Cytomorphological characteristics of low-grade papillary urothelial carcinoma for differential diagnosis from benign papillary urothelial lesions: logistic regression analysis in SurePath(™) liquid-based voided urine cytology.

OBJECTIVE: The diagnosis of low-grade papillary urothelial carcinoma (LGPUC) in urine cytology specimens is challenging because of its subtle, minimally atypical findings. Furthermore, as SurePath(™) liquid-based cytology (LBC) is becoming a widely used method in urine cytology, the inevitable cytomorphological alterations resulting from this technique call for new morphological diagnostic criteria in LGPUC. METHODS: Logistic regression analysis was carried out on SurePath slides from surgically proven voided urine specimens. The study was designed to include a test set (n = 141) and a validation set (n = 61), and evaluated significant discriminative parameters between LGPUC and benign papillary urothelial neoplasm (BPUN). RESULTS: Of the seven cytological findings that were found to have statistical significance in univariate analysis, five were found to be independent variables: loss of polarity of papillaroid clusters, irregular contours, absence of columnar cells, hobnail features and hyperchromasia. These independent variables had an area under the curve (AUC) of 0.781. CONCLUSIONS: The distinctive cytological criteria identified above may prove to be helpful in cases in which other conventional criteria for LGPUC are insufficient for diagnosis.

Lack of positive effect of intravitreal bevacizumab in central serous chorioretinopathy: meta-analysis and review.

PURPOSE: To review and evaluate the effects of intravitreal bevacizumab injection (IVB) in centralserous chorioretinopathy (CSC) by meta-analysis. PATIENTS AND METHODS: Clinical controlled studies that evaluated the effect of IVB in CSC were identified through systematic searches of Embase, PubMed, and the Cochrane Central Register of Controlled Trials. Data on the best-corrected visual acuity (BCVA) in logMAR and central macular thickness (CMT) in µm at baseline and 6 months after IVB were extracted and compared with those treated by simple observation. RESULTS: Four clinical controlled studies were included in the meta-analysis. The IVB injection group achieved better BCVA at a follow-up of 6 months. However, the analysis showed that there were no significant differences of BCVA at 6 months after injection between IVB group and the observation group (-0.02 logMAR, 95% CI -0.14 to 0.11, P=0.80). The analysis of the reduction in CMT revealed that the difference between groups was not statistically significant (-8.37 µm, 95% CI -97.26 to 80.52, P=0.85). No report assessed severe complications or side effects of IVB in patients with CSC. CONCLUSIONS: Meta-analysis failed to verify the positive effect of IVB in CSC based on the epidemiological literature published to date.

Cloning of a Family 11 Xylanase Gene from Bacillus amyloliquefaciens CH51 Isolated from Cheonggukjang.

Bacillus amyloliquefaciens CH51, an isolate from cheonggukjang, Korean fermented soyfood, secretes several enzymes into culture medium. A gene encoding 19 kDa xylanase was cloned by PCR. Sequencing showed that the gene encoded a glycohydrolase family 11 xylanase and named xynA. xynAHis, xynA with additional codons for his-tag, was overexpressed in Escherichia coli BL21(DE3) using pET-26b(+). XynAHis was purified using HisTrap affinity column. Km and Vmax of XynAHis were 0.363 mg/ml and 701.1 µmol/min/mg, respectively with birchwood xylan as a substrate. The optimum pH and temperature were pH 4 and 25 °C, respectively. When xynA was introduced into Bacillus subtilis WB600, active XynA was secreted into culture medium.

Blood pressure changes after intravitreal bevacizumab in patients grouped by ocular pathology.

PURPOSE: We evaluated the effects of intravitreal injection of bevacizumab (Avastin; Novartis, Basel, Switzerland) on blood pressure (BP) in the context of ocular vascular pathology. METHODS: This study retrospectively examined 135 consecutive patients treated with intravitreal injections of 1.25 mg bevacizumab for retinal vascular disease; there were 61 cases of diabetic retinopathy, 30 of retinal vein occlusion, 35 of choroidal neo-vascularization (CNV), and 9 of other retinal vascular diseases. BP was measured before injection and at 30 min, 1 day, 1 week, 3 weeks, and thereafter monthly over a 6-month period. RESULTS: In the CNV group, 30-min post-injection systolic values were significantly higher than baseline, and systolic and diastolic values after 1 day, 1 week, and 3 weeks were significantly lower than before injection. No other pressure measurement differed significantly from baseline values in the other groups. DISCUSSION: Intravitreal bevacizumab injection is safe in terms of its effect on BP, regardless of ocular pathology.

Expression of exo-polygalacturonases in Botrytis cinerea.

The pathogenic fungus, Botrytis cinerea, causing gray mold disease in a variety of plant species, secretes at least four polygalacturonases (PGs), cell wall degrading enzymes. Among them, we prepared polyclonal antibody against purified 66-kDa exo-PG in rabbit. Immunoblot analysis revealed that the antibody recognized two exo-PGs, 66 kDa and 70 kDa in molecular mass, secreted from B. cinerea cultured in the medium containing citrus pectin as a carbon source. By immunohistochemical analysis, the expression of exo-PGs was identified in cucumber leaves inoculated with spores of B. cinerea. The exo-PGs were observed 9 h after inoculation, and the amount of exo-PGs increased with time in the leaves. The exo-PGs were induced by polygalacturonic acid as well as its monomer, galacturonic acid, in vitro. The expression of 66-kDa exo-PG (exo-PG I) increased with time of culture, while 70-kDa exo-PG (exo-PG II) was transiently expressed soon after the start of culture. Therefore, exo-PGs might play an important role in pathogenesis at an early stage of infection as well as in tissue maceration of host plant.

Paenibacillus koreensis sp. nov., a new species that produces an iturin-like antifungal compound.

A bacterial strain, YC300T, that produces an iturin-like antifungal antibiotic was isolated from compost and identified as member of the genus Paenibacillus. Gram reaction of the strain was variable depending upon growth stages and culture media. Three different types of colonies were developed on tryptic soy agar. The organism was facultatively anaerobic and grew at 50 degrees C. The DNA G+C content was 54 mol % and anteiso-C15:0 was the major fatty acid. A 0.9 kb fragment was produced by PCR amplification of strain YC300T DNA using primers PAEN515F and 1377R. Levels of 16S rDNA similarity between strain YC300T and other Paenibacillus species were between 89.8 and 94.8%. Phylogenetically, strain YC300T formed a significant monophyletic clade with Paenibacillus validus. It is clear from polyphasic evidence that the isolate should be classified as Paenibacillus koreensis sp. nov., the type strain of which is YC300T (= KCTC 2393T, KCCM 40903T).

Kitasatospora cheerisanensis sp. nov., a new species of the genus Kitasatospora that produces an antifungal agent.

An actinomycete, strain YC75T, which produced bafilomycin-like antifungal compounds, was identified as a member of the genus Kitasatospora on the basis of morphological and chemotaxonomic characteristics. The strain produced the aerial and fragmenting vegetative mycelia consisting of straight chains of 20 or more smooth-surfaced spores. Submerged spores were formed in tryptic soy broth. No soluble pigments were formed. Whole-cell hydrolysates contained glucose and mannose, but not galactose. The 16S rDNA sequence of YC75T was compared with those of the other representative kitasatosporae and streptomycetes. Strain YC75T formed a significant monophyletic clade with Kitasatospora phosalacinea. The levels of DNA relatedness between strain YC75T and representatives of the genus Kitasatospora ranged from 16 to 59% including K. phosalacinea (28 and 40%). It is clear from polyphasic evidence that the isolate should be classified as Kitasatospora cheerisanensis sp. nov., whose type strain is YC75T (= KCTC 2395T). The presence of galactose in whole-cell hydrolysates may not be a stable chemical marker for the genus Kitasatospora.

Biological control of fusarium wilt of cucumber by chitinolytic bacteria.

ABSTRACT Two chitinolytic bacterial strains, Paenibacillus sp. 300 and Streptomyces sp. 385, suppressed Fusarium wilt of cucumber (Cucumis sativus) caused by Fusarium oxysporum f. sp. cucumerinum in nonsterile, soilless potting medium. A mixture of the two strains in a ratio of 1:1 or 4:1 gave significantly (P < 0.05) better control of the disease than each of the strains used individually or than mixtures in other ratios. Several formulations were tested, and a zeolite-based, chitosan-amended formulation (ZAC) provided the best protection against the disease. Dose-response studies indicated that the threshold dose of 6 g of formulation per kilogram of potting medium was required for significant (P < 0.001) suppression of the disease. This dose was optimum for maintaining high rhizosphere population densities of chitinolytic bacteria (log 8.1 to log 9.3 CFU/g dry weight of potting medium), which were required for the control of Fusarium wilt. The ZAC formulation was suppressive when added to pathogen-infested medium 15 days before planting cucumber seeds. The formulation also provided good control when stored for 6 months at room temperature or at 4 degrees C. Chitinase and beta-1,3-glucanase enzymes were produced when the strains were grown in the presence of colloidal chitin as the sole carbon source. Partial purification of the chitinases, followed by sodium dodecyl sulfate polyacrylamide gel electrophoresis and activity staining, revealed the presence of five bands with molecular masses of 65, 62, 59, 55, and 52 kDa in the case of Paenibacillus sp. 300; and three bands with molecular masses of 52, 38, and 33 kDa in the case of Streptomyces sp. 385. Incubation of cell walls of F. oxysporum f. sp. cucumerinum with partially purified enzyme fractions led to the release of N-acetyl-D-glucosamine (NAGA). NAGA content was considerably greater when pooled enzyme fractions (64 to 67) from Paenibacillus sp. were used, because they contained high beta-1,3-glucanase activity in addition to chitinase activity. Suppression of Fusarium wilt of cucumber by a combination of these two bacteria may involve the action of these hydrolytic enzymes.

Identification of putative phosphoinositide-specific phospholipase C genes in filamentous fungi.

Five putative phosphoinositide-specific phospholipases C (PLC) genes were identified in three species of filamentous fungi. Using polymerase chain reaction with degenerate oligonucleotide primers, gene fragments encoding amino acid sequences homologous to PLCs of mammals and other organisms were amplified: one sequence from Botryotinia fuckeliana, one from Aspergillus nidulans, and three from Neurospora crassa. The molecular cloning and sequencing of a putative PLC gene (BCPLC1) from B. fuckeliana showed that it encoded a polypeptide containing X and Y domains, the two conserved regions found in all known PLCs. The hypothetical gene product of BCPLC1 was of delta type in its primary structural organization. The identification of three PLC genes in N. crassa shows that multiple PLC isozymes also occur in microbial eukaryotes.

Isolation and characterization of a benomyl-resistant form of beta-tubulin-encoding gene from the phytopathogenic fungus Botryotinia fuckeliana.

As an initial step to develop a DNA-mediated transformation system using benomyl resistance as a dominant selectable marker in the phytopathogenic fungus Botryotinia fuckeliana (anamorph, Botrytis cinerea), we have constructed a phage lambda genomic DNA library of a benomyl-resistant strain 91T-1, and a beta-tubulin-encoding gene benA was isolated, cloned and sequenced. Southern blot analysis suggested that a single copy of benA is present in the genome of B. fuckeliana. The benA gene is composed of seven exons which are separated by six introns of 52 to 135 bp. The intron consensus sequences are similar to those of other fungal genes. The deduced amino acid sequence (447 amino acid residues) is highly homologous to those of other fungal beta-tubulin-encoding genes. Comparison of the sequences around codons 198 and 200 in benomyl-resistant and sensitive strains revealed that the benAHR allele from the benomyl-resistant strain 91T-1 contained a mutation at codon 198 from GAG (Glu) to GCG (Ala), which has been correlated with high resistance to benzimidazole fungicides including benomyl in various filamentous fungi.