Synchrospora gen. nov., a New Peronosporaceae Genus with Aerial Lifestyle from a Natural Cloud Forest in Panama.

During a survey of Phytophthora diversity in Panama, fast-growing oomycete isolates were obtained from naturally fallen leaves of an unidentified tree species in a tropical cloud forest. Phylogenetic analyses of sequences from the nuclear ITS, LSU and ßtub loci and the mitochondrial cox1 and cox2 genes revealed that they belong to a new species of a new genus, officially described here as Synchrospora gen. nov., which resided as a basal genus within the Peronosporaceae. The type species S. medusiformis has unique morphological characteristics. The sporangiophores show determinate growth, multifurcating at the end, forming a stunted, candelabra-like apex from which multiple (8 to >100) long, curved pedicels are growing simultaneously in a medusa-like way. The caducous papillate sporangia mature and are shed synchronously. The breeding system is homothallic, hence more inbreeding than outcrossing, with smooth-walled oogonia, plerotic oospores and paragynous antheridia. Optimum and maximum temperatures for growth are 22.5 and 25-27.5 °C, consistent with its natural cloud forest habitat. It is concluded that S. medusiformis as adapted to a lifestyle as a canopy-dwelling leaf pathogen in tropical cloud forests. More oomycete explorations in the canopies of tropical rainforests and cloud forests are needed to elucidate the diversity, host associations and ecological roles of oomycetes and, in particular, S. medusiformis and possibly other Synchrospora taxa in this as yet under-explored habitat.

Differences in the Proteomic and Metabolomic Response of Quercus suber and Quercus variabilis During the Early Stages of Phytophthora cinnamomi Infection.

Phytophthora cinnamomi Rands is a cosmopolite pathogen of woody plants which during the last couple of centuries has spread all over the world from its center of origin in Southeast Asia. In contrast to Chinese cork oak (Quercus variabilis Blume) forests native to Asia, which are generally healthy despite the presence of the pathogen, the populations of Cork oaks (Quercus suber L.) in Europe have been severely decimated by P. cinnamomi. The present study aims at identifying the differences in the early proteomic and metabolomic response of these two tree species that lead to their differences in susceptibility to P. cinnamomi. By using micropropagated clonal plants, we tried to minimize the plant-to-plant differences in the defense response that is maximized by the high intraspecific genetic variability inherent to the Quercus genus. The evolution on the content of Phytophthora proteins in the roots during the first 36 h after inoculation suggests a slower infection process in Q. variabilis plants. These plants displayed a significant decrease in sugars in the roots, together with a downregulation of proteins related to carbon metabolism. In the leaves, the biggest changes in proteomic profiling were observed 16 h after inoculation, and included increased abundance of peroxidases, superoxide dismutases and glutathione S-transferases in Q. variabilis plants, which probably contributed to decrease its susceptibility to P. cinnamomi.

In-situ preparation of plant samples in ESEM for energy dispersive x-ray microanalysis and repetitive observation in SEM and ESEM.

The Extended Low Temperature Method (ELTM) for the in-situ preparation of plant samples in an environmental scanning electron microscope enables carrying out repetitive topographical and material analysis at a higher resolution in the vacuum conditions of a scanning electron microscope or in the low gas pressure conditions of an environmental scanning electron microscope. The method does not require any chemical intervention and is thus suitable for imaging delicate structures rarely observable with common treatment methods. The method enables both sample stabilization as close to their native state as possible, as well as the transfer of the same sample from a low vacuum to an atmospheric condition for sample storage or later study. It is impossible for wet samples in the environmental scanning electron microscope. Our studies illustrate the high applicability of the ELTM for different types of plant tissue, from imaging of plant waxes at higher resolution, the morphological study of highly susceptible early somatic embryos to the elemental microanalysis of root cells. The method established here provides a very fast, universal and inexpensive solution for plant sample treatment usable in a commercial environmental scanning electron microscope equipped with a cooling Peltier stage.

Effects of copper and arsenic stress on the development of Norway spruce somatic embryos and their visualization with the environmental scanning electron microscope.

Somatic embryogenesis is an important biotechnological technique which can be used in studies associated with environmental stress. Four embryogenic cell lines of Norway spruce were grown on media enriched with copper and arsenic in concentration ranges 50-500 µM and 10-50 µM, respectively. The effects were observed during subsequent stages of somatic embryogenesis, the characteristics evaluated being proliferation potential, average number of somatic embryos obtained per g/fresh weight, morphology of developed somatic embryos, metal uptake, and microanalysis of macro- and micronutrients uptake. Copper and arsenic at higher concentrations significantly reduced the growth of early somatic embryos. In almost all treatments, the cell line V-1-3 showed the best performance compared with the other lines tested. Environmental scanning electron microscopy was used to visualize and identify morphological abnormalities in the development of somatic embryos. Abnormalities observed were classified into several categories: meristemless somatic embryos, somatic embryos with disrupted meristem, reduced number of cotyledons, single cotyledon and fused cotyledons. With the application of a low temperature method for the environmental scanning electron microscope, samples were stabilized and whole meristems could be investigated in their native state. As far as we are aware, this is the first report of the effect of copper and arsenic during the process of somatic embryogenesis and the first to evaluate the content of macro and micronutrients uptake in Norway spruce.

Bottlenecks in bog pine multiplication by somatic embryogenesis and their visualization with the environmental scanning electron microscope.

Somatic embryogenesis (SE) is an important biotechnological technique used for the propagation of many pine species in vitro. However, in bog pine, one of the most endangered tree species in the Czech Republic, limitations were observed, which negatively influenced the development and further germination of somatic embryos. Although initiation frequency was very low-0.95 %, all obtained cell lines were subjected to maturation. The best responding cell line (BC1) was used and subjected to six different variants of the maturation media. The media on which the highest number of early-precotyledonary/cotyledonary somatic embryos was formed was supplemented with 121 µM abscisic acid (ABA) and with 6 % maltose. In the end of maturation experiments, different abnormalities in formation of somatic embryos were observed. For visualization and identification of abnormalities in meristem development during proliferation and maturation processes, the environmental scanning electron microscope was used. In comparison to the classical light microscope, the non-commercial environmental scanning electron microscope AQUASEM II has been found as a very useful tool for the quick recognition of apical meristem disruption and abnormal development. To our knowledge, this is the first report discussing somatic embryogenesis in bog pine. Based on this observation, the cultivation procedure could be enhanced and the method for SE of bog pine optimized.

[The role of colposcopy and typization of human papillomavirus in further diagnostic proceedings in patients with ASC-US cytological finding of the uterine cervix].

BACKGROUND/AIM: Bethesda system of classification of cytological findings was introduced in 2001 two subcategories in the category of atypical squamous cells (ASC) findings: ASC of undetermined significance (ASC-US) and ASC which cannot exlude high-grade intraepithelial lesions (ASC-H). The aim of our study was to assess a possible association of these two subcategories with pathologic biopsy finding and to find out the best further diagnostic proceedings. METHODS: At the Clinic of Gynecology and Obstetrics, Nis 130 patients with ASC findings were analyzed. Colposcopy was performed in all study participants. Patients with pathological colposcopic findings underwent cervical biopsy. In 10 patients with pathologic histologic and 15 with benign findings human papilloma virus (HPV) typization was done using the Hybrid Capture method. RESULTS: Patients with ASC-H finding had significantly more pathologic biopsies compared with patients with ASC-US finding (57.84: 20.72). CONCLUSION: Colposcopy was exhibited somewhat higher sensitivity compared to HPV typization (94.7 : 90), but lower sensitivity (79.27 : 86.6). The usage of HPV typization in the triage of patients with ASC cytologic smear induces statistically significant reduction of unnecessary percentage of cervical biopsies.

Computer-assisted quantitative analysis of Ki-67 antigen in dysplasia--associated lesions or masses in ulcerative colitis.

BACKGROUND/AIM: The aim of this study was to apply computer-assisted methodology in assessment of Ki-67 positivity in "adenoma-like" dysplasia associated lesions or masses (DALMs), and carcinoma in ulcerative colitis (UC), and to determine a new approach to grading of Ki-67 staining intensity. METHODS: Immunohistochemical slides were quantitatively analyzed for estimation of proportion and intensity of Ki-67 positive-stained cells in a total of 50 "adenoma-like" DALMs (27 with low-grade dysplasia and 23 with high-grade dysplasia), and 17 adenocarcinomas associated with UC. The four grades of immunohistochemical staining intensity were established by an automated classification of nuclear optical densities. RESULTS: The Ki-67 labeling index (LI) in low-grade dysplasia was significantly lower than in high-grade dysplasia, and carcinoma (p < 0.001). The Ki-67 LI of carcinomas was not significantly different from the value obtained in high-grade dysplasia (p > 0.05), however having the difference in percentage values of the moderate stained nuclei (p < 0.05). The overall average values of chromogene nuclear optical density, showed statistically significant differences between DALMs and carcinoma (p < 0.05), although not between normal mucosa and low-grade dysplasia (p > 0.05). CONCLUSION: The obtained results imply, according to the overall percentage of labeled nuclei, that high-grade dysplasia is very close to carcinoma, while there is the difference in the percentage of moderately stained nuclei. We showed that Ki-67 positivity have a different internal distribution which could be useful in analysing these lesions. These findings also, indicate the important biological differences between low-grade dysplasia and carcinoma in UC, and a low proliferative potential of the former. Automated image analysis permits an objective [corrected] estimation of Ki-67 immunohistochemical staining in UC-associated dysplasia and carcinoma.

[Influence of the primary malignant tumor site on the incidence and features of metastatic ovarian tumors].

BACKGROUND/AIM: Ovary is the organ of the female reproductive system most commonly affected by metastases. The aim of the study was to determine the frequency and features of metastatic ovarian tumors (MOT) depending on the site of the primary malignant tumor. METHODS: The study group consisted of 488 patients with histopathologically confirmed ovarian cancers treated at the Clinic of Oncology, Clinical Center Nis, in the period from 1 January 1998 to 31 December 2005. MOT were found in 41 patients. Regarding the site of the primary malignant tumor, those with secondary ovarian tumor were divided into two groups: group A--primary malignant tumor involving the genital organs (n = 30) and group B--primary malignant tumor of extragenital origin (n = 11). RESULTS: MOT were confirmed in 8.40% (41/488) of the patients. Secondary ovarian malignancies were the consequence of endometrial carcinoma spreading in 73.17%, breast carcinoma in 19.51%, stomach carcinoma in 4.88% and colon carcinoma in 2.44% of the cases. No significant differences were found between the group A and group B by the factors of age, body mass index, parity and menopausal status. Contrary to the group A, metastatic tumors in the group B patients were more commonly asymptomatic (p < 0.001), bilateral (p < 0.05), with larger ovarian diameter (p < 0.05), associated with ascites (p < 0.001) and abdominal metastases (p < 0.01), all of statistical significance. CONCLUSIONS: Metastatic tumors made up 8.40% of ovarian neoplasmas. With non-genital primary tumors, secondary ovarian deposits were frequently asymptomatic, bilateral, associated with larger ovarian diameter, ascites and abdominal metastatic deposits, compared to malignant tumors of genital origin.

[Preoperative and postoperative histopathological findings in patients with endometrial hyperplasia].

INTRODUCTION: The aim of this study was to analyze and compare the histopathological findings in curettage and hysterectomy specimens, to evaluate the accuracy of histopathological diagnosis in curettage specimens, and to determine the frequency of coexisting endometrial carcinoma in patients with histopathological diagnosis of endometrial hyperplasia. MATERIAL AND METHODS: Curettage and hysterectomy specimens of 135 female patients with initially diagnosed endometrial hyperplasia were retrospectively analyzed and compared. RESULTS: Simple hyperplasia was found in 49 patients (36.3%), complex hyperplasia in 14 (10.4%), simple atypical hyperplasia in 24 (17.8%), and complex atypical hyperplasia in 48 (35.5%) patients. After hysterectomy, 59 (43.7%) patients were found to have simple hyperplasia, 12 (8.9%) complex hyperplasia, 15 (11.1%) simple atypical hyperplasia, 18 (20.7%) complex atypical hyperplasia, and 21 (15.5%) endometrial carcinoma. The accuracy of histopathological diagnosis of endometrial hyperplasia in curettage specimens was 82.2-89.6% and dependent on the types of hyperplasia. The frequency of coexisting endometrial carcinoma was significantly higher (p < 0.001) in patients with atypical hyperplasia than in patients with hyperplasia without cytological atypia. After hysterectomy, coexisting endometrial carcinoma was found in 27.8% of patients with histopathological diagnosis of atypical hyperplasia in curettage specimens. In contrast to simple atypical hyperplasia, the frequency of coexisting endometrial carcinoma was significantly higher (p < 0.05) in complex atypical hyperplasia. CONCLUSION: The frequency of coexisting endometrial carcinoma in hysterectomy specimens in patients with histopathological diagnosis of atypical hyperplasia in curettage specimens was relatively high and it should be taken into account when planning therapy.

Differential influence of open surgery and sepsis on the circulating insulin-like growth factors and their binding proteins as representative metabolic markers.

OBJECTIVES: In critical illnesses and stress conditions many endocrine systems are disturbed. In the current study we determined the influence of open surgery, post-operative sepsis and its early therapy on the components of the insulin-like growth factor (IGF) system in patients with malignant gastric or pancreatic tumors. DESIGN AND METHODS: Twenty-one patients and eighty-one age- and sex-matched healthy subjects were included in this study. IGF-I, IGF-II, IGF binding proteins (IGFBPs), cortisol, insulin and protein concentrations (total, albumin and IgG) were determined pre-operatively, post-operatively, when sepsis was diagnosed and 48 h after initiating therapy. RESULTS: The concentrations of circulating IGF-I, IGF-II and IGFBP-3 were significantly lower in pre-operative patients compared to healthy subjects. Sepsis caused a further decrease in IGF-I and IGFBP-3 but an increase in IGFBP-1, IGFBP-2 and IGFBP-4 resulting in a redistribution of IGF molecules from ternary to binary complexes. CONCLUSIONS: The presence of malignant gastric or pancreatic tumors followed by post-operative sepsis caused a serious misbalance in components of the IGF system which failed to recover during the time of our longitudinal study.

[Expression of nuclear Ki-67 antigen in prostatic high grade intraepithelial neoplasia and prostatic carcinoma].

BACKGROUND/AIM: Prostatic intraepithelial high grade neoplasia (PINHG) is accepted as preneoplastic lesion in prostatic carcinoma. One of the fundamental events in early oncogenesis is the disruption of proliferative activity. One of the numerous regulatory proteins is Ki-67 expressed in all proliferating cells. Index Ki-67 is considered to have prognostic significance. The aim of the study was to compare the level of proliferation in hyperplastic epithelium, prostatic carcinoma (Gleason score > 6) and PINHG. METHODS: Micromorphological examination was done in 85 patients. Pathohistological analysis was performed on standard histologic specimens with the estimation of Gleason score and the presence of PINHG in its surroundings. Nuclear proliferative activity was analyzed immunohistochemically in 19 cases, using a monoclonal anti-Ki-67 antibody. RESULTS: PINHG was found in prostatic carcinoma surrounding in 30% of the patients. In hyperplastic epithelia Ki-67 proliferative activity was 1,08, in PINHG 2,25 (p < 0,05), while in prostatic cancer, Ki-67 index was 17,64. Proliferative activity in prostatic carcinoma was significantly higher than in PINHG (p < 0,001) and hyperplasia (p < 0,001). CONCLUSION: This study confirmed that high grade PIN lesion predominately appears in the surrounding of poor or moderately differentiated prostate carcinoma, and that it represents progressive disorder of proliferation in preneoplastic and neoplastic prostatic epithelium.

[Clinico-pathological characteristics of colonic carcinoma in relation to localization and histologic type].

BACKGROUND/AIM: A significant increase in colonic carcinoma has been analyzed in numerous clinico-pathological studies and genetics models. The aim of this study was to determinate the differences in clinico-pathological parameters of colonic carcinoma regarding localization and histological type. METHODS: The study enrolled 124 patients who had undergone surgery for colorectal carcinoma at the Clinic of Surgery in the town of Nis within 2005. Paraffin sections were stained with Hematoxilin-eosin (H&E), High iron diamine-Alcian blue (HID-AB) (pH-2.5) and Alcian blue-Preiodic arid Shiff (AB-PAS) (pH-2.5) methods. From pathological report we used data regarding: localization (right or left colon), histological type, histological grade, and parameters which determinate the tumor stage. RESULTS: The total number of patients with right-sided colonic carcinoma was 40 (32.36%) and 84 (67.74%) with left-sided colonic carcinoma. Histopathologically, 96 (77.42%) adenocarcinomas and 28 (22.58%) mucinous adenocarcinomas were verified. There were no statistically significant differences between the right-sided and left-sided colonic carcinoma regarding sex, age, histological grade and tumor stage (p > 0.05). Mucinous adenocarcinomas was statistically significantly more frequent in right-sided colon (35.00%) than in left-sided colon (16.67%) (p < 0.05). There were no statistically significant differences between adenocarcinomas and mucinous adenocarcinomas regarding sex and disease stage. In younger patients the percentage of mucinous adenocarcinomas (28.57%) compared to non-mucinous ones (11.46%) was significantlly higher (p < 0.05) than in older patients. Mucinous adenocarcinomas had statistically significantly more frequently poor differentiation in comparision to adenocarcinomas (46.43% versus 9.37%, p < 0.001). CONCLUSION: According to the presented results it can be concluded that the lower grade of differentiation of the colon adenocarcinoma and mucinous secretion are significantly often present in younger patients.

[Prognostic parameters in FIGO stage I endometrial carcinoma of endometrioid type].

BACKGROUND/AIM: Endometrial carcinoma is the most common malignant neoplasm of the female genital tract in developed countries. Endometrioid carcinoma represents about three-fourths of all endometrial carcinoma. The aim of this study was to examine pathologic parameters, age, and the 5-year survival of the patients with FIGO stage I endometrial carcinoma of endometrioid type and to assess the prognostic utility of age, depth of myometrial invasion, hystologic type (endometrioid or variant), histologic grade, nuclear grade, and lymph-vascular space invasion. METHODS: Age, pathologic parameters, and survival data were retrospectively collected on 236 patients with FIGO stage I endometrial carcinoma of endometrioid type. All the patients underwent hysterectomy between 1996 and 2000 and follow-up until December 2005. RESULTS: A total of 236 patients (mean age 58.0, range 40-79) were analyzed. During the 5-year period of follow-up, 59 (25.0%) patents died from the disease. An univariate analysis revealed that age, depth of myometrial invasion, histologic grade, nuclear grade, and lymph-vascular space invasion were associated significantly with the 5-year survival of the patients. A multivariate analysis revealed that age, lymph-vascular space invasion, and depth of myometrial invasion were associated significantly with the 5-year survival. CONCLUSION: Age, lymph-vascular space invasion, and depth of myometrial invasion are independent prognostic parameters for the 5-year survival of the patients with FIGO stage I endometrial carcinoma of endometrioid type.

[Successful resuscitation of a patient with prolonged tocolytic therapy and an emergency cesarean section]. / Uspesna reanimacija bolesnice na produzenoj tokolitickoj terapiji kod koje je uraden hitan carski rez.

Ritodrine is the only medicament approved by FDA in the USA as well as in our country for prevention of the threatening preterm labor. Its adverse effects upon the respiratory and cardiovascular systems, including pulmonary oedema and myocardial ischemia, occur more frequently during the intravenous therapy than during the oral maintenance therapy. The aim of this report was to present a patient with cardiovascular adverse effects of ritodrine, who had her pregnancy terminated by an urgent cesarean section under general anesthesia. In the course of operation, the patient had two cardiac arrest (total of 70 min). Resuscitation was performed by direct and indirect heart massage. The patient's condition was stabilized during the next six hours. The patient was transferred to the coronary unit, where the treatment was continued for 30-days period, after which the patient was released home as completely recovered.

[Co-induction in ambulatory anesthesia]. / Koindukcija u ambulantnoj anesteziji.

BACKGROUND: Co-induction in anesthesia is very useful: synergistic effects of two inductional drugs may lower the dose regimen and the incidence of adverse effects. The aim of this study was to investigate and compare two anesthesiological techniques for short-lasting gynecological procedures in outpatient anesthesia. A total of 80 patients scheduled for surgical termination of pregnancy were randomly assigned into two equal groups--control and co-induction group. METOHDS: The first group of patients received atropine 0.5 mg i.v., alfentanil 0.5 mg i.v. and propofol as a fractionated i.v. bolus until the loss of eyelash reflex. The second group received atropin 0.5 mg, alfentanil 0.5 mg, midazolam 3 mg and propofol in the same manner as the first group. Anesthesia was maintained with propofol increments. Cardiovascular parameters, parameters of post anesthesia recovery and the adverse effects were registered. RESULTS: In patients receiving midazolam inductional dose of propofol was significantly lower, whereas cardiovascular parameters were not significantly different. The recovery after anesthesia was slightly longer after co-induction, but it was not of great clinical significance. The reduction of the adverse effects was found in the co-induction group. CONCLUSION: The results of the study showed that co-induction of midazolam-propofol in comparison with propofol alone for outpatient anesthesia had the following advantages: the reduction of propofol dose, better quality of anesthesia and the reduction of the adverse effects. Recovery was faster in the group that didn't receive midazolam, but it was not of great clinical significance. The conclusion is that co-induction with the combination midazolam-propofol has the advantage in outpatient procedures.