[Evaluation of classification of petroclival meningiomas and proposed selection of microsurgical approach: a single center experience of 179 cases].

Objective: To evaluate the classification of petroclival meningiomas(PCM), proposed selection of microsurgical approach and therapeutic outcomes. Methods: Retrospectively analyzed clinical data of 179 cases of PCM from Department of Neurosurgery, Xiangya Hospital, Central South University between January 2011 and November 2020. There were 28 males and 151 females with an age of(49.9±10.2) years(range: 22 to 75 years) and the tumor size of(44.8±10.3)mm(range: 15 to 80 mm). The mean duration of symptom(M(QR)) was 18.0(40.6)months(range:1 week to 320 months) and the mean preoperative Karnofsky performance scale(KPS) was 78.6±13.3(range: 40 to 100). The PCM were classified into 5 types according to the difference in the origin of dural attachment, involvement of adjacent structures and growth patterns through preoperative MRI. The surgical approaches were selected based on the proposed classification, and the clinical characteristics, surgical record, and follow-up data of each type were reviewed. Results: The PCM were divided into clivus type(CV, 4 cases), petroclival type(PC, 60 cases), petroclivosphenoidal type(PC-S, 62 cases), sphenopetroclival type with 2 subtypes(S-PC, 50 cases) and central skull base type(CSB, 3 cases). All of 176 cases were obtained microsurgical treatment except CSB type. The gross total resection reached in 124 cases(70.5%) with 112 cases of retrosigmoid approach(RSA), 27 cases of subtemporal transtentorial transpetrosal approach, 13 cases of pretemporal trancavernous anterior transpetrosal approach(PTCA), 12 cases of extended pterional transtentorial approach(EPTA) and presigmoid combined supra-infratentorial approach, respectively. The RSA could be adopted in both of CV type and PC type and most of PC-S type(71.0%). S-PC subtype Ⅰ and subtype Ⅱ were mainly underwent EPTA(40.8%) and PTCA(52.2%), respectively. Seventy-two cases(40.9%) gained new neurological dysfunctions mainly with the cranial nerve paralysis. The postoperative morbidity and complications were recovered or improved with conservative and positive symptomatic and supportive treatment. There was no intraoperative and postoperative death case. One hundred and sixty four cases(93.2%) of operative patients were followed with the duration of 24(48)months(range:3 to 108 months). Tumor recurrence and progress were identified in 14 cases(10.4%) and 14 cases(28.6%), respectively. Compared with postoperative neurological status, 89 patients(54.3%) had improved and 38 patients(23.2%) were still suffering various degrees of neurological dysfunctions during the follow-up. The recent KPS was 84.2±11.4(range: 50 to 100) without statistical difference from preoperative KPS(t=-1.356,P=0.125). As for each type, there were statistically significant differences in brain stem edema(χ2=3.482,P=0.038), gross total resection(χ2=9.127,P=0.001), surgical duration(F=8.954, P=0.013), postoperative length of stay(F=3.652, P=0.025), postoperative complications(χ2=1.550,P=0.024), postoperative KPS(F=2.856, P=0.042) and tumor recurrence/progress(χ2=4.824,P=0.013). Conclusion: The precise and comprehensive classification of PCM and specific individual treatment strategy are benefit to evaluate the diverse clinical prognosis, choose optimal surgical approaches, elevate gross total resection, diminish neurological dysfunctions and restraint tumor recurrence, so as to improve the quality of life for patients.

[A homozygous variant in a consanguineous pedigree with inherited protein S deficiency].

Objective: To observe the clinical feature of familiar hereditary protein S deficiency, and to explore the related gene mutation. Methods: The blood samples were obtained from the proband and the family memebers(3 generations,6 persons). PROS1 gene of the proband and the family members was analyzed. The 15 exons and flanking sequence of PROS1 gene were analyzed by PCR and DNA sequencing. Results: Five out of 6 family members were diagnosed as having hereditary protein S deficiency. The proband suffered from pulmonary embolism. The others had no obvious thrombotic event. The gene sequencing revealed that the proband carried a c.-168C>T homozygous variant in the promoter of exon 1. His parents, brother and son all carried c.-168C>T heterozygosis variant at the same position. The gene of his wife was a wild type. Conclusion: A gene variant (c.-168C>T) of PROS1 was discovered in this Chinese family. Gene variant of PROS1 may result in protein S deficiency. Patients with protein S deficiency may suffer from vein thrombosis and(or) pulmonary embolism.

[Effects of human leucocyte antigen-G expression on invasion and proliferation of chorionic trophoblastic cell line JEG-3].

Objective: To investigate the effects of human leukocyte-associated antigen-G (HLA-G) expression in silencing trophoblast cell line JEG-3 under normal and hypoxic conditions on invasion and proliferation of JEG-3 cells. Methods: Inhibition of HLA-G expression in JEG-3 cells by transfection of small interfering RNA (siRNA),the transfected JEG-3 cells were divided into 4 groups: normoxia control group, hypoxia control group, normoxia inhibition group and hypoxia inhibition group. The levels of HLA-G mRNA and protein in 4 groups of cells were detected by real-time quantitive PCR and western blot. The proliferation activity and invasion ability of 4 groups of cells were determined by methylthiazolyl tetrazolium (MTT) assay and invasion assay. Results: (1) Real-time quantitive PCR technology showed: the level of HLA-G mRNA in the hypoxic inhibition group (0.220±0.050) was significantly different (P<0.05), when compared with that in the hypoxic control group (0.630±0.030) and normoxic inhibition group (0.400±0.020). (2) Western blot analysis showed: the expression level of HLA-G protein in the hypoxic inhibition group was 0.260±0.010, statistically different from that in the hypoxic control group (0.850±0.100) and the normoxic inhibition group (0.560±0.020; P<0.05).(3) MTT showed: proliferative activity of JEG-3 cells in the normoxic inhibition group was 0.490±0.070, the ability of cell proliferation was reduced. When compared with that in the normoxic control group (0.850±0.050), the differences was statistically significant (P<0.05). The proliferative activity of JEG-3 cells in the hypoxic inhibition group (0.330±0.070) was lower than that in the normoxic inhibition group (0.490±0.070), and there was a significant difference (P<0.05). (4) Invasion assay showed: compared with the normoxic control group (98±7), the invasive ability of JEG-3 cells in the normoxic inhibition group (73±7) was weakened, and the difference was statistically significant (P<0.05). The number of transmembrane cells (52±11) of JEG-3 cells in the hypoxic inhibition group was lower than that in the hypoxic control group (72±7), and the difference was statistically significant (P<0.05). Compared with the normoxic inhibition group, the invasion ability of JEG-3 cells in the hypoxic inhibition group decreased, and the difference was statistically significant (P<0.05). Conclusion: Under hypoxia, using siRNA technology to down-regulate the expression of HLA-G may affect the proliferation and invasion ability of trophoblast cells, which may be involved in the occurrence of hypertensive disorder of pregnancy.

[Application of multiplex PCR assay to study early multiple herpesviruses infection during HSCT].

Objective: To investigate herpesvirus infection in early stage of hematopoietic stem cell transplantation (HSCT) by multiplex polymerase chain reaction (PCR), and to explore the association between multiple herpesviruses infection and clinical characteristics in HSCT patients and its impact on post-transplant complications and prognosis. Methods: A total of 734 peripheral blood samples were collected from 90 patients undergoing HSCT in the Department of Hematology, the First Affiliated Hospital of Soochow University between February 2017 and August 2017. The peripheral blood specimens were obtained before and within 90 days after transplantation at different time points. Lab-Aid824 Nucleic Acid Extraction Mini Reagent was used to extract DNA and multiplex PCR assay was used to simultaneously detect 8 kinds of human herpesviruses from genomic DNA. The incidence of various herpesvirus infections, its correlation with clinical features and effects on post-transplant complications and prognosis were analyzed. Results: The median follow-up time was 192 (range: 35-308) days. Among the 90 patients before transplantation, the incidence of herpes virus infection was 35.6% (32/90), including 12.2% (11/90) with one herpes virus infection and 23.3% (21/90) with multiple viruses infection. The incidence of herpes virus infection after transplantation was 77.8% (70/90), including 20.0% (18/90) with one herpes virus infection and 57.8% (52/90) with multiple herpes virus infection. Among the 52 patients with multiple herpes viruses infection, 30 (57.7%) patients were infected by 2 kinds of viruses, 18 (34.6%) patients by 3 kinds of viruses and 4 (7.7%) patients by 4 kinds of viruses. There was a correlation between HHV-6 and HHV-7 herpesvirus infection (OR=13.880, Q=0.026). EBV infection was related to HHV-7 infection (OR=0.093, Q=0.044). The age of patients was correlated with the incidence of HHV-1 infection before transplantation. There were 24 patients in our study experienced clinical symptoms associated with viral infection. The main manifestations were hemorrhagic cystitis (HC), interstitial pneumonia, enteritis, viral encephalitis and fever of unknown origin. EBV infection was related to HLA incompatibility and the inconsistent of the ABO blood group and grade Ⅱ-Ⅳ aGVHD after transplantation. HLA incompatibility and the unrelated donor and grade Ⅱ-Ⅳ aGVHD were related to multiple viruses infection. Conclusion: Multiple herpesviruses were common in patients undergoing HSCT, which were closely related to HLA mismatch, unrelated donor and grade Ⅱ-Ⅳ aGVHD.

[Paracondylar-lateral cervical approach for resection of jugular foramen schwannomas: a retrospective analysis of 15 cases].

Objective: To explore the effects of paracondylar-lateral cervical approach for resection of the jugular foramen schwannoma(JFS). Methods: A total of 15 patients with JFS operated via the paracondylar-lateral cervical approach between December 2011 and March 2016 at Department of Neurosurgery in Xiangya Hospital of Central South University were retrospectively analyzed. There were 7 males and 8 females, aging from 22 to 77 years with a mean age of (41.9±15.8) years.There were 12 patients who accepted primary surgery, 3 patients who accepted secondary surgery. There were 10 patients with tinnitus or hearing loss, 8 patients with dysphagia, 9 patients with hoarseness, 7 patients with tongue hemiparesis, 8 patients with ataxia, 1 patient with Pyramidal signs, 2 patients with facial hypesthesia or pain, 1 patient with facial paresis. According to Samii JFS grading system, 2 patients were type B, 4 were type C and 9 were type D. All patients were followed-up through outpatient and telephone by MRI in 3, 9, 12 months postoperation. Results: Gross total removal of tumors were achieved in 14 patients and subtotal removal in 1 patient. Two patients had new hoarseness, 2 had new dysphagia and 3 had more serious dysphagia, 1 patient had more serious facial paresis after the operation. There were 2 patients with intracranial infection, 2 with pneumonia, 1 with subcutaneous effusion, 1 with cerebrospinal fluid rhinorrhea, 5 with gastric intubation during perioperative period. There were no death, intracranial hematoma and decreased hearing patients. All patients were followed up, the follow-up time were 3 to 33 months with a mean of (26.9±11.2) months. Till to the latest follow up, dysphagia improved in 2 cases, hoarseness and tongue hemiparesis improved in 3 cases, hearing loss and tinnitus improved in 9 cases, balance function improved in 7 cases, facial hypesthesia and pain improved in 2 cases, pyramidal signs disappeared in 1 case, facial nerve function improved to normal in 1 case. There was no recurrence and progressed case. Conclusion: Paracondylar-lateral cervical approach is an alternative approach for both intracranial and extracranial JFS.

Lack of Association between SLC6A11 Genetic Polymorphisms and Drug Resistant Epilepsy in Chinese Han Population.

BACKGROUND: Epilepsy is one of the most common chronic neurological diseases. A recent study has implicated that the genetic variants of the SLC6A11 gene encoding GAT-3, an astrocytic GABA transporter, may influence the efficacy of antiepileptic drugs (AEDs) in the Korean population. METHODS: This study aims to investigate the possible associations between SLC6A11 gene and drug resistance in Chinese epilepsy patients. Genomic DNA from 240 drug resistant epilepsy (DRE) patients and 336 drug responsive epilepsy patients was tested for the polymorphisms using Illumina GoldenGate assay. RESULTS: None of the 14 tagSNP alleles and genotypes were found to be related to DRE. The frequencies of haplotype 5 was obviously lower in DRE patients than that in drug responsive epilepsy patients (1% vs. 4%, OR = 2.56 [0.107 - 0.763], p = 0.01). However, after the correction of multiple comparisons with Bonferroni's method, we found that haplotype 5 was not associated with DRE. CONCLUSIONS: Our study suggested no existing association between the 14 SNPs of SLC6A11 and AEDs efficacy in the Chinese Han population.

[Size exclusionchromatography-high-performance liquid chromatography-inductively coupled plasma mass spectrometry for measuring the stability of cadmium telluridequantum dots].

Objective: To investigate the peak time and peak area of elements in cadmium telluride quantum dots (CdTe QDs) using size exclusion chromatography-high-performance liquid chromatography-inductively coupled plasma mass spectrometry, as well as the biological stability of CdTe QDs in vivo and in vitro. Methods: Transmission electron microscope and ultraviolet fluorescence were used for characterization and synthesis of water-soluble CdTe QDs, and CdTe QDs were added to double-distilled water, mobile phase, or bovine serum medium to observe the change in stability after different periods of time. CdTe QDs were injected into the vein of mice, and the changes in the morphology of CdTe QDs in serum and the liver were measured at 1, 24, and 72 hours after exposure. Size exclusion chromatography-high-performance liquid chromatography was used for the elution of the compounds in the solution based on their volume, and then inductively coupled plasma mass spectrometry was performed for the eluent. The flow time of (114)Cd and (130)Te and molar ratio were used for qualitative analysis of CdTe QDs, and the peak area was used to judge whether CdTe QDs were degraded. Results: CdTe QDs were diluted to a concentration of 0.5 mmol/L with double-distilled water and then placed in a dark place at room temperature; CdTe QDs were completely degraded after 60 minutes. CdTe QDs were diluted to a concentration of 0.005 mmol/L with a mobile phase, and the peak of CdTe QDs was not detected. After CdTe QDs were placed in a dark place at room temperature for 48 hours at a concentration of 0.005 mmol/L in bovine serum mediumin vitro, the peak area of (114)Cd was 6179841-7346084, and the peak area of (130)Te was 1077913-1191066. CdTe QDs had the highest peak area at 1 hour after exposure, and the peak areas of (114)Cd and (130)Te were 18183894 and 25187987, respectively. CdTe QDs were quickly degraded in the liver; at 1 hour after exposure, the degradation products of CdTe QDs containing Cd were observed in liver tissue homogenate, and CdTe QDs were largely degradedat 24 hours. Conclusion: This method can be used to investigate the biological stability of CdTe QDs. CdTe QDs are degraded in the liver and produce Cd(2+), which may cause toxic reaction.

Drug-drug interaction between valproic acid and meropenem: a retrospective analysis of electronic medical records from neurosurgery inpatients.

WHAT IS KNOWN AND OBJECTIVE: A series of studies have indicated that valproic acid (VPA) plasma concentration decreased rapidly when used concomitantly with carbapenem antibiotics, including meropenem (MEPM), imipenem and panipenem, which may increase the risk of seizure breakthrough. However, the cause for the change in VPA pharmacokinetics is unclear. A retrospective analysis of VPA therapeutic drug monitoring (TDM) records was performed to investigate this VPA pharmacokinetics drug-drug interaction. METHODS: Three hundred and eighty one VPA TDM records from the Department of Neurosurgery of Xiangya Hospital from January 2012 to December 2014 were collected. The VPA TDM records were categorized by VPA and MEPM daily dosages in grams/day (g/day). A comparison of VPA plasma levels among different groups was performed to investigate the change in VPA level in this drug interaction. RESULTS AND DISCUSSION: Remarkable decreases in VPA plasma level were observed when the drug was used concomitantly with MEPM in both 1.2 g/d and 1.6 g/d VPA groups (67·3 ± 4·6 µg/mL, n = 21 vs. 15·3 ± 1·9 µg/mL, n = 15, P < 0·001; 67·6 ± 1·2 µg/mL vs. 18·1 ± 2·6 µg/mL, n = 38, P < 0·001). No significant difference in VPA plasma concentrations was observed between the 1·2 g/day VPA + MEPM, 1·6 g/day VPA + MEPM and 2·0 g/day VPA + MEPM groups (15·3 ± 1·9 µg/mL, n = 15 vs. 18·1 ± 2·6 µg/mL, n = 38 vs. 9·0 ± 3·0 µg/mL, n = 7; P = 0·252). The decrease in VPA concentration was independent of MEPM daily dose (14·0 ± 5·1 µg/mL, n = 4 for high MEPM daily dose vs. 16·5 ± 1·9 µg/mL, n = 56 for low MEPM daily dose; P = 0·729). After discontinuation of MEPM for more than 7 days, VPA plasma concentration recovered to a value comparable to that before MPEM initiation (69·7 ± 4·2 µg/mL, n = 21 vs. 51·2 ± 8·1 µg/mL, n = 9; P = 0·48). WHAT IS NEW AND CONCLUSION: This is the first study using a large number of VPA TDM records to investigate the change in VPA levels caused by concomitant use of MEPM. Our results imply that the decrease in drug concentration cannot be reversed by increasing VPA dose. Moreover, MEPM daily dose did not influence the drop in VPA plasma level. At least 7 days are required for the recovery of VPA plasma concentration after discontinuation of MEPM.

Classification and regression tree analysis of acute-on-chronic hepatitis B liver failure: Seeing the forest for the trees.

At present, there is no ideal model for predicting the short-term outcome of patients with acute-on-chronic hepatitis B liver failure (ACHBLF). This study aimed to establish and validate a prognostic model by using the classification and regression tree (CART) analysis. A total of 1047 patients from two separate medical centres with suspected ACHBLF were screened in the study, which were recognized as derivation cohort and validation cohort, respectively. CART analysis was applied to predict the 3-month mortality of patients with ACHBLF. The accuracy of the CART model was tested using the area under the receiver operating characteristic curve, which was compared with the model for end-stage liver disease (MELD) score and a new logistic regression model. CART analysis identified four variables as prognostic factors of ACHBLF: total bilirubin, age, serum sodium and INR, and three distinct risk groups: low risk (4.2%), intermediate risk (30.2%-53.2%) and high risk (81.4%-96.9%). The new logistic regression model was constructed with four independent factors, including age, total bilirubin, serum sodium and prothrombin activity by multivariate logistic regression analysis. The performances of the CART model (0.896), similar to the logistic regression model (0.914, P=.382), exceeded that of MELD score (0.667, P<.001). The results were confirmed in the validation cohort. We have developed and validated a novel CART model superior to MELD for predicting three-month mortality of patients with ACHBLF. Thus, the CART model could facilitate medical decision-making and provide clinicians with a validated practical bedside tool for ACHBLF risk stratification.

Association analysis of IL-17A and IL-17F polymorphisms in Chinese women with cervical cancer.

We selected six tagged single nucleotide polymorphisms (SNPs) in the interleukin 17A (IL-17A) and IL-17F genes, and evaluated the relationship between the six common SNPs and environmental factors in cervical cancer patients. Polymerase chain reaction-restriction fragment length polymorphism was used to detect the IL-17A (rs2275913, rs3748067, and rs3819025) and IL-17F (rs763780, rs9382084, and rs1266828) SNPs. The associations between IL-17A and IL-17F gene polymorphisms and risk of cervical cancer were estimated by conditional logistic regression. Compared with the control subjects, the cervical cancer patients had a lower age at first live birth, a habit of smoking, a family history of cancer, and a greater incidence of human papillomavirus-16 or 18 infections. The logistic regression analysis showed that the variant AA genotype of rs2275913 was associated with a significantly higher risk of cervical cancer than the wild-type GG genotype (OR = 1.99, 95%CI = 1.12-3.50). However, no evidence of the association was observed between rs3748067, rs3819025, rs763780, rs9382084, and rs1266828 polymorphisms and the risk of cervical cancer. We suggest that rs2275913 may play a role in the etiology of cervical cancer. These findings could be helpful in identifying individuals at increased risk of developing cervical cancer.

BAC transgenic mice provide evidence that p53 expression is highly regulated in vivo.

p53 is an important tumor suppressor and stress response mediator. Proper control of p53 level and activity is tightly associated with its function. Posttranslational modifications and the interactions with Mdm2 and Mdm4 are major mechanisms controlling p53 activity and stability. As p53 protein is short-lived and hardly detectable in unstressed situations, less is known on its basal level expression and the corresponding controlling mechanisms in vivo. In addition, it also remains obscure how p53 expression might contribute to its functional regulation. In this study, we established bacterial artificial chromosome transgenic E.coli ß-galactosidase Z gene reporter mice to monitor p53 expression in mouse tissues and identify important regulatory elements critical for the expression in vivo. We revealed preferentially high level of p53 reporter expressions in the proliferating, but not the differentiated compartments of the majority of tissues during development and tissue homeostasis. In addition, tumors as well as regenerating tissues in the p53 reporter mice also expressed high level of ß-gal. Furthermore, both the enhancer box sequence (CANNTG) in the p53 promoter and the 3' terminal untranslated region element were critical in mediating the high-level expression of the reporter. We also provided evidence that cellular myelocytomatosis oncogene was a critical player regulating p53 mRNA expression in proliferating cells and tissues. Finally, we found robust p53 activation preferentially in the proliferating compartment of mouse tissues upon DNA damage and the proliferating cells exhibited an enhanced p53 response as compared with cells in a quiescent state. Together, these results suggested a highly regulated expression pattern of p53 in the proliferating compartment controlled by both transcriptional and posttranscriptional mechanisms, and such regulated p53 expression may impose functional significance upon stress by setting up a precautionary mode in defense of cellular transformation and tumorigenesis.

Application of 64-slice spiral computed tomography angiography in extremity vascular injuries.

The objective of the current study was to assess the utility of 64-row helical computed tomography angiography (CTA) in the evaluation of extremity vascular traumas. The extremities from 17 clinical cases of suspected traumatic vascular damage were evaluated using 64-row helical CTA. To evaluate extremity vascular traumas using CTA, volume rendering, multiple planar reconstruction, and curved planar reconstruction technology were applied to accurately and rapidly indicate the type and extent of blood vessel damage, as well as any relationship with injuries to adjacent bones, joints, soft tissue swelling, or hematomas. The types of extremity vascular traumas evaluated included damaged arteries, artery spasms or block, blood vessels shifted because of pressure, pseudo aneurysms, arteriovenous fistula, and vein occlusion. The results of the study indicated that 64-row helical CTA could be highly efficient and accurate in the evaluation of extremity vascular traumas, and could aid in making clinical assessments.

Transverse mode control in high-contrast grating VCSELs.

This paper presents an extensive numerical analysis of a high-contrast grating VCSEL emitting at 0.98 µm. Using a three-dimensional, fully vectorial optical model, we investigate the influence of a non-uniform grating with a broad range of geometrical parameters on the modal behavior of the VCSEL. Properly designed and optimized, the high-contrast grating confines the fundamental mode selectively in all three dimensions and discriminates all higher order modes by expelling them from its central region. This mechanism makes single mode operation possible under a broad range of currents and could potentially enhance the single-mode output power of such devices. The high-contrast grating design proposed here is the only design for a VCSEL with three-dimensional, selective, optical confinement that requires relatively simple fabrication.

Three-dimensional visualization of rat brain microvasculature following permanent focal ischaemia by synchrotron radiation.

OBJECTIVE: Identifying morphological changes that occur in microvessels under both normal and ischaemic conditions is crucial for understanding and treating stroke. However, conventional imaging techniques are not able to detect microvessels on a micron or sub-micron scale without angiography. In the present study, synchrotron radiation (SR)-based X-ray in-line phase contrast imaging (ILPCI) was used to acquire high-resolution and high-contrast images of rat brain tissues in both normal and ischaemic states. METHODS: ILPCI was performed at the Shanghai Synchrotron Radiation Facility, Shanghai, China, without the use of contrast agents. CT slices were reformatted and then converted into three-dimensional (3D) reconstruction images to analyse subtle details of the cerebral microvascular network. RESULTS: By using ILPCI, brain vessels up to 11.8 µm in diameter were resolved. The number of cortical and penetrating arteries detected were found to undergo a remarkable decrease within the infarct area. 3 days after permanent ischaemia, vascular masses were also observed in the peripheral region of the infarcts. CONCLUSION: SR-based ILPCI-CT can serve as a powerful tool to accurately visualize brain microvasculature. The morphological parameters of blood vessels in both CT slices and 3D reconstructions were determined, and this approach has great potential for providing an effective diagnosis and evaluation for rehabilitation therapy for stroke. ADVANCES IN KNOWLEDGE: In the absence of contrast agent, the 3D morphologies of the brain microvasculature in normal and stroke rats were obtained using SR-based ILPCI. SR imaging is a sensitive and promising method which can be used to explore primary brain function.

Summer Blight of White Clover (Trifolium repens) Caused by Rhizoctonia solani AG-1-IB in China.

White clover (Trifolium repens), also known as Dutch clover (family Fabaceae), is an herbaceous, perennial plant, widely planted as pasture crop and occasionally used as lawn plant. From June to September in 2012 and 2013, approximately 5 to 8% of the plants as garden lawn were infected in the areas surveyed in Tonghua County, Jinlin Province. Ash green and water-soaked lesions appeared initially on the petiole and leaves. Subsequently, petioles collapsed with soft watery rot followed by collapse of leaves and eventually the entire plant. Aerial hyphae appeared on all the infected parts, followed by production of light brown to brown sclerotia. Seven isolates with the morphological characteristics of Rhizoctonia solani Kühn were isolated from symptomatic petioles and leaves which were surface disinfested in 70% alcohol for 30 s and 0.5% sodium hypochlorite for 1 min and plated on potato dextrose agar (PDA). Hyphal tips were transferred to a fresh plate of PDA and the cultures were examined for morphological characters microscopically. Mycelia of all isolates were branched at right angles with a septum near the branch and a slight constriction at the branch base. Hyphal cells were determined to be multinucleate when stained with 1% safranin O and 3% KOH solution (1) and examined at 400× magnification with a microscope. The internal transcribed spacer (ITS) region of the nuclear rDNA was amplified by using the primers ITS4 and ITS5 (2). The ITS sequences of isolates BSYJ14 (GenBank Accession No. HF678123), BSYJ31 (HF571130), BSYY21 (HF678126), and BSY22 (KC572140) exhibited 100% identity with that of R. solani AG 1-1B (AB122138 and HQ185364). ITS sequences of another three isolates, BSYJ11 (HF678122), BSYJ32 (HF678125), and BSYJ12 (HF678121) exhibited 99% identity with the ITS sequence of R. solani AG 1-1B. Pathogenicity tests were performed on healthy, potted T. repens. Five potted plants were inoculated at the base of the petiole with a 0.6-cm diameter mycelial plug from 3-day-old PDA cultures for each isolate, and the inoculation sites were covered with moistened sterile absorbent cotton. Another five potted plants were inoculated with sterile PDA plugs as controls. All plants in the experiments were covered with plastic bags and kept in a greenhouse at 20 to 25°C for 72 h, then the plastic bags were removed. After 5 to 7 days, the symptoms of watery rot were observed on petioles and leaves of all plants inoculated with these isolates, while control plants remained healthy. R. solani AG 1-IB was re-isolated from all plants inoculated with the isolates. The isolates were confirmed by morphological characteristics of the hyphae and hyphal fusions with the original isolates. The pathogenicity test was carried out twice with similar results. R. solani has been reported to cause root rot on T. pratense in northwestern China (4) and summer blight on T. pratense in Japan (3). To our knowledge, this is the first report of R. solani AG 1-IB causing summer blight on T. repens in China. References: (1) R. J. Bandoni. Mycologia 71:873, 1979. (2) D. E. L. Cooke et al. Mycol. Res. 101:667, 1997. (3) T. Tsukiboshi et al. Bull. Natl. Grassl. Res. Inst. 39:50, 1988. (4) W.-Y. Zhuang (ed.). Fungi of Northwestern China. Mycotaxon Ltd. Ithaca, NY, 2005.

Veronica sibirica Leaf Spots Caused by Phacellium veronicae, a New Disease in China.

Veronica sibirica (Veronicastrum sibiricum) is an erect perennial herb, an ornamental, and a traditional Chinese medicine plant distributed mostly in northeastern, northern, and northwestern China. It has dehumidifying and detoxifying properties, and is mainly used for the treatment of cold, sore throat, mumps, rheumatism, and insect bites (4). In June 2008 through 2012, leaf spots of V. sibirica were observed in the Medicinal Herb Garden of Jilin Agricultural University (43°48'N, 125°23'E) and the medicinal plantations of Antu County (43°6'N, 128°53'E), Jilin Province. Leaf spots were amphigenous, subcircular, angular-irregular, brown, and 1 to 10 mm in diameter; they occasionally merged into a larger spot with an indefinite margin or with a pale center and dark border. Pale conidiomata were hypophyllous and scattered on the spots. The conidiophores were 100 to 400 µm high and clustered together to form synnemata 20 to 50 µm in diameter, which splayed out apically and formed loose to dense capitula. Conidiophores occasionally emerged through the stomata individually and produced conidia on the surface of the infected leaves. The conidiogenous cell terminal was geniculate-sinuous with somewhat thickened and darkened conidial scars. Conidia were solitary or catenulate, ellipsoid-ovoid or subcylindric-fusiform, hyaline and spinulose, 4.01 to 7.18 × 11.16 to 20.62 µm with obtuse to somewhat attenuated ends, and slightly thickened, darkened hila. Six isolates were obtained from necrotic tissue of leaf spots and cultured on potato dextrose agar at 25°C. After incubation for 14 days, colony surfaces were white to pinkish. The colony diameter increased by 12 mm after 21 days' incubation. Hyphae were hyaline, septate, and branched. Conidiophores grew individually or fascicularly. The symptoms and morphological characteristics were consistent with previous descriptions (1,2), and the fungus was identified as Phacellium veronicae (Pass.) (U. Braun 1990). The internal transcribed spacer (ITS) region of the nuclear rDNA was amplified using primers ITS4/ITS5 (3). The ITS was identical among all six isolates (HE995799) and 98% identical to that of P. veronicae (JQ920427, HQ690097). Pathogenicity was confirmed by spraying five 1-year-old V. sibirica seedlings with a conidial suspension (106 conidia/ml) of each isolate and five seedlings with sterile water as a control treatment. Plants were grown in the greenhouse at 20 to 25°C and were covered with plastic bags to maintain humidity on the foliage for 72 h. After 15 days, the same symptoms appeared on the leaves as described earlier for the field-grown plants; the control plants remained healthy. The same fungus was reisolated from the leaf spots of inoculated plants. Currently, the economic importance of this disease is limited, but it may become a more significant problem, as the cultivated area of V. sibirica is increasing. To our knowledge, although P. veronicae was recorded on the other species of Veronica (V. austriaca, V. chamaedrys, V. grandis, V. longifolia, V. paniculata, and V. spicata ssp. incana) in Europe (Germany, Denmark, Ireland, Romania) and V. wormskjoldii in North America (Canada) (1), this is the first report of V. sibirica leaf spots caused by P. veronicae in the world, and it is a new disease in China. References: (1) U. Braun. A monograph of Cercosporella, Ramularia and allied genera (phytopathogenic Hyphomycetes) 2, IHW-Verlag, Germany, 1998. (2) U. Braun. Nova Hedwigia 50:499, 1990. (3) D. E. L. Cooke et al. Mycol. Res. 101:667, 1997. (4) Jiangsu New Medical College. Dictionary of Chinese Materia Medica. Shanghai: Shanghai Scientific and Technical Publishers, China, 1977.

First Report of Daylily Leaf Streak Caused by Kabatiella microsticta in China.

Daylily (Hemerocallis spp.) is an herbaceous, perennial plant, cultivated for its flowers. Daylily is sold in Asian markets as fresh or dried flowers (the flowers of some species, e.g., Hemerocallis citrina, are edible) or as the corm, which is used for medicinal purposes. In June 2011, daylily leaf streak was found in a nursery of Jilin Agricultural University, Jilin Province, China. Symptoms included water-soaked, irregular spots along the leaf midvein that turned orange to reddish brown and eventually enlarged to coalesce into extensive, necrotic streaks along the length of the leaf, as previously reported (2). Heavily infected leaves often withered and died. Four isolates were recovered from necrotic tissue of leaf spots and cultured on potato dextrose agar (PDA) at 25°C. All colonies were initially cream to peach colored and appeared slimy. With the maturation of the culture, the colonies became dark brown to black with sparse aerial hyphae. Blastic conidia formed simultaneously on intercalary or terminal, undifferentiated conidiogenous cells, and were scattered in dense sections on culture surface. When the conidia dropped from conidiogenous cell, an indistinct scar or a denticle remained. Conidia were hyaline, one-celled, smooth, ellipsoidal, and variable in size (2.73 to 6.01 × 8.45 to 19.36 µm), and all morphological characteristics were consistent with Kabatiella microsticta Bubak (syn. Aureobasidium microstictum; 2,4). The internal transcribed spacer (ITS) region of the nuclear rDNA was amplified using primers ITS4/ITS5 (1). ITS (534 bp) was identical among all four isolates (GenBank Accession No. HE798117) and 100% identical to that of K. microsticta CBS 114.64 (FJ150873). Pathogenicity was confirmed by spraying 20 seedlings of daylily, propagated in tissue-culture medium, with a conidial suspension (106 conidia/ml) of each isolate. A second set of 20 seedlings was sprayed with the same volume of sterile water as the noninoculated control treatment. Plants were grown in the greenhouse at 20 to 25°C and were covered with plastic bags to maintain humidity on the foliage for 72 h. After 5 days, the foliar symptoms described earlier for the field plants appeared on the leaves, whereas the control plants remained healthy. K. microsticta was reisolated from the leaf spots of all 20 inoculated plants. Leaf streak is the most destructive disease of daylily, and was previously reported in Japan and the United States (Illinois, Kentucky, Mississippi, Louisiana, Pennsylvania, Maryland, Virginia, Florida, North Carolina, and Georgia) (3). To our knowledge, this is the first report of the disease caused by K. microsticta in China. References: (1) D. E. L. Cooke et al. Mycol. Res. 101:667, 1997. (2) E. J. Hermanides-Nijhof. Stud. Mycol. 15:153, 1977. (3) R. M. Leahy et al. Plant Pathology Circular No. 376, 1996. (4) P. Zalar et al. Stud. Mycol. 61:21, 2008.