Novel score for prediction of malignant bile duct obstruction based on biochemical and clinical markers.

BACKGROUND: Early differentiation of malignant from benign bile duct obstruction is of utmost importance. AIM: To identify biochemical and clinical predictors for malignancy in patients with bile duct obstruction, and establish a predictive model by combining pre-treatment patient characteristics. A web-based application was developed for easy assessment of malignant bile duct probability (www.pmal-score.org). METHODS: One thousand hundred and thirty-five patients [median age 66 (52-75) years, 53% male] with bile duct obstruction of various aetiologies were retrospectively evaluated at our tertiary referral centre. Multivariate logistic regression analysis identified factors as independently significant for malignant bile duct obstruction. A predictive risk score was established using ROC analysis and applied to an external validation cohort of 101 patients. RESULTS: Three hundred and ninety-four patients had malignant bile duct obstruction proven by surgery, while in 741 patients benign obstruction was observed. Multivariate analysis identified various clinical factors to be predictive for malignancy. On the basis of eight predictors, a risk score for malignancy was developed [X = 0.025 * [age] + 1.239 * [1 if weight loss, otherwise 0] - 0.235 * [1 if pain, otherwise 0] + 0.649 * [1 if diabetes, otherwise 0] + 0.896 * [1 if jaundice, otherwise 0] + 0.109 * [bilirubin] + 0.0007 * [γ-GT] + 0.0003 * [AP] - 4.374]: A significant correlation between the predicted malignancy and the actual malignancy was found by ROC (AUC: 0.862; 95% CI 0.838-0.886, P < 0.0001). CONCLUSIONS: This predictive risk score estimates the risk of malignancy in patients with bile duct obstruction, and it seems to be very accurate. A better prediction enables both earlier diagnosis of malignant obstructive disease and improved management of patients with bile duct obstruction, which may result in reduced morbidity and mortality.

Highly water-soluble prodrugs of anthelmintic benzimidazole carbamates: synthesis, pharmacodynamics, and pharmacokinetics.

Highly water-soluble prodrugs 1a- g of anthelmintic benzimidazole carbamates 2a- g were synthesized. These prodrugs combine high aqueous solubility and stability with high lability in the presence of alkaline phosphatases. The veterinary utility of 1a was shown by a pharmacodynamic and pharmacokinetic study performed in swine. Comparable anthelmintic efficacy was observed with prodrug 1a or the parent fenbendazole 2a. The pharmacokinetic results showed that 2a is better absorbed when derived from 1a than when applied as such.

Outbreaks of Soybean Frogeye Leaf Spot in Iowa.

Frogeye leaf spot of soybean, caused by Cercospora sojina, is typically a disease of warm and humid regions (2). Although the disease was reported in the Midwest in the 1920s (1), no outbreaks have been recorded in Iowa. Outbreaks of frogeye leaf spot occurred during 1999 in soybean fields in Ames and Grand Junction in central Iowa. During the 2000 growing season, the disease occurred in southwestern, southcentral, central, southeastern, and east-central Iowa. Occurrences of the disease with severity (reduction of green leaf area) greater than 50% were observed in production soybean fields at Grand Junction in central Iowa and Central City in eastern Iowa. In a 12-ha no-till field planted with cv. Asgrow 2501, the disease was noticeable and uniformly distributed in the entire field in mid July. Disease severity in this field was greater than 70% by the end of August. Disease incidence, however, was less than 10% in three adjacent soybean fields. In a soybean performance test at a central Iowa location where the disease occurred in 1999 and 2000, the disease was observed on all 80 varieties, with four having a severity equal to or greater than 40%. Fourteen entries had less than a 10% disease severity and 19 entries had a disease severity equal to or greater than 30%. Infected leaves in these locations had typical lesions of frogeye leaf spot, which appeared as reddish brown margins surrounding light brown or ash gray centers. On the infected tissues, hyaline, straight, and multiseptate conidia from clustered conidiophores were found, isolated, and identified to C. sojina. The relatively warm winter temperatures in 1998 to 1999 and 1999 to 2000 were associated with frogeye leaf spot epidemics. Because of the seedborne nature of C. sojina, efforts are warranted to monitor and survey the occurrence of frogeye leaf spot in Iowa, an important seed production state in the northern soybean production region. References: (1) K. Athow and A. H. Probst. Phytopathology 42:660-662, 1952. (2) D. V. Phillips. 1999. Pages 20-21 in: Soybean Disease Compendium. Hartman et al. eds, American Phytopathological Society. St. Paul, MN.

Soybean Varietal Response and Yield Loss Caused by Sclerotinia sclerotiorum.

Soybean Sclerotinia stem rot, caused by Sclerotinia sclerotiorum, has recently emerged from being a minor problem in areas where soybeans of maturity groups 0 to I are grown to a significant cause of soybean yield losses in the north-central region, which produces 80% of soybean in the United States. Studies were conducted in Iowa to quantify varietal response to S. sclerotiorum for cultivars of maturity groups I to III in fields that had uniform infestation histories. Over the course of the study, disease incidence was generally high at the northern Iowa sites but low in central Iowa, with disease incidence of susceptible standards >60% and <30%, respectively. Regression analysis showed that maturity class significantly affected disease incidence, with greater effects in environments where susceptible standard cultivars had high disease incidences. Consistency of varietal response among the environments was quantified using Pearson correlation analysis. When disease incidence was high, varietal responses measured by disease ratings and yield were consistent among locations, but the responses were inconsistent when disease incidence was low. Pearson correlation coefficients ranged from 0.80 to 0.94 for disease incidence and 0.58 to 0.81 for yield among the experiments having high disease incidence in susceptible standards. The relationship between disease incidence and yield was well described by linear regression models with coefficients of determination (r2) ranging from 0.59 to 0.83. Based on regression slopes (significant at P < 0.0001), yield losses are estimated to range from 170 to 335 kg/ha for each 10 percentage points of disease incidence. Regression analysis also showed that maturity groups had a linear relationship with disease incidence (r2 = 0.18 to 0.39, P < 0.01).

Treatment and Outcome of Adolescents with Anorrexia Nervosa.

Primary care providers need to be aware of the therapeutic partnership required for successful treatment of adolescents with anorexia nervosa. This partnership, based on a biopsychological model, addresses multiple aspects of the disorder rather than isolated goals, such as weight gain. Early, established, and severe anorexia are discussed, as well as options for outpatient care and hospitalization and possible outcome.

A modified method for experimental candidosis in mice avoiding lethality.

A model is presented which selected one out of 150 Candida albicans strains for the evaluation of antifungal agents. The mice were inoculated with 6 x 10(5) CFU of strain 352 into the tail vein. The strain has a moderate phospholipase B (PLB) activity in vitro and was originally isolated from a stool sample from a patient in an intensive care unit. This infection leads to very little suffering in the infected animals during the 6-day observation period. Kidney counts at day 5 after infection can give a first indication for a possible fungistatic mechanism. Possible interesting drugs can then be evaluated by a second set of experiments using a longer observation time to investigate the compounds for fungicidal properties. The model suggests that screening for systemic antifungals by avoiding lethality of mice in the first place can be done.

Increase of DNA content in tissue stages of Entamoeba histolytica strain SFL 3.

The DNA content of culture forms and tissue stages of pathogenic E. histolytica strain SFL 3 were measured photometrically after the nuclei had been stained with the fluorochrome BAO. As a control, the DNA guartity of E. histolytica strain HK 9 and E. invadens were determined by the same method and compared with reference values. Tissue stages were obtained from hamsters experimentally infected by intrahepatic injection of SFL 3 amoebae. Further studies concerning possible changes in the DNA content of tissue stages involved the following methods: (a) isolation of tissue stages from the liver, followed by distinct suspension periods. (b) Infected liver pieces were directly transferred into culture medium; amoebae emigrating therefrom were cultivated. The study demonstrated that tissue stages contained up to 4 times more DNA than did culture forms. After 3 h cultivation, the DNA content of tissue stages decreased to the level of culture forms. Possible reasons for this change are discussed.

Antimalarial activity of new floxacrine-related acridinedione derivatives: studies on blood schizontocidal action of potential candidates against P. berghei in mice and P. falciparum in vivo and in vitro.

Deoxyfloxacrine derivatives (1-hydrazone: S 83 0083; 1-imine: S 84 7277) and floxacrine derivatives (10-methoxy-floxacrine: L 84 7667; 1-imine: L 84 7693) selected from a series of newly synthesized 3-aryl-7-chloro-3,4-dihydro-1,9(2H,10H)-acridinediones were evaluated for blood schizontocidal activities in mice infected with asexual stages of various drug-resistant lines of P. berghei and in New World monkeys infected with blood schizonts of different chloroquine-resistant strains of P. falciparum. All compounds tested showed high activity against drug-resistant lines of P. berghei (ED50: 1.0-4.4 mg/kg x 5, per os) and were distinctly superior in their antimalarial potency to floxacrine. Compounds L 84 7667 and L 84 7693 proved to be highly active against the FCBR strain of P. falciparum in vitro (IC50: 0.73-1.78 nmol); they effected temporary clearance of parasitemias due to the Palo Alto strain of P. falciparum in squirrel monkeys at oral doses of 15 mg/kg given daily for 5 consecutive days. Compounds S 83 0083 and S 84 7277, showing moderate in vitro effects (12.9-24.8 nmol), cleared parasitemias of the FCBR strain of P. falciparum in owl monkeys at oral doses of 20 mg/kg (S 84 7277) given daily for 5 or 7 consecutive days (follow-up period, 17 and 30 days, respectively) or at doses of 20 mg/kg (x 4) (S 83 0083) followed by doses of 40 mg/kg (x 3) within a follow-up period of 30 days. These observations suggest that the range of doses required for the cure of established P. falciparum infections is probably too large to cover infections with strains of the least susceptibility and might evoke toxic reactions by the potential candidates tested.

Effects of dimethylsulfoxide and the deep-freezing process on the infectivity, motility, and ultrastructure of Trypanosoma cruzi.

The effects of dimethylsulfoxide (DMSO, final concentration 5%) and the deep-freezing process on the infectivity (ID50), motility, and ultrastructure of nontreated and DMSO-treated Trypanosoma cruzi suspensions (PSG-3 buffer with 10% horse serum) were investigated prior to and after cryopreservation in liquid nitrogen. DMSO equilibration caused distinct suppression of motility and characteristic, fine structural alterations in numerous organelles, such as myelin-like structures in the cytoplasm and/or inside the mitochondrial apparatus, enlargement of the perinuclear space, endoplasmic reticulum, and mitochondrial cristae, as well as condensation of the kinetoplast with loss of its lamellar structure. There was no evidence of loss of infectivity in DMSO-treated parasites. DMSO-treated and deep-frozen organisms showed, however, very similar fine structural alterations, although damage occurring during freezing and thawing was more pronounced. Apart from the frequently enlarged kinetoplast and the loosening of its mitochondrial matrix, numerous trypanosomes revealed total disintegration of the kinetoplast-mitochondrion complex with loss of its whole matrix. Deep-frozen trypanosomes were significantly less infective to mice than nontreated organisms, and their motility was strongly suppressed. These results suggest that cryopreservation and thawing of T. cruzi may lead to severe damage of the mitochondrial apparatus and thus to heavy disorders of metabolic function, exhaustion of the metabolic pool, and finally, to death of such damaged trypanosomes, despite the use of DMSO as a cryoprotective agent.

Ultrastructural changes of Trichomonas vaginalis prior and after cryopreservation.

Fine structural changes of Trichomonas vaginalis are described prior and after the freezing process in liquid nitrogen. Dimethyl sulfoxide (DMSO) used as the cryoprotectant caused distinct alterations of the cytoplasm when trichomonads were equilibrated with 5% DMSO under various experimental conditions. Changes were bubble-like protrusions, fissuration and/or vacuolation of the cytoplasm, doubling and removal or/and rupture of the cell membrane. Apart from these findings cryopreservation caused marked alterations on the hydrogenosomes, such as condensation and flocculence of the usually homogeneous contents; in addition numerous hydrogenosomes fused while loosing membrane at the site of fusion. However, several parasites revealed normal hydrogenosomes after the freezing process. It is assumed that these organisms survived freezing and thawing as demonstrated by successful cultivation of recovered trichomonads.

[Fine-structure changes in Toxoplasma gondii trophozoites after deep-freezing with dimethyl sulphoxide (author's transl)]. / Feinstrukturelle Veränderungen von Toxoplasma gondii-Trophozoiten nach Tiefgefrierung mit Dimethylsulfoxid.

The changes observed in trophozoites of Toxoplasma gondii after deep-freeze preservation were examined by electron microscopy. Toxoplasmas (strain BK) from peritoneal exudate of infected NMRI mice were supended in Ringer's solution, deep-frozen in liquid nitrogen with 5% dimethylsulphoxide (DMSO), and compared after thawing with control samples with and without the addition of DMSO. Slight structural changes such as widening of endoplasmic reticulum, formation of fissures in the cytoplasm, and loosening of chromatin were only observed in some of the free toxoplasmas of the DMSO control. Among the deep-frozen parasites, about 1/5 of the free stages showed no or only slight morphological changes. In contrast to this, almost all intracellular forms found in macrophages showed lesions. The most remarkable change was a partial destruction of the inner cell membrane complex. The outflow of ribosome-containing protoplasm with ballon-like swelling of the outer elementary membrane was observed as a consequence of this frequent lesion. The outflow of protoplasm induced a drastic decrease in the electronic density of the whole cytoplasm. Other characteristic degenerative signs were vacuolation of cytoplasm up to formation of great optically empty spaces, widening of the perinuclear space, swelling of mitochondria, disintegration of rhoptria, micronemata, and Golgi zone, coarse-plaque loosening, and displacement of electron-dense areas of the nucleus up to disintegration with maintenance of the karyoplasm. In some almost completely disintegrated trophozoites, enlarged mitochondria with remarkable electronic density were observed. Apart from the cell membrane, the conoid was the longest-persisting organelle. The alterations observed after deep-freezing permit the conclusion that the free cells, which were only slightly impaired or not at all, remained infective.

[Invasion of erythrocytes by toxoplasma gondii (author's transl)]. / Invasion von Erythrozyten durch Toxoplasma gondii.

The in vitro-invasion of mouse erythrocytes by Toxoplasma gondii could be detected and analysed by electron microscopy. The sequence of events observed during erythrocyte invasion led to the assumption of an actively penetrating parasite into the non-phagocytic host cell.

Structural changes on Entamoeba histolytica trophozoites after cryopreservation in liquid nitrogen.

Trophozoites of Entamoeba histolytica cultures which had been deep-frozen in the presence of 5% DMSO, along with untreated cells and cells treated with DMSO (5%), were examined for fine-structural changes. After deep-freezing in liquid nitrogen only a few amoebae exhibited normal nuclear and cytoplasmic structure. One frequently observed but unspecific finding pertaining to recovered cells is the separation of the cytoplasm into large vacuolated (coarse-granular) and electron-optically fine-granular (hyaline) zones. The glycogen which normally lies in the cytoplasm is always eluted. In many cases numerous short RNP helices are scattered unevenly in the vesicular plasma, but they are also found in larger masses adjacent to the membranes of still intact and already damaged nuclei. Moderately damaged nuclei have a poorly folded membrane and their chromatin is markedly denatured. More heavily damaged nuclei have a membrane which has partly fibrillated or ruptured and then formed conspicuous folds, where the nuclear membrane has ruptured nucleoplasmic remnants of chromatin and button-like bodies appear to pour into the surrounding cytoplasm. The final destruction of the cell is marked by coalescing autolytic zones, first in the vacuolated and later in the fine-granular cytoplasm. Finally only remnants of the nuclear membrane and of the membranes of numerous vacuoles remain. It is assumed that most of the changes in the cytoplasm are of a secondary nature and are caused by the early functional disturbance of the nucleus.