenzene (DSB) is simulated based on linear-response time dependent density functional theory (LR-TDDFT). It is found that distyrylbenzene shows a strong reverse saturable absorption effect around the near-infrared range. Combining the calculations of cubic response functions to simulate the three-photon absorption in distyrylbenzene, we are able to show that distyrylbenzene is a promising ultrafast optical limiter for the light with wavelengths around 775 nm. The primary mechanism for the optical limiting behavior can be well understood by the three-photon induced excited state absorption (3PA-ESA). This result in that DSB has high transmittance for low-intensity ambient light levels and the ultrafast response of optical-limiting. In addition, the limited optical window can be tuned by changing the length of the π-electron conjugated structure. It was also discovered that the molecular aggregation has an inhibitory effect on the optical limiting efficiency of distyrylbenzene. The present results may serve as a theoretical guideline for the design of distyrylbenzene-based optical limiting materials.

Genetic diversity analysis of grapevine rupestris stem pitting-associated virus from grapevine by colony PCR-SSCP and -RFLP.

We have developed methods for detecting the genetic diversity of grapevine rupestris stem pitting-associated virus (GRSPaV) based on restriction fragment length polymorphism (RFLP) and single stranded conformational polymorphism (SSCP) in the 905 nt 3' sequence. The amplicons were cloned from six grapevine cultivars, and colony polymerase chain reaction (colony PCR) using recombination bacteria was subsequently analyzed by RFLP and SSCP. Four haplotypes of SSCP and six haplotypes of Sac I RFLPs were defined. The two methods had a 40% discrepancy rate in showing the degree of diversity. All clones were sequenced and were used to construct a phylogenetic tree with seven previously reported GRSPaV sequences. In the tree, all the newly acquired sequences were divided into three clusters, I, II, and III, which corresponded to haplotypes I, II, and III of SSCP, respectively. Haplotype IV of SSCP was grouped into cluster II. A recombination analysis showed that haplotype IV has undergone a recombination event. Together, these results indicate that the SSCP assay is useful for the rapid identification of genetic diversity of GRSPaV. This is the first report of an analysis of the large fragment of GRSPaV by colony PCR-SSCP. Keywords: grapevine; grapevine rupestris stem pitting-associated virus (GRSPaV); RFLP; SSCP; genetic diversity analysis.

Self-Assembly of [3]Catenane and [4]Catenane Based on Neutral Organometallic Scaffolds.

Transition metal-mediated templating and self-assembly have shown great potential to construct mechanically interlocked molecules. Herein, we describe the formation of the bimetallic [3]catenane and [4]catenane based on neutral organometallic scaffolds via the orthogonality of platinum-to-oxygen coordination-driven self-assembly and copper(I) template-directed strategy of a [2]pseudorotaxane. The structures of these bimetallic [3]catenane and [4]catenane were characterized by multinuclear NMR {1H and 31P} spectroscopy, electrospray ionization time-of-flight mass spectrometry (ESI-TOF-MS), and PM6 semiempirical molecular orbital theoretical calculations. In addition, single-crystal X-ray analyses of the [3]catenane revealed two asymmetric [2]pseudorotaxane units inside the metallacycle. It was discovered that tubular structures were formed through the stacking of individual [3]catenane molecules driven by the strong π-π interactions.

Variations in Behavioral and Physiological Traits in Yearling Tibetan Sheep (Ovis aries).

Temperament is a consistent behavioral difference among individuals over time or in different contexts. A comprehensive understanding of temperament and complex behavioral interactions enhances knowledge on animal evolution, welfare, and productivity. However, reports on the development of behavioral consistency over ontogeny are vague. Here, we tested the ontogeny of the temperament and physiological traits of Tibetan sheep (Ovis aries) in three crucial age stages. The mean level of the risk-taking variable increased, while that of the vocalizations variable decreased. The exploration variable was stable over ontogeny. The novelty decreased and the heart rate increased from the juvenile to the adolescent stage but stabilized at the adult stage. The fecal cortisol concentration (CORT) variable was stable at the juvenile and adolescent stages but decreased at the adult stage. Stable correlations were reported for the juvenile and adolescent stages and for the behavioral variables and heart rate. However, some correlations emerged only after maturation, whereas others disappeared over ontogeny. Moreover, CORT was independent of temperament and heart rate at different ages. These results demonstrate that age affects temperament and physiology and their correlations. Hence, developmental aspects should be incorporated into future temperament studies.

Prenatal, perinatal, and postnatal factors associated with autism spectrum disorder cases in Xuzhou, China.

BACKGROUND: The aim of the present study was to explore the prenatal, perinatal, and postnatal risk factors in children with autism spectrum disorder (ASD) from Xuzhou, China by comparing them with healthy children. METHODS: Children with ASD who received rehabilitation training at special education schools and rehabilitation institutions in Xuzhou were selected as the ASD group, and healthy children during the same period were selected as the healthy non-ASD group. A questionnaire based on the possible causes and susceptibility factors of ASD in children was issued and given to all children in this study. RESULTS: The findings of the present study revealed a higher prevalence of prenatal, perinatal, and postnatal factors in children with ASD compared with healthy children. There were significantly more males than females in the ASD group, and the proportion of boys to girls was 5.81:1 (P<0.05). Logistic regression analysis suggested that the risk factors of male children developing ASD were feeding difficulties, poor living environment during pregnancy, maternal exposure to cigarette smoking during pregnancy, and perinatal hypoxia. Factors associated with ASD risk among were identified, such as living environment during pregnancy, delivery method, feeding difficulties, and epilepsy (P<0.05). Feeding difficulties and living in the countryside during pregnancy might be risk factors for ASD in girls according to the logistic regression analysis. CONCLUSIONS: This survey confirmed the high prevalence of prenatal, perinatal, and postnatal factors in children with ASD. Some of these factors may be effective entry points for the prevention and treatment of ASD.

Differential Expression of Metabolism-Related Genes in Plateau Pika (Ochotona curzoniae) at Different Altitudes on the Qinghai-Tibet Plateau.

According to life history theory, animals living in extreme environments have evolved specific behavioral and physiological strategies for survival. However, the genetic mechanisms underpinning these strategies are unclear. As the highest geographical unit on Earth, the Qinghai-Tibet Plateau is characterized by an extreme environment and climate. During long-term evolutionary processes, animals that inhabit the plateau have evolved specialized morphological and physiological traits. The plateau pika (Ochotona curzoniae), one of the native small mammals that evolved on the Qinghai-Tibet Plateau, has adapted well to this cold and hypoxic environment. To explore the genetic mechanisms underlying the physiological adaptations of plateau pika to extremely cold ambient temperatures, we measured the differences in resting metabolic rate (RMR) and metabolism-related gene expression in individuals inhabiting three distinct altitudes (i.e., 3,321, 3,663, and 4,194 m). Results showed that the body mass and RMR of plateau pika at high- and medium-altitudes were significantly higher than those at the low-altitude. The expression levels of peroxisome proliferator-activated receptor α (pparα), peroxisome proliferator-activated receptor-γ coactivator-1α (pgc-1α), and the PR domain-containing 16 (PRDM16) in white (WAT) and brown (BAT) adipose tissues of plateau pika from high- and medium-altitudes were significantly higher than in pika from the low-altitude region. The enhanced expression levels of pgc-1α and pparα genes in the WAT of pika at high-altitude showed that WAT underwent "browning" and increased thermogenic properties. An increase in the expression of uncoupling protein 1 (UCP1) in the BAT of pika at high altitude indicated that BAT increased their thermogenic properties. The gene expression levels of pparα and pgc-1α in skeletal muscles were significantly higher in high-altitude pika. Simultaneously, the expression of the sarcolipin (SLN) gene in skeletal muscles significantly increased in high-altitude pika. Our results suggest that plateau pika adapted to an extremely cold environment via browning WAT, thereby activating BAT and enhancing SLN expression to increase non-shivering thermogenesis. This study demonstrates that plateau pika can increase thermogenic gene expression and energy metabolism to adapt to the extreme environments on the plateau.

Epidemiology of burns in pediatric patients of Beijing City.

BACKGROUND: This study aimed to assess the epidemiological characteristics of pediatric burns in Beijing City. METHODS: This was a retrospective study of pediatric patients (n = 400) admitted to four burn centers in Beijing City between June 2010 and May 2011. Burn severity was determined according to total body surface area (TBSA) percentage and degree. Patients were followed up for one year. Multivariate analyses were carried out to determine the factors (burn etiology, time and place of injury, living conditions, hospital type, first-aid treatment methods, and parent/guardian knowledge of burns, educational level, occupation) affecting burn properties (severity and pigmentation/scar). RESULTS: 191/400 (47.8 %) patients were aged 2-3 years, and scalding was the leading etiology (355/400, 88.8 %). Burn incidence peaked in May (14.8 %), at 10:00-12:00 and 17:00-20:00. Most burn events occurred indoors (272/400, 68.0 %), especially in the kitchen (180/400, 45.0 %). Roughly half of them involved head and neck; 188 (47.0 %) patients had mild burns, 140 (35.0 %) moderate, 44 (11.0 %) extensive, and 28 (7.0 %) critical burns; 184 (46.0 %) patients were treated only with cold-water rinsing or compress; 120 (30.0 %) received no first aid. Only 32 (8.0 %) patients visited a specialized institution. 164 patients underwent surgery. Hospitalization lasted for 14.8 ± 8.1 days. Independent risk factors for burn severity were occurrence month, living conditions, occupation of the mother, and first aid. 288 (72.0 %) patients developed pigmentation and scar within a year while no independent risk factors was observed. CONCLUSIONS: Pediatric burns often occurred indoors, especially in the kitchen, and a substantial proportion receives no first aid.

Expression and immunogenicity of recombinant glycoprotein D of herpes simplex virus 1 in Drosophila S2 cells.

Herpes simplex virus type 1 (HSV-1) is responsible for cold sores in the general population, but also contributes to the development of other more serious diseases in some circumstances. The viral glycoprotein D (gD) is essential for virus entry into host cells. In the present study, the Drosophila melanogaster Schneider 2 (S2) expression system (DES) was evaluated for the expression of recombinant gD1. The DNA sequences encoding the full-length gD1 (369aa, FLgD1) and a truncated gD1 form corresponding to the ectodomain (314aa, EgD1) were cloned into S2 expression vector pMT/BiP/V5-HisA to generate pMT-EgD1 and pMT-FLgD1, respectively. Two forms of gD1 gene were fitted with a hexahistidine tag to facilitate their purification. Cell populations expressing the highest gD1 levels were selected by using a limiting dilution assay. Western blot, flow cytometry (FACS), and confocal immunofluoresence assay demonstrated that the full-length form is restrained in the lipid membranes of the cell and the ectodomain form is secreted into the medium. Recombinant ectodomain gD1 was scaled up and purified from the culture medium using nickel nitrilotriacetic acid affinity chromatography, and a maximum production level of 56.8 mg/L of recombinant gD1 was obtained in a shake-flask culture of S2 cells after induction with 5 µM CdCl2 for 4 days. Mice were then immunized with recombinant purified gD1 and produced high titers of antibody measured by enzyme-linked immunosorbent assay (ELISA; 1:5,120,000) as well as high plaque neutralization titer (1:320). Overall, the data indicated that stable expression in S2 cells is a practical way of synthesizing gD1 for use in structural and functional studies in the further study.

Molecular characterizations of two grapevine rupestris stem pitting-associated virus isolates from China.

The complete nucleotide sequences of two isolates of grapevine rupestris stem pitting-associated virus (LSL and JF) collected from grapevine of Xingcheng in Liaoning Province, China, were determined. The genomes of both LSL and JF were found to contain five open reading frames (ORFs). Sequence alignments showed that the genomic sequences of JF were 76.1 %-83.5 % identical to the other ten GRSPaV isolates that have been reported previously and that the nucleotide sequence identity of isolate LSL to other isolates was no more than 78 %. Phylogenetic analysis based on the complete genome sequence indicated that JF belongs to group III and that LSL belongs to a new group (group IV). The average genetic distances of the new genetic lineage from groups I, II and III were 0.34, 0.32 and 0.33, respectively.