First Report of Leaf Spot Caused by Alternaria alternata on Chard (Beta vulgaris var. cicla L.) in China.

Chard (Beta vulgaris var. cicla L.) is popular vegetable in China. In June 2023, a leaf spot disease was observed on Chard plants in Hunan Province (27°46'10.99″N, 112°05'52.80″E), China. The disease incidence was 30% in a surveyed of about 500 plants. Symptoms began as many light brown round- to polygon-shaped spots on chard leaves, then developed and enlarged into grayish-white lesions, with the edge of the spots brown to dark brown. A total of 10 symptomatic samples were randomly collected. To identify the pathogen, symptomatic tissues (0.5 × 0.5 cm) from the lesion margin surface were sterilized with 75% ethanol for 30 s and 2% NaClO for 1 min, rinsed 3 times with sterile water, air dried. The sterile pieces were placed on potato dextrose agar (PDA) and incubated at 25°C. A total of nine isolates were obtained. Fungal colonies cultured on potato carrot agar (PCA) were almost the same as each other, and two representative isolates (TC0, TC10) were used for further identification. On PCA, the fungal hyphae were initially white and finally gray-brown with flocculent aerial mycelia. Conidia were solitary or in chains, with various shapes, mostly subglobose, the size was 13.2 to 28.0 µm long and 5.8 to 13.0 µm wide (n = 30). The cultural and morphological characteristics of isolates were similar to those of Alternaria sp (Simmons et al. 2007). For molecular identification, four loci, ITS (White et al. 1990), RPB2 (O'Donnell, 2022), H3 (Zheng et al. 2015), and GAPDH (Berbee et al. 1999), were sequenced from two representative isolates (TC0, TC10). Compared with a reference isolate, Alternaria alternata strain CBS 107.27, GenBank accession nos. KP124300.1 (ITS), KP124768.1 (RPB2), KP124157.1 (GAPDH). The ITS, RPB2, and GAPDH sequences of TC0 and TC10 showed 99% (502 of 504 bp ), 100% (753 of 753 bp), and 99% (560 of 561 bp) similarity, respectively. Compared with a reference isolate, A. alternata isolate 21-5, GenBank accession no. MN840996.1 (H3), H3 sequences of TC0 and TC10 showed 99% (399 of 401 bp) similarity. The sequences of two isolates (TC0, TC10) were deposited in GenBank with accession numbers PP837733.1, PP565404.1(ITS), PP839298.1, PP573905.1(RPB2), PP839299.1, PP573904.1 (GAPDH), and PP839297.1, PP573903.1(H3). Phylogenetic trees were constructed using the sequences and showed that isolates (TC0, TC10) were in the same clade with A. alternata strains. TC0 and TC10 were identified as A. alternata based on the morphological characteristics and molecular phylogeny. Pathogenicity testing was conducted on six-month-old healthy plants, (cv. Green Stalk), three plants were inoculated by spraying spore solution (1 × 106 conidia/mL), and three plants were sprayed with sterile water as a control. The pathogenicity test was performed 3 times. Plants were maintained at 28°C and >80% RH. Plants showed symptoms after 30 days, symptoms were observed similar to those of the original infected plants, control plants were asymptomatic. The fungus was reisolated, confirmed as A. alternata based on conidial characteristics, no pathogenic fungus was isolated from the control plants. A. alternata has previously been reported on beet (also Beta vulgaris) in China (Tai, F. L. 1979; Zhuang, W. Y. 2005). To our knowledge, this is the first report of leaf spot caused by A. alternata on chard in China. This result may expand the etiological study of A. alternata and the control strategy of Chard leaf spot.

[Clinical characteristics of biliary obstruction patients with Clonorchis sinensis infections].

OBJECTIVE: To analyze the clinical characteristics of biliary obstruction patients with Clonorchis sinensis infections, so as to provide insights into the clinical diagnosis and therapy of this comorbidity. METHODS: A total of 45 biliary obstruction patients with C. sinensis infections that were admitted to the Second Affiliated Hospital of Harbin Medical University from October 2012 to September 2019 were enrolled, and 45 patients with biliary obstruction alone at the hospital during the same study period were recruited as controls. Univariate analysis was performed to analyze the epidemiological characteristics, clinical manifestations, laboratory examination results and imaging manifestations related to C. sinensis infection, and the statistically significant univariate was used as an independent variable for multivariate logistic regression analysis. RESULTS: Compared with biliary obstruction alone, males (91.11% vs. 46.67%; χ2 = 20.737, P < 0.01) and rural areas (62.22% vs. 22.22%; χ2 = 14.757, P < 0.01) showed a significantly higher proportion in biliary obstruction patients with C. sinensis infections. The major clinical symptoms involved in had jaundice (45 cases, 100%) and abdominal pain (40 cases, 88.89%) in C. sinensis-infected patients. Logistic regression analysis showed that males [odds ratio (OR) = 10.717, 95% confidential interval (CI): (2.571, 44.662)] and drinking alcohol [OR = 4.474, 95% CI: (1.019, 19.642)] were risk factors for biliary obstruction patients with C. sinensis infections, while living in city [OR = 0.128, 95% CI: (0.038, 0.435)] was a protective factor. Additionally, in biliary obstruction patients with C. sinensis infections, lower total bilirubin (Z = -2.566, P <0.05) and direct bilirubin (Z = -3.454, P <0.05), higher indirect bilirubin (Z = -3.821, P < 0.05), thickening of the bile duct wall and dilatation of the intrahepatic bile duct were detected. CONCLUSIONS: A comprehensive diagnosis requires to be made based on clinical symptoms, laboratory and imaging examinations, in order to improve the diagnosis of biliary obstruction patients with C. sinensis infections.

Manual contouring based volumetric evaluation for colorectal cancer with liver limited metastases: a comparison with RECIST.

BACKGROUND: To compare response evaluation criteria in solid tumours (RECIST) and volumetric evaluation (VE) for colorectal cancer with liver-limited metastasis. PATIENTS AND METHODS: VE of liver metastases was performed by manual contouring before and after chemotherapy on 45 pairs of computed tomography (CT) images in 36 patients who suffered from metastatic colorectal cancer (mCRC) with liver metastasis only. Cohen kappa was used to compare the agreement between VE and RECIST. Pearson correlation was performed for their comparison after cubic root transformation of the aggregate tumor volumes. Logistic regression was done to identify clinical and radiographic factors to account for the difference which may be predictive in overall response (OR). RESULTS: There were 16 partial response (PR), 23 stable disease (SD) and 6 progressive disease (PD) cases with VE, and 14 PR, 23 SD and 8 PD with RECIST. VE demonstrated good agreement with RECIST (κ=0.779). Discordant objective responses were noted in 6 pairs of comparisons (13.3%). Pearson correlation also showed excellent correlation between VE and RECIST (r2=0.966, p<0.001). Subgroup analysis showed that VE was in slightly better agreement with RECIST for enlarging lesions than for shrinking lesions (r2=0.935 and r2=0.780 respectively). No factor was found predictive of the difference in OR between VE and RECIST. CONCLUSIONS: VE exhibited good agreement with RECIST. It might be more useful than RECIST in evaluation shrinking lesions in cases of numerous and conglomerate liver metastases.

Mc-hES, a novel plasmid carrying human endostatin gene, inhibits nasopharyngeal carcinoma growth.

Conventional plasmids for gene therapy produce low-level and short-term gene expression. Here, we first created minicircle carrying endostatin (mc-hES) for measurement of transfection efficiency. Compared with pcDNA-hES, MC-mediated endostatin gene transfer in vitro resulted in seven-fold greater endostatin expression levels in transfected cells and inhibited the growth of Human umbilical vein endothelial cells (HUVEC) more efficiently. HUVEC cell migration and tube-formation assays suggested that MC-mediated endostatin gene has significant anti-migration and anti-tube-formation capacity than that in pcDNA-hES. In vivo experiments showed that after transfection, mc-hES inhibited the growth of nasopharyngeal carcinoma xenografts. The tumor inhibition rates of mc-hES and pcDNA-hES were 60.8% and 26.9%, respectively (P<0.05). MC-mediated intratumoral endostatin expression in vivo was 2.2-17.9 times higher than pcDNA-hES in xenografted mice and lasted for 20 days. Our results suggest that minicircle DNA vectors might be a promising vector for biotherapy and should be further investigated.

Detection of let-7a microRNA by real-time PCR in colorectal cancer: a single-centre experience from China.

Colorectal cancer, a heterogeneous disease arising from a complex series of molecular changes, is one of the world's leading causes of cancer deaths. MicroRNAs (miRNAs), an extensive class of small non-coding RNAs, have been implicated in cancer development and progression. One of the first miRNAs to be identified was let-7 miRNA, which has recently been found to be expressed at reduced levels in human lung cancer cells. We used a rapid stem-loop reverse transcription polymerase chain reaction method to quantify human let-7a miRNA expression in samples of human colorectal cancer. This method was able to detect let-7a miRNA in as little as 0.05 ng of total RNA from colorectal mucosa and its specificity was high (100%). Our results showed that the expression of let-7a miRNA was considerably reduced in two of eight patients. To our knowledge, this is the first study of Chinese patients to show reduced expression of endogenous let-7 miRNA in colorectal cancer.

Phase II trial of a 2-h infusion of gemcitabine plus carboplatin as first-line chemotherapy for advanced non-small-cell lung cancer.

PURPOSE: To evaluate the efficacy and safety of the combination of using gemcitabine as a rate infusion of 10 mg/m(2) per min with carboplatin in front-line chemonaive patients with advanced non-small-cell lung cancer (NSCLC). PATIENTS AND METHODS: Fifty-four chemonaive patients with stage IIIB or IV NSCLC have been included, 44 males and 10 females, with a median age 63 years (range 19-75). Thirty-two (59%) patients had adenocarcinoma, 13 (24%) squamous cell, 1 (2%) large cell carcinoma and 8 (15%) others. Eight (15%) had stage IIIB and 46 (85%) stage IV. Treatment was consisted of 1,200 mg/m(2) gemcitabine given as a 2-h continuous infusion (10 mg/m(2) per min) on days 1 and 8 of each cycle an AUC 5 carboplatin as on day 1, repeating each cycle for every 21 days. A total of 223 chemotherapy cycles were administered, with a median of four cycles per patient (range 1-6), and 15 (28%) patients received all six cycles. RESULTS: Of the 54 patients enrolled, all were evaluated for toxicity and 51 assessed for response. The overall response rate was 41% (95% confidence interval, 28-57%) with complete and partial responses of 4 and 37%, respectively. The median time to disease progression was 5.0 months (95% CI, 3.7-6.3 months), and median overall survival time was 11.5 months (95% CI, 9.9-13.1 months). One-year survival was 42%. The main grade 3-4 toxicity (according to the WHO scale) consisted of neutropenia (56%) and thrombocytopenia (57%). Patients were required platelet transfusion in 27 cycles (12%) and hematopoietic growth factors support care in 56 (25%) cycles. No bleeding episodes were recorded. Grade 3 nausea/vomiting occurred in 6% and grade 1-2 skin rash occurred in 43%. CONCLUSIONS: Prolonged gemcitabine infusion combined with carboplatin is manageable and tolerated, and its efficacy is similar to that of other chemotherapeutic schemes used for NSCLC treatment.

Optimization and characterization of chitosan-coated alginate microcapsules containing albumin.

In order to obtain small microcapsules with high protein encapsulation efficiency and extended release characteristics various processing factors were studied. Bovine serum albumin-loaded alginate microcapsules were prepared by an emulsion method and further incubated in chitosan. Many process factors were tested including the concentration and molecular weight of alginate, the concentration and pH of chitosan, and surfactants, etc. Microcapsules were achieved with diameters less than 2 microm, high encapsulation efficiency (> 80%) and high loading rate (> 10% w/w). The results also showed that the initial BSA amount of 20%-30% loaded alginate microcapsules coated with 0.2%-0.5% chitosan solutions at pH 4 by the two-stage procedure present the best sustained releasing characteristics.

A heparin-binding growth factor secreted from breast cancer cells homologous to a developmentally regulated cytokine.

We report purification of an 18-kDa heparin-binding growth factor secreted from human cancer cells which is homologous to a developmentally regulated, neurotrophic factor, heparin-binding growth-associated molecule/pleiotrophin (HB-GAM/PTN; Merenmies, J., and Rauvala, H. (1990) J. Biol. Chem. 265, 16721-16724; Li, Y. S., Milner, P. G., Chauhan, A. K., Watson, M. A., Hoffman, R. M., Kodner, C. M., Milbrandt, J., and Deuel, T. F. (1990) Science 250, 1690-1694). We have purified the protein from tissue culture supernatants of human breast cancer cells (MDA-MB 231) and have used soft agar cloning of an epithelial cell line (SW-13) to detect its growth stimulating activity. A 32,000-fold purification was achieved by isoelectric focusing, heparin affinity chromatography, and reversed phase high pressure liquid chromatography. The molecular mass of the protein was confirmed by gel filtration chromatography in the presence of detergent and bioassay of the fractions. The N-terminal sequence was homologous to HB-GAM/PTN, and polymerase chain reaction amplification and DNA sequencing confirmed that the respective transcript was present in the cancer cells. We conclude that HB-GAM/PTN can function as a tumor growth factor in addition to its role during neuronal development.