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Aim: The multicentre non-interventional AVANTI study assessed safety, effectiveness and patient-reported outcomes with approved first-line bevacizumab-containing regimens for HER2-negative locally recurrent/metastatic breast cancer (LR/MBC) in German routine oncology practice., Methods: Eligible patients had HER2-negative LR/MBC, no bevacizumab contraindications and no prior chemotherapy for LR/MBC. Chemotherapy schedule, diagnostics and follow-up were at physicians' discretion. Data were collected for 1 year after starting bevacizumab, then every 6 months for 1.5 years (maximum follow-up: 2.5 years). Patients and physicians rated treatment satisfaction. Subgroup analyses were prespecified in clinically relevant populations, including triple-negative breast cancer (TNBC)., Results: Between November 1, 2009 and April 30, 2016, 2065 eligible patients at 346 centres received bevacizumab with paclitaxel or capecitabine. Patients receiving bevacizumab-capecitabine were less likely to have de novo disease and more likely to have TNBC, age ≥60 years and prior anthracycline/taxane and/or endocrine therapy. Median PFS was 12.6 (95% CI 11.9-13.2) months (12.8 with bevacizumab-paclitaxel, 10.5 with bevacizumab-capecitabine); median OS was 23.9 (95% CI 22.2-25.1) months. Outcomes were worse in patients with TNBC, prior anthracycline/taxane or prior endocrine therapy. Grade ≥3 adverse events occurred in 27% of patients. Treatment was discontinued for adverse events in 15%. Treatment satisfaction was rated as good or better by 304/394 responding patients (77%) at week 54 and in 1393/2065 patients (67%) by physicians overall., Conclusions: In routine clinical practice, effectiveness and safety of first-line bevacizumab-containing therapy for LR/MBC were consistent with experience from phase III trials. Patient and physician treatment satisfaction showed high concordance., Competing Interests: Declaration of interest statement V.M. reports honoraria from Amgen, AstraZeneca, Daiichi-Sankyo, Eisai, Pfizer, MSD, Novartis, Roche, Teva and Seattle Genetics (for advisory boards) and from Genomic Health, Hexal, Roche, Pierre Fabre, Amgen, ClinSol, Novartis, MSD, Daiichi-Sankyo, Eisai, Lilly, Tesaro, Seattle Genetics and Nektar (for consultancy); research funding to his institution from Novartis, Roche, Seattle Genetics and Genentech; and travel grants from Roche, Pfizer and Daiichi-Sankyo. O.H. reports honoraria from Roche, Novartis, Pfizer, MSD, Daiichi-Sankyo, AstraZeneca, Eisai, Hexal, Amgen and Riemser Pharma; advisory/consultancy roles for Roche, Novartis, MSD, Daiichi-Sankyo and Hexal; and travel/accommodation/expenses from Novartis, Pfizer, MSD, Daiichi-Sankyo, Eisai, Hexal and Amgen. A.-K.S.-J. is an employee of Roche Pharma AG and holds shares in Roche. A.S. reports honoraria from Roche, AstraZeneca, Celgene, Pfizer, Novartis, MSD, Tesaro and Lilly; expert testimony for Roche and AstraZeneca; research funding to his institution from Roche, Celgene and AbbVie; and travel/accommodation/expenses from Roche, Celgene and Pfizer. The remaining authors declare no conflicts of interest., (Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Anthropometry provides information on the physical status of the individual and can be associated with aspects of health including nutritional status. Currently, the stratification of the arm and calf circumferences is classified into only two situations: "malnourished" and "well-nourished". A total of 513 interviews were conducted, and 391 elderly people (≥ 65 years) completed the assessment using the Mini Nutritional Assessment (MNA) and anthropometry of selected samples of the population of Cuiabá-MT. The body mass index (BMI, kg/m2) was calculated for the elderly people, establishing five new reference values for circumferences, arm relaxed (RAC), abdomen (AC), and calf (CC) in centimeters (cm). The median age was 71 years (64% women and 36% men) and was correlated to the RAC (r=-0.180, p<0.001) and CC (r=-0.202, pp<0.001). The BMI obtained the median of 27 (15% malnourished, risk of malnutrition 13%, eutrophic 24%; overweight 33%, obese 16%), and it was correlated to the RAC (r=0.798, pp<0.001), AC (r=0.823 p p<0.001) and CC (r=0.605, pp<0.001). The MNA was 26 (malnourished 13.8%, risk of malnutrition 12.3%, well-nourished, 73.9%). The BMI stratification by morbidity vs no morbidity was 27.50 (n=287) and 24.4 (n=104) to total sample respectively (pp<0,05). The RAC x AC (r=0.798, pp<0.001), RAC x CC (r=0.648, pp<0.001), and CC x AC (r=0.496, pp<0.001) were correlated between themselves. The eutrophic classification by circumference for both genders: RAC=27.1-29.00 cm, AC=88.1-95.00 cm, CC=32.60-33.00 cm. There are more overweight and obese than malnourished, which is a risk factor for morbidity and MNA only identifies malnutrition. Circumferences showed good association with BMI and are easy to apply. Therefore, the proposal of the circumferences can simplify and expand the nutritional assessment.

Invasive lobular carcinomas (ILC) show better clinical behaviour compared with other histological types, but significantly lower pathological complete response (pCR) rates after neoadjuvant chemotherapy (NACT). We investigated whether factors influencing pCR rate in ILC after NACT can be identified and whether clinical outcome is different. 9,020 breast cancer patients from nine German neoadjuvant trials with known histological type were pooled. 11.7 % of tumours were ILC. Endpoints were: pCR rate, surgery type and survival. ILC was associated with older age, larger tumour size, lymph node negativity, lower grade and positive hormone-receptor-status (HR). Patients with ILC achieved a significantly lower pCR rate compared with non-ILC patients (6.2 vs. 17.4 %, P < 0.001). The pCR rate was 4.2 % in ILC/HR+/G1-2, 7.0 % in ILC with either HR- or G3, and 17.8 % in ILC/HR-/G3. Mastectomy rate was higher in ILC compared with non-ILC patients irrespective of response to NACT (pCR: 27.4 vs. 16.6 %, P = 0.037 and non-pCR: 41.8 % vs. 31.5 %, P < 0.0001). Age and HR independently predicted pCR in ILC. In ILC patients, pCR did not predict distant disease free (DDFS) and loco-regional disease free survival (LRFS), but overall survival (OS). Non-pCR patients with ILC had significantly better DDFS (P = 0.018), LRFS (P < 0.0001) and OS (P = 0.044) compared with non-ILC patients. Patients with ILC had a low chance of obtaining a pCR and this is not well correlated with further outcome. The mastectomy rate was considerably high in ILC patients even after obtaining a pCR. We, therefore, suggest to offer NACT mainly to ILC patients with HR-negative tumours.

Background: Preoperative chemotherapy is a recommended treatment of both primary operable and locally advanced breast cancer. Strategies to improve efficacy include the use of anthracyclines, taxanes, and intensified dose with bone marrow support., Patients and Methods: Patients received neoadjuvant epirubicin 90 mg/m(2) plus cyclophosphamide 600 mg/m(2) followed by paclitaxel 175 mg/m(2) (EC→T), each 3-weekly for four cycles (n = 370), or epirubicin 150 mg/m(2) followed by paclitaxel 225 mg/m(2) with pegfilgrastim followed by CMF (cyclophosphamide 500 mg/m(2), methotrexate 40 mg/m(2), fluorouracil 600 mg/m(2)) on days 1 and 8 (E(dd)→T(dd)→CMF), each 2-weekly and for three cycles (n = 363). Patients were randomly allocated to either simultaneous darbepoetin alfa (DA) (n = 356) or none (n = 377)., Results: Pathological complete response (pCR) rate (breast) was higher with E(dd)→T(dd)→CMF, 18.7% versus 13.2% with EC→T; P = 0.043, ypT0/Tis; ypN0 was reported in 20.9% versus 14.3% respectively; P = 0.019. Patients with grade 3 tumors and negative hormone receptor status had a significantly higher pCR rate. Mean hemoglobin values maintained higher with DA (13.6 versus 12.6 g/dl). E(dd)→T(dd)→CMF regimen showed more grade 3-4 mucositis, sensory neuropathy, and neurological complaints. Thromboembolic events were more frequent on DA (3% versus 6%; P = 0.055)., Conclusion: Dose-dense and -intensified neoadjuvant chemotherapy with E(dd)→T(dd)→CMF was potentially superior to EC→T in terms of pCR. Primary use of DA did not affect pCR.

Purpose: To evaluate efficacy and safety of epirubicin and cyclophosphamide followed by paclitaxel and trastuzumab as neoadjuvant treatment in patients with human epidermal growth factor receptor 2 (HER2)-overexpressing breast cancer., Patients and Methods: Patients with centrally confirmed HER2-overexpressing breast cancer (≥ 2 cm or inflammatory) received four 3-week cycles epirubicin and cyclophosphamide (90/600 mg/m(2)) followed by four 3-week cycles paclitaxel (175 mg/m(2)) and trastuzumab (6 mg/kg) before surgery. Trastuzumab was continued after surgery to complete 1 year of treatment. Primary end point was pathologic complete response (pCR) defined as no residual invasive tumor in breast and lymphatic tissue., Results: Thirty-nine percent of 217 enrolled patients achieved a pCR. Breast conservation was possible in 64% of patients. Three-year disease-free survival (DFS) was 88% in patients with pCR compared to 73% in patients without pCR (P = .01). Three-year overall survival (OS) was 96% in patients with pCR compared to 86% in patients without pCR (P = .025). pCR was the only significant prognostic factor for DFS (hazard ratio [HR] 2.5; 95% CI, 1.2 to 5.1; P = .013) and OS (HR, 4.9; 95% CI, 1.4 to 17.4; P = .012) in multivariable analysis. Cardiac toxicity was reported in eight patients (3.7%) of whom six presented with an asymptomatic left ventricular ejection fraction decrease and two with symptomatic chronic heart failure., Conclusion: Neoadjuvant combination of trastuzumab and chemotherapy resulted in a high pCR rate in HER2-overexpressing primary breast cancer. Patients with a pCR after neoadjuvant anti-HER2 therapy in combination with chemotherapy followed by maintenance trastuzumab have an improved long-term outcome. Patients without a pCR had an increased risk for relapse and death.

Background: The objective of this study was to compare the effect of dose-intensified neoadjuvant chemotherapy with that of standard epirubicin plus cyclophosphamide followed by paclitaxel in combination with or without darbepoetin on survival in primary breast cancer., Patients and Methods: A total of 733 patients received either four cycles of neoadjuvant epirubicin 90 mg/m(2) plus cyclophosphamide 600 mg/m(2) every 3 weeks followed by four cycles of paclitaxel 175 mg/m(2) every 3 weeks (EC→T), or three cycles of epirubicin 150 mg/m(2) every 2 weeks followed by three cycles of paclitaxel 225 mg/m(2) every 2 weeks followed by three cycles of combination chemotherapy with cyclophosphamide, methotrexate, and fluorouracil (E(dd)→T(dd)→CMF). The patients were randomly assigned to receive darbepoetin or none. The primary objective was to demonstrate a superior disease-free survival (DFS) of E(dd)→T(dd)→CMF compared with EC→T., Results: Estimated 3-year DFS was 75.8% with EC→T versus 78.8% with E(dd)→T(dd)→CMF [hazard ratio (HR) 1.14; P = 0.37] and overall survival (OS) 88.4% versus 91.5% (HR 1.26; P = 0.237). Three-year DFS was 74.3% with darbepoetin versus 80.0% without (HR 1.31; P = 0.061) and OS 88.0% versus 91.8% (HR 1.33; P = 0.139). Patients with a pathologically documented complete response [pathological complete response (pCR)] had a significantly better DFS compared with those without achieving a pCR (estimated 3-year DFS: 89.2% versus 74.9%; HR 2.27; P = 0.001)., Conclusion: Neoadjuvant dose-intensified chemotherapy compared with standard chemotherapy did not improve DFS, whereas the addition of darbepoetin might have detrimental effects on DFS.

Pathological complete response (pCR) to neoadjuvant treatment correlates with outcome in breast cancer. We determined whether characteristics of neoadjuvant therapy are associated with pCR. We used multi-level models, which accounted for heterogeneity in pCR across trials and trial arms, to analyze individual patient data from 3332 women included in 7 German neoadjuvant trials with uniform protocols. PCR was associated with an increase in number of chemotherapy cycles (odds ratio [OR] 1.2 for every two additional cycles; P = 0.009), with higher cumulative anthracycline doses (OR 1.6; P = 0.002), higher cumulative taxane doses (OR 1.6; P = 0.009), and with capecitabine containing regimens (OR 1.62; P = 0.022). Association of pCR with increase in number of cycles appeared more pronounced in hormone receptor (HR)-positive tumors (OR 1.35) than in HR-negative tumors (OR 1.04; P for interaction = 0.046). Effect of anthracycline dose was particularly pronounced in HER2-negative tumors (OR 1.61), compared to HER2-positive tumors (OR 0.83; P for interaction = 0.14). Simultaneous trastuzumab treatment in HER2-positive tumors increased odds of pCR 3.2-fold (P < 0.001). No association of pCR and number of trastuzumab cycles was found (OR 1.20, P = 0.39). Dosing characteristics appear important for successful treatment of breast cancer. Longer treatment, higher cumulative doses of anthracyclines and taxanes, and the addition of capecitabine and trastuzumab are associated with better response. Tailoring according to breast cancer phenotype might be possible: longer treatment in HR-positive tumors, higher cumulative anthracycline doses for HER2-negative tumors, shorter treatment at higher cumulative doses for triple-negative tumors, and limited number of preoperative trastuzumab cycles in HER2-positive tumors.

Background: Pegylated liposomal doxorubicin (PLD) is active in metastatic breast cancer. This observational study evaluated the efficacy and safety of PLD in patients treated during routine clinical practice., Methods: Eligible patients had metastatic breast cancer and were treated with PLD according to the dose and schedule determined by their physician as part of routine practice. The primary objectives were to analyze the efficacy and toxicity of PLD therapy., Results: 125 patients were assessable. Median age was 62 years, 78% had performance status 0-1, and 60% had estrogen-receptor-positive disease. PLD treatment was second- or third-line in 69% of patients. Prior anthracyclines (adjuvant or metastatic) had been used in 56% of patients. The majority of patients (79%) received PLD every 4 weeks at a median dose of 40 mg/m2. Overall response rate was 43% in all patients and 34% in those previously treated with anthracyclines. The most common grade 3/4 adverse events were skin toxicity/hand-foot syndrome (6%), and leukopenia (3%)., Conclusions: This observational study supports the activity and tolerability of PLD in metastatic breast cancer as demonstrated in PLD clinical trials.

Pectate lyase (PL) is a potent cell wall-degrading enzyme known to play a role in the microbial infection of plants. We re-examined the pectolytic property of seven representative pathovars of Pseudomonas syringae. None of the 10 P. syringae pv. glycinea strains examined exhibited pectolytic activity. However, the PL gene (pel) was detected by Southern hybridization in four out of four P. syringae pv. glycinea strains examined. A P. syringae pv. glycinea pel gene was cloned, sequenced, and predicted to encode a protein sharing 70%-90% identity in amino acid sequence with PLs produced by pectolytic pseudomonads and xanthomonads. A series of amino acid and nucleotide sequence analyses reveal that (i) the predicted P. syringae pv. glycinea PL contains two regions in the amino acid sequence that may affect the formation of a beta-helix structure important for the enzyme activity, and (ii) the P. syringae pv. glycinea pel gene contains a single-base insertion, a double-base insertion, and an 18-bp deletion, which can lead to the synthesis of an inactive PL protein. The function of P. syringae pv. glycinea PL could be restored by removing the unwanted base insertions and by filling in the 18-bp deletions by site-directed mutagenesis. The altered pel sequence was also detected by polymerase chain reaction and nucleotide sequencing in the genomes of other pathovars of P. syringae, including phaseolicola and tagetis.

This study observed the effect of a dietary re-education plus regular physical activity on body composition, risk factors and physical test performance of sedentary overweight/obese women and to correlate these variables one with each other. Fifty women (36 +/- 10 yrs; 31 +/- 6 body mass index (BMI, kg/m2)) volunteered for the study. Body compositions were obtained by anthropometry and bioimpedance and some body indexes were established. One-repetition maximum (1-RM) and treadmill VO2max tests were carried out and blood samples were obtained for lipid, glucose and uric acid analyses before (T1) and after two months of intervention (T2). Diet was established by indirect calorimetry. Body fat, glucose, uric acid, total cholesterol, HDL-cholesterol and systolic blood pressure were significantly reduced. The 1-RM and VO2max tests were significantly increased. Neck circumference (NC) was correlated with body composition, back muscle 1-MR, HDL and LDL cholesterol, total cholesterol/HDL ratio, uric acid, and resting energy expenditure. BMI was found to be significantly correlated with waist/hip ratio, circumference sum, and body fat percentage by anthropometry and bioimpedance. Body fat percentage determined by bioimpedance and anthropometry was significantly correlated with arm fat area and arm fat area corrected respectively, and both with BMI at T1 and T2. This study suggests that a dietary reeducation plus physical activity around 200 min/week improved body composition and the health of these women. Many anthropometry measurements have correspondence to risk factors and NC could be a simple approach to reflect these results, without other more complex techniques.

Rhamnolipids, naturally occurring biosurfactants constructed of rhamnose sugar molecules and beta-hydroxyalkanoic acids, have a wide range of potential commercial applications. In the course of a survey of 33 different bacterial isolates, we have identified, using a phenotypic assay for rhamnolipid production, a strain of the nonpathogenic bacterial species Pseudomonas chlororaphis that is capable of producing rhamnolipids. Rhamnolipid production by P. chlororaphis was achieved by growth at room temperature in static cultures of a mineral salts medium containing 2% glucose. We obtained yields of roughly 1 g/liter of rhamnolipids, an amount comparable to the production levels reported in Pseudomonas aeruginosa grown with glucose as the carbon source. The rhamnolipids produced by P. chlororaphis appear to be exclusively the mono-rhamnolipid form. The most prevalent molecular species had one monounsaturated hydroxy fatty acid of 12 carbons and one saturated hydroxy fatty acid of 10 carbons. P. chlororaphis, a nonpathogenic saprophyte of the soil, is currently employed as a biocontrol agent against certain types of plant fungal diseases. The pathogenic nature of all bacteria previously known to produce rhamnolipids has been a major obstacle to commercial production of rhamnolipids. The use of P. chlororaphis therefore greatly simplifies this matter by removing the need for containment systems and stringent separation processes in the production of rhamnolipids.

Aims: To investigate the resuscitation of acid-injured Salmonella enterica in selected enrichment broths, in apple juice and on cut surfaces of apple and cucumber slices., Methods and Results: Following exposure to 2.4% acetic acid for 7 min, S. enterica (serovars Mbandaka, Chester and Newport) cells were used to inoculate enrichment broths, phosphate-buffered saline (PBS), apple juice and fruit slices. Injured Salmonella cells resuscitated and regained the ability to form colonies on selective agar (Xylose-Lysine-Tergitol 4) if they were incubated in lactose broth (LB), universal pre-enrichment broth (UPB) or buffered peptone water (BPW), but not in tetrathionate broth, PBS or apple juice. The resuscitation occurred at a significantly (P > 0.05) faster rate in UPB than in LB or BPW. The resuscitation also occurred on the surfaces of fresh-cut cucumber at 20 degrees C, but not at 4 degrees C., Conclusions: Acid-injured Salmonella cells resuscitated in nonselective enrichment broths at different rates, but not in selective enrichment broth, apple juice, PBS or on fresh-cut apple., Significance and Impact of the Study: Pre-enrichment of food samples in UPB prior to selective enrichment is recommended. Injured Salmonella cells have the ability to resuscitate on fresh-cut surfaces of cucumber when stored at abusive temperatures.

A study was done to determine the efficacy of aqueous ozone treatment in killing Listeria monocytogenes on inoculated alfalfa seeds and sprouts. Reductions in populations of naturally occurring aerobic microorganisms on sprouts and changes in the sensory quality of sprouts were also determined. The treatment (10 or 20 min) of seeds in water (4 degrees C) containing an initial concentration of 21.8 +/- 0.1 microg/ml of ozone failed to cause a significant (P < or = 0.05) reduction in populations of L. monocytogenes. The continuous sparging of seeds with ozonated water (initial ozone concentration of 21.3 +/- 0.2 microg/ml) for 20 min significantly reduced the population by 1.48 log10 CFU/g. The treatment (2 min) of inoculated alfalfa sprouts with water containing 5.0 +/- 0.5, 9.0 +/- 0.5, or 23.2 +/- 1.6 microg/ml of ozone resulted in significant (P < or = 0.05) reductions of 0.78, 0.81, and 0.91 log10 CFU/g, respectively, compared to populations detected on sprouts treated with water. Treatments (2 min) with up to 23.3 +/- 1.6 microg/ml of ozone did not significantly (P > 0.05) reduce populations of aerobic naturally occurring microorganisms. The continuous sparging of sprouts with ozonated water for 5 to 20 min caused significant reductions in L. monocytogenes and natural microbiota compared to soaking in water (control) but did not enhance the lethality compared to the sprouts not treated with continuous sparging. The treatment of sprouts with ozonated water (20.0 microg/ml) for 5 or 10 min caused a significant deterioration in the sensory quality during subsequent storage at 4 degrees C for 7 to 11 days. Scanning electron microscopy of uninoculated alfalfa seeds and sprouts showed physical damage, fungal and bacterial growth, and biofilm formation that provide evidence of factors contributing to the difficulty of killing microorganisms by treatment with ozone and other sanitizers.

Attachment and survival of Listeria monocytogenes on external surfaces (rind) of inoculated cantaloupe, resistance of the surviving bacteria to chlorine or hydrogen peroxide treatments, transfer of the pathogen from unsanitized and sanitized rinds to fresh-cut tissues during cutting and growth, and survival of L. monocytogenes on fresh-cut pieces of cantaloupe were investigated. Surface treatment with 70% ethanol to reduce the native microflora on treated melon, followed by immersion in a four-strain cocktail of L monocytogenes (10(8) CFU/ml) for 10 min, deposited 4.2 log10 CFU/cm2 and 3.5 log10 CFU/cm2 of L monocytogenes on treated and untreated cantaloupe rinds, respectively. L. monocytogenes survived on the treated or untreated cantaloupe rinds for up to 15 days during storage at 4 and 20 degrees C, but populations declined by approximately 1 to 2 log10 CFU/cm2. Fresh-cut pieces prepared from inoculated whole cantaloupes stored at 4 degrees C for 24 h after inoculation were positive for L. monocytogenes. Washing inoculated whole cantaloupes in solutions containing 1,000 ppm of chlorine or 5% hydrogen peroxide for 2 min at 1 to 15 days of storage at 4 degrees C after inoculation resulted in a 2.0- to 3.5-log reduction in L. monocytogenes on the melon surface. Fresh-cut pieces prepared from the sanitized melons were negative for L. monocytogenes. After direct inoculation onto fresh-cut pieces, L. monocytogenes survived, but did not grow, during 15 days of storage at 4 degrees C. Growth was evident by 4 h of storage at 8 and 20 degrees C. It is concluded that sanitizing with chlorine or hydrogen peroxide has the potential to reduce or eliminate the transfer of L. monocytogenes on melon surfaces to fresh-cut pieces during cutting.

The native microflora of three types of produce (green bell peppers, Romaine lettuce, and prepeeled baby carrots) and two types of sprouting seeds (alfalfa and clover) were investigated. Aerobic plate count (APC) for each produce or seed type as determined on Pseudomonas agar F (PAF) with incubation at 28 degrees C was in the range of 4 to 7 log CFU per g of tissue or seed. There was no significant difference (P > or = 0.05) in APC when the determinations were made with three agar media including PAF, brain heart infusion agar, and plate count agar. However, the APC as determined from plates that were incubated at 28 degrees C was significantly (P < or = 0.05) higher than with incubation at 37 degrees C. Fluorescent pseudomonads accounted for 23 to 73% of APC and 6 to 18% of APC recovered from carrots, pepper, and lettuce were pectolytic. Forty-eight strains of pectolytic bacteria were randomly isolated and identified, respectively, as members of the genera of Pseudomonas, Erwinia, Bacillus, Xanthomonas, or Flavobacterium. Lactic acid bacteria and/or yeast were consistently isolated from baby carrots, lettuce, and sprouting seeds (alfalfa or clover) but not from green bell peppers. Approximately 120 strains of indigenous microflora were tested for their ability to inhibit the growth of Salmonella Chester, Listeria monocytogenes, Escherichia coli, or Erwinia carotovora subsp. carotovora on PAF. Six isolates capable of inhibiting the growth of at least one pathogen were isolated and identified, respectively, as Bacillus spp. (three strains), Pseudomonas aeruginosa (one strain), Pseudomonas fluorescens (strain A3), and yeast (strain D1). When green pepper disks were inoculated with strains A3 and D1, the growth of Salmonella Chester and L. monocytogenes on the disks was reduced by 1 and 2 logs, respectively, over a period of 3 days. Application of strains A3 and D1 as potential biopreservatives for enhancing the quality and safety of fresh produce is discussed.

Thirty-eight strains of filamentous bacteria, many of which are thermophilic or thermotolerant and commonly found in composts and mouldy fodders, were examined for their ability to produce cutinolytic esterase (cutinase) in culture media supplemented with cutin, suberin or cutin-containing agricultural by-products. Initially, the ability of culture supernatants to hydrolyse the artificial substrate p-nitrophenyl butyrate was determined by spectrophotometric assays. Only one bacterium, Thermoactinomyces vulgaris NRRL B-16117, exhibited cutinolytic esterase production. The enzyme was highly inducible, was repressed by the presence of glucose in the medium and hydrolysed both apple and tomato cutins. Inducers included apple cutin, apple pomace, tomato peel, potato suberin and commercial cork. Unlike similar fungal enzymes, the T. vulgaris cutinolytic esterase was not inducible by cutin hydrolysate. The cutinolytic esterase exhibited a half-life of over 60 min at 70 degrees C and a pH optimum of >/= 11.0. This study indicates that thermophylic filamentous bacteria may be excellent commercial sources of heat-stable cutin-degrading enzymes that can be produced by fermentation of low cost feedstocks.

Scanning electron microscopy was used to examine the cotyledons, hypocotyls, and roots of alfalfa, broccoli, clover, and sunflower sprouts purchased from retail outlets as well as alfalfa sprouts grown in the laboratory using a tray system equipped with automatic irrigation. Biofilms were observed on all plant parts of the four types of commercially grown sprouts. Biofilms were also commonly observed on alfalfa sprouts grown in the laboratory by 2 days of growth. Rod-shaped bacteria of various sizes were predominant on all sprouts examined both as free-living cells and as components of biofilms. Occasionally, cocci-shaped bacteria as well as yeast cells were also present in biofilms. The microbes contained in the biofilms appeared to be attached to each other and to the plant surface by a matrix, most likely composed of bacterial exopolysaccharides. Biofilms were most abundant and of the largest dimensions on cotyledons, sometimes covering close to the entire cotyledon surface (approximately 2 mm in length). Naturally occurring biofilms on sprouts may afford protected colonization sites for human pathogens such as Salmonella and Escherichia coli O157:H7, making their eradication with antimicrobial compounds difficult.

The introduction of size-exclusion chromatography (SEC) analysis of polysaccharides prior to MALDI mass spectroscopy accounts for the determination of the molecular mass of the repeating unit when neutral homopolymers are investigated. In the case of natural polysaccharides characterised by more complicated structural features (presence of non-carbohydrate substituents, charged groups, etc.), this mass value usually is in agreement with more than one sugar composition. Therefore, it is not sufficient to give the correct monosaccharidic composition of the polysaccharide investigated. To solve this problem, MALDI spectra were recorded on the permethylated sample and post-source decay experiments were performed on precursor ions. In this way, the composition (in terms of Hex, HexNAc, etc.), size and sequence of the repeating unit were determined.

The influence of pyruvate ketals and acetyl groups on the conformational behaviour of the exopolysaccharide produced by Pseudomonas 'gingeri' strain Pf9 has been investigated experimentally through studies of intrinsic viscosity and circular dichroism experiments. A conformational variation was detected as a function of the ionic strength. Measurements carried out on the native polymer, as well as on both de-pyruvated and de-acetylated samples, suggested a critical role for the acetyl group on the solution conformation of the polysaccharide. Molecular mechanics calculations indicated the possibility of intramolecular hydrogen bonding between acetyl substituents on the mannose and the C(2)OH group of the preceding saccharidic unit. NMR linewidth measurements, carried out as a function of temperature, on the low molecular weight de-pyruvated sample indicated different polymeric backbone dynamics in aqueous solutions with respect to that observed in 0.3 M NaCl solutions.

Cutin in tomato peels was depolymerized in methanolic base to yield cutin monomers or a mixture of cutin oligomers. These products were isolated by typical solvent extraction methods or by precipitation, and the isolates were characterized by chromatographic and spectroscopic analyses. It was determined that the compositions of the isolates from both isolation procedures were similar, although solvent extraction gave higher yields. However, the precipitation method, which is easy to carry out and avoids the use of undesirable organic solvents, may be preferable in commercial processes for recovering these compounds.

Pseudomonas flavescens strain B62 (NCPPB 3063) is a recently described bacterium isolated from walnut blight cankers. This strain has been designated as the type strain of a Pseudomonas rRNA group-I species. Strain B62 produced a mixture of two exopolysaccharides, differing in weight average relative molecular mass and composition. Only the most abundant exopolysaccharide (90% by mass), corresponding to the one with the lower molecular mass, was investigated by use of methylation analysis, partial acid hydrolysis, and NMR spectroscopy. The polysaccharide was depolymerised by the action of the cellulase produced by Penicillum funiculosum and the oligosaccharide obtained, corresponding to the repeating unit, was characterised by NMR spectroscopy and ion-spray mass spectrometry. The repeating unit of the B62 exopolysaccharide is [structure in text] where X is glucose (75%) or mannose (25%), and Lac is lactate. The O-acetyl groups are present only on 75% of the repeating units, and they are linked to the C6 of the hexose residues in non-stoichiometric amounts.

Zymomonas mobilis (ATCC 29191) was grown either aerobically or anaerobically in the presence of 2% (wt/vol) glucose and 0, 3, or 6% (vol/vol) ethanol. The rates of growth and the composition of hopanoids, cellular fatty acids, and other lipids in the bacterial membranes were quantitatively analyzed. The bacterium grew in the presence of 3% and 6% ethanol and was more ethanol tolerant when grown anaerobically. In the absence of ethanol, hopanoids comprised about 30% (by mass) of the total cellular lipids. Addition of ethanol to the media caused complex changes in the levels of hopanoids and other lipids. However, there was not a significant increase in any of the hopanoid lipid classes as ethanol concentration was increased. As previously reported, vaccenic acid was the most abundant fatty acid in the lipids of Z. mobilis, and its high constitutive levels were unaffected by the variations in ethanol and oxygen concentrations. A cyclopropane fatty acid accounted for 2.6-6.4 wt % of the total fatty acids in all treatments.

An acidic exopolysaccharide was isolated from P. fluorescens strain H13. The structure of the polysaccharide repeating unit was determined using chemical methods and 1D and 2D NMR techniques. The repeating unit was characterized as a trisaccharide composed of D-glucose, 2-acetamido-2-deoxy-D-glucose and 4-O-acetyl-2-acetamido-2-deoxy-D-mannuronic acid.

Pseudomonas marginalis is an important postharvest pathogen capable of causing soft rot in a wide variety of harvested fruits and vegetables. Following transposon mutagenesis, we isolated two groups of P. marginalis CY091 mutants deficient in production of pectate lyase (Pel) and soft-rot pathogenicity in plants. The first group, designated Pel-, was caused by the insertion of Tn5 into a pel structural gene, and the second group, designated LemA-, was caused by the insertion of Tn5 into a regulatory locus corresponding to the lemA gene previously identified in other Gram-negative bacteria. The LemA- mutants also exhibited alteration in colony morphology and showed deficiency in production of protease (Prt). A cosmid clone pCIC carrying the P. marginalis lemA gene was isolated and characterized. pCIC was capable of restoring Pel production and soft-rot pathogenicity in LemA- mutants of P. marginalis and Pseudomonas viridiflava, indicating that the function of lemA gene in these two pseudomonads was similar and interchangeable. Using MudI-mediated mutagenesis, we isolated a third group of P. marginalis mutants deficient in production of Pel, Prt, and soft-rot pathogenicity. Mutants in this group (designated GacA-1) contained an insertion of MudI in a locus corresponding to the gacA gene of P. viridiflava. Like LemA- mutants, GacA- mutants also exhibited alteration in colony morphology and showed deficiency in production of Pel and Prt. However, GacA- mutants produced much lower levels of levan and fluorescent pyoverdine siderophore than the wild type and LemA- mutants. These results provide the first genetic evidence that P. marginalis produces a single alkaline Pel for maceration of plant tissue and demonstrate that production of Pel, Prt, levan, and pyoverdin by this bacterium is mediated by the two-component lemA/gacA gene system.

The relevance of quantitative determinations of urinary deoxypyridinolines (DPY) and pyridinolines (PY), and of serum type I collagen carboxyterminal cross-linked telopeptides (ICTP), has been evaluated for patient monitoring in multiple myeloma (MM). In 178 untreated MM patients, a clear correlation was found between ICTP concentrations, bone destructions and serum calcium levels. Furthermore, serum ICTP, urinary DPY and PY concentrations were estimated before and during treatment in a further 33 MM patients randomly allocated to four groups receiving intravenous melphalan/prednisone (MivP) chemotherapy alone, or MivP in combination with three different doses of i.v. clodronate. 1800 mg of i.v. clodronate combined monthly with MivP induced a rapid and sustained reduction in bone resorption parameters to the normal range, a result not obtained with either MivP alone, or with a lower clodronate dose. While confirming the relevance of determining pyridinium cross-links for estimating bone resorption in MM, our data indicate that measurements of these parameters could be useful for dose finding and monitoring of bisphosphonate therapy.

It is the long-term goal of the German CML Study Group and of the Süddeutsche Hämoblastosegruppe (SHG) to improve survival of patients with chronic myelogenous leukemia (CML). In a first randomized study (CML Study I) monotherapies with hydroxyurea or interferon alpha (IFN-alpha) were compared with a standard busulfan regimen with regard to duration of the chronic phase and survival. The main results of this study were published, 1-3 and a long-term follow up is planned. In a second randomized study the effect of the combination of IFN-alpha and hydroxyurea versus hydroxyurea monotherapy on survival is being investigated. This paper provides a first preliminary report on the study concept, patient recruitment, state of documentation and initial patients' characteristics 9 months after closure of the study.

Purpose: The prognosis of patients with locally advanced esophageal cancer (LAEC) remains poor when treated with local modalities. An intensive preoperative program with chemoradiotherapy was used to evaluate the curative resection rate, pathologic response, and survival of patients with LAEC., Patients and Methods: Ninety patients with LAEC were treated preoperatively with chemotherapy (three courses of fluorouracil, leucovorin, etoposide, and cisplatin [FLEP]) followed by concurrent chemoradiotherapy (one course of cisplatin plus etoposide in combination with 40 Gy of radiation). Transthoracic esophagectomy was performed 4 weeks after the end of radiation., Results: Seventy-two patients were included in this evaluation. Forty-four (61%) underwent a complete tumor resection, and 16 (22%) had no tumor in the resected specimen (pathologic complete response [PCR]). The operative mortality rate was 15%. At a median follow-up time of 22 months (range, 12 to 41), the median survival duration of all 72 patients was 17 months (range, 1 to 41+). The calculated survival rates at 3 years were 33%, 42%, and 68% for all patients, patients after complete resection, and patients with PCR, respectively., Conclusion: This combined treatment modality is active in LAEC, with a PCR in 33% of the patients undergoing surgery. The results appear improved compared with those reported with surgery alone, by approximately doubling the 3-year survival rate. The high efficacy of preoperative chemoradiation warrants evaluation of the role of surgery in LAEC.

The structure of the acidic exopolysaccharide produced by the mushroom pathogen Pseudomonas "gingeri" strain Pf9, a bacterium which causes ginger blotch, was investigated by chemical analysis, mass spectrometry and 1D and 2D NMR spectroscopy. The polysaccharide consists of the linear trisaccharide repeating unit [formula: see text] where the cyclic pyruvic acetal groups at O-4 and O-6 of the mannopyranosyl residues have the S-configuration. Methylation analysis under neutral conditions and NMR data showed that the mannose residues are acetylated at O-2. This exopolysaccharide has the same structure as the E. coli K55 capsular polysaccharide and differs from the Klebsiella K5 capsular polysaccharide only in the position of acetylation (C-2 of the glucopyranose residue).

Objective: To evaluate potential changes in the ratio of whole-body/large-vessel hematocrit (f-cell ratio) during isovolemic hemodilution and to compare the volume effects of 2 different plasma exchange solutions (hydroxyethyl starch 200,000/0.62 6% and human albumin 5%)., Design: Prospective, randomized, controlled trial., Setting: Operating theater in a university hospital., Patients: 24 gynecological patients scheduled for elective surgery., Interventions: Isovolemic hemodilution was performed using 2 different plasma exchange solutions. Plasma volume was determined using dye dilution technique before and after hemodilution. The volume of withdrawn blood was measured from the change in weight of the blood bags taking into account the specific gravity of blood., Results: The volume of administered plasma exchange solutions exceeded the amount of withdrawn blood by 80 +/- 47 ml (p < 0.001). Plasma volume was 3,067 +/- 327 ml before and 3,517 +/- 458 ml after hemodilution. Using red cell volumes calculated from measured plasma volumes and peripheral hematocrit, a deficit of 249 +/- 133 ml (p < 0.0001) in red cells after hemodilution appeared with the measured withdrawn red cell volumes taken into account. This finding can be explained by a change in the f-cell ratio during isovolemic hemodilution. The volume effect of the exchange solutions was 1.05 for hydroxyethyl starch and 0.95 for albumin., Conclusions: The results demonstrate that a change in the f-cell ratio occurs during isovolemic hemodilution. The estimation of red cell volume or plasma volume changes by using either the hematocrit or plasma or red cell volume determinations together with the hematocrit may lead to erroneous results.

In vitro data have demonstrated autologous T-lymphocytes with anti-tumour activity in multiple myeloma (MM). Therefore a phase I/II trial was conducted to study the feasibility, the effect on several immunological parameters, and the tumour response induction of low-dose recombinant interleukin-2 (rIL-2) in MM patients. 18 MM patients of advanced stages in progress, who had failed on standard chemotherapy received 9 x 10(6) IU/m2 rIL-2 twice daily on days 1 and 2 and 0.9 x 10(6) IU/m2 twice daily for 5 subsequent days per week subcutaneously from days 3 to 56 (repeated every 12 weeks until progression). Patients were treated for between 8 and 1086 + d (mean 241 d) without serious side-effects. 6/17 patients experienced tumour response (2/17 objective tumour mass reduction, 4/17 long-lasting stable disease following tumour progression before initiation of rIL-2 treatment). During therapy the number of eosinophils increased 15-fold, CD4+ T lymphocytes were activated as demonstrated by enhanced CD25 antigen expression, and CD56+ NK cells expanded in the peripheral blood. Furthermore, a diminished pre-treatment ratio of CD4+/CD8+ lymphocytes was normalized during rIL-2 treatment. NK cell activity and lymphokine activated killer (LAK) cell activity was significantly enhanced. Endogenous IL-2 production and elevated soluble IL-2 receptor serum concentrations were induced. Low-dose rIL-2 can stimulate immune enhancement in MM despite the characteristic tumour-induced immunodeficiency. The treatment has proven though limited efficacy in advanced MM. Because most of the responders experienced termination of tumour progression rather than tumour regression, rIL-2 maintenance of chemotherapy-induced remissions should be investigated.

The acidic exopolysaccharides (EPSs) from 63 strains of mushroom production-associated fluorescent pseudomonads which were mucoid on Pseudomonas agar F medium (PAF) were isolated, partially purified, and characterized. The strains were originally isolated from discolored lesion which developed postharvest on mushroom (Agaricus bisporus) caps or from commercial lots of mushroom casing medium. An acidic galactoglucan, previously named marginalan, was produced by mucoid strains of the saprophyte Pseudomonas putida and the majority of mucoid strains of saprophytic P. fluorescens (biovars III and V) isolated from casing medium. One biovar II strain (J1) of P. fluorescens produced alginate, a copolymer of mannuronic and guluronic acids, and one strain (H13) produced an apparently unique EPS containing neutral and amino sugars. Of 10 strains of the pathogen "P. gingeri," the causal agent of mushroom ginger blotch, 8 gave mucoid growth on PAF. The "P. gingeri" EPS also was unique in containing both neutral sugar and glucuronic acid. Mucoid, weakly virulent strains of "P. reactans" produced either alginate or marginalan. All 10 strains of the pathogen P. tolaasii, the causal agent of brown blotch of mushrooms were nonnmucoid on PAF. Production of EPS by these 10 strains plus the 2 nonmucoid strains of "P. gingeri" also was negative on several additional solid media as well as in two broth media tested. The results support our previous studies indicating that fluorescent pseudomonads are a rich source of novel EPSs.

Hopanoids and other lipids were extracted from Zymomonas mobilis and quantitatively analyzed by high-performance liquid chromatography. Previous methods for hopanoid analysis required derivatization of the hopanoids via periodate oxidation or acetylation. The current method employs a normal-phase silica gel column, a ternary gradient of hexane-isopropanol-water-triethylamine, and detection with a flame ionization detector. Three major hopanoid classes were separated and quantified by this new method, and together they comprised 30 to 40% of the total lipid in these cells. Mass spectrometry confirmed that chemical structures of these hopanoids were consistent with those previously proposed. In addition, several other common lipid classes (free fatty acids and six classes of common phospholipids), comprising the remaining 60 to 70% of the total lipids, were also separated and quantified. This new chromatographic method represents the first technique for the analysis and purification of intact hopanoids. This method should be useful for the analysis and purification of hopanoids from other bacterial species.

406 untreated multiple myeloma patients of stage I (n = 54), II (n = 148) and III (n = 204) were enrolled in the trial. 51/54 stage I and 60/148 stage II patients were asymptomatic and followed without treatment until disease progression (progression free survival: 60% after 4 years for stage I versus 50% after 1 year for stage II). Symptomatic patients of stage I (n = 3/54) and II (n = 88/148) presenting with tumour progression, received melphalan 15 mg/m2 intravenously (i.v.) and prednisone 60 mg/m2 oral days 1-4 (MP). Stage II disease remission rate was 59%, and 50% tumour related survival (TRS) was 59 months. Stage III patients were randomised to receive MP or VBAMDex (vincristine/BCNU/doxorubicin/melphalan/dexamethasone) treatment. 43% of MP treated patients responded compared with 64% of the VBAMDex group. 50% TRS was 36 months in both groups without a detectable difference. 117 responders of stage II and III with stable disease were randomised to receive either IFN-alpha (5 x 10(6) IU, subcutaneous (S.C.) 3 times per week) or no maintenance treatment. The relapse rate in both groups was 50% after 13 months. No survival benefit for IFN alpha treated patients was observed (50% TRS: 45 months).

Four pleiotropic mutants of Pseudomonas viridiflava strain PJ-08-6A that were deficient in production of both pectate lyase (Pel) and protease (Prt) were isolated following transposon mutagenesis. Unlike secretion-defective (Out-) mutants, these four showed no accumulation of enzymes within the cells. Southern hybridization analysis revealed that each mutant had Tn5 inserted in one of two EcoRI genomic fragments. These EcoRI fragments (5.2- and 6.3-kb) appeared to contain two distinct gene loci, designated repA and repB, which were required for production of extracellular enzymes in this bacterium. Cosmid clones carrying the functional repA and repB DNA fragments were identified in a genomic library of strain PJ-08-6A. After analysis of repA+ plasmids by restriction mapping and marker-exchange mutagenesis, the repA gene was located in a joint region between the 1.8-kb EcoRI-HindIII and 2.8-kb EcoRI fragments cloned. Nucleotide sequence analysis of the repA region revealed the presence of an open reading frame consisting of 2,790 bases. The RepA protein predicted from the DNA sequence showed 93% similarity in amino acid sequence to the LemA protein of P. syringae pv. syringae, which was previously identified as a member of a two-component global regulatory system. A plasmid carrying the lemA gene of P. syringae pv. syringae was capable of complementing the RepA- mutation in P. viridiflava. The functions of the repA and lemA genes thus appear to be similar and interchangeable. Mutants of P. viridiflava strain SF312A deficient in production of Pel, Prt, and the exopolysaccharide alginate also were identified.(ABSTRACT TRUNCATED AT 250 WORDS)

Mid-latency auditory evoked potentials (MLAEP) reflect the primary cortical processing of auditory stimuli. They are suppressed widely during general anesthesia. Under ketamine, in contrast, MLAEP seem to be preserved. Ketamine exists in two optical isomers, S (+) ketamine and R (-) ketamine, which differ in their pharmacodynamic properties. S (+) ketamine has a higher anesthetic-hypnotic and analgesic potency than R (-) ketamine or the racemic mixture of S (+) ketamine and R (-) ketamine. In a blinded, randomized evaluation we compared the effect of induction of general anesthesia with the more potent ketamine compound--S (+) ketamine--to induction with the racemic ketamine on MLAEP in 60 patients scheduled for minor gynecologic procedures. Anesthesia was induced with S (+) ketamine (1 mg/kg Group I, n = 30) or an equi-anesthetic dose of racemic ketamine (2 mg/kg, Group II, n = 30). Auditory evoked potentials (AEP) were recorded before, during, and after induction of general anesthesia. Latencies of the peaks V, Na, Pa, Nb, and P1 and amplitudes Na/Pa, Pa/Nb, and Nb/P1 were measured. A fast-Fourier transform was used to calculate the power spectra of the AEP. The baseline MLAEP peaks of the awake patients were of normal amplitude and demonstrated a characteristic periodic wave form morphology. Power spectra indicated high energy in the 30-40 Hz frequency range. After induction of general anesthesia with S (+) ketamine or racemic ketamine, there was no increase in latencies of peaks V, Na, Pa, Nb, or P1. No decrease in amplitudes Na/Pa, Pa/Nb, or Nb/P1 could be observed. There was no significant change in the power spectra.(ABSTRACT TRUNCATED AT 250 WORDS)

Mid-latency auditory evoked potentials (MLAEP) reflect the primary cortical processing of auditory stimuli. They are widely suppressed during general anaesthesia with volatile anaesthetics. Under ketamine, in contrast, they seem to be preserved, which has been interpreted as indicating insufficient suppression of consciousness during ketamine anaesthesia. Ketamine exists in two optical isomeres, S-(+)-ketamine und R-(-)-ketamine, which differ in their pharmacodynamic properties: S-(+)-ketamine has higher anaesthetic-hypnotic and analgesic potency than R-(-)-ketamine. It thus appears obvious to question whether S-(+)-ketamine has a different effect on the primary cortical processing of sensory, i.e., auditory stimuli. We therefore studied the effects of S-(+)-ketamine versus ketamine-racemate on MLAEP. PATIENTS AND METHODS. Institutional approval and informed consent were obtained for 40 patients scheduled for minor gynaecological procedures. The patients were assigned randomly to one of the two experimental groups. All experimental evaluations were conducted under double-blind conditions. Anaesthesia was induced with S-(+)-ketamine 1 mg/kg (group I, n = 20) or ketamine-racemat 2 mg/kg (group II, n = 20). MLAEP were recorded before, during, and after induction of general anaesthesia from the vertex (positive) and mastoids on both sides (negative). Auditory clicks were presented binaurally at 70 dBnHL at a rate of 9.3 Hz. Using the electrodiagnostic system Pathfinder I (Nicolet), 1000 successive stimulus responses were averaged over a 100-ms poststimulus interval and analysed off-line. Latencies of the peak V, Na, Pa, Nb, P1, N1, and amplitudes Na/Pa, Pa/Nb, and Nb/P1 were measured. V belongs to the brainstem-generated potentials, which demonstrates that auditory stimuli were correctly transduced. Na, Pa, Nb, P1, and N1 are generated in the primary auditory cortex of the temporal lobe and are the electrophysiological correlate of the primary cortical processing of the auditory stimuli. A Fast-Fourier transformation calculated powerspectra of the AEP. RESULTS. In the awake state, AEP peak latencies were in the normal range. MLAEP had high amplitudes and a periodic wave form. Powerspectra indicated high energy in the 30-40-Hz frequency range. After induction of general anaesthesia with (S+)-ketamine or ketamine-racemat, there was no increase in the latencies of the peaks V, Na, Pa, Nb, P1, and N1. No decrease in amplitudes Na/Pa, Pa/Nb, or Nb/P1 could be observed. In the power spectra, frequencies in the range of 30-40 Hz retained high energy. CONCLUSIONS. MLAEP do not change in amplitude or latency during induction of general anesthesia with S-(+)-ketamine or ketamine-racemat. Primary cortical processing of auditory stimuli seems to preserved under S-(+)-ketamine and ketamine-racemat. This must be viewed in connection with dreams and hallucinations and could be interpreted as inadequate suppression of auditory information processing during general anaesthesia with S-(+)-ketamine and ketamine-racemat.

We investigated the feasibility of using indocyanine green (ICG) for plasma volume (PV) determination in man. Duplicate PV measurements were carried out in 23 healthy subjects to test repeatability. ICG (0.25 mg/kg) was injected intravenously into one arm and venous blood was withdrawn from the opposite arm. Optical density of plasma samples from minute 3 to 9 was measured in a densitometer. ICG concentration at injection time was determined by monoexponential extrapolation. The mean (SD) difference (MD) was -23 ml (183) or -0.6% (5.7%). Linear regression revealed PV2 = 0.92.PV1 + 226 (r = 0.92). The PV values corresponded well with data from other studies. In 26 surgical patients PV was determined using two methods: 1) the same as in healthy subjects and 2) using a modification of this method in whole blood (PVB). For PVB measurement blood was drawn through a cuvette-densitometer from an arterial line. Calculations were the same as in PV determination except for the use of hematocrit to achieve plasma concentrations of ICG from whole blood. In patients MD were -53 ml (144) or -1.3% (4.3) for PV and -19 ml (161) or -0.3% (5.1) for PVB. Comparing PVB and PV revealed MD = -113 ml (149) or -3.3% (4.2). The whole blood method is easier to perform and reduces blood waste to almost zero. In conclusion, ICG is a suitable tracer for PV determination.

Between October 1988 and October 1991, 104 patients with multiple myeloma and 6 with plasma cell leukaemia were studied cytogenetically. Abnormal karyotypes were found in bone marrow cells of 33 patients (30%). Most pathological karyotypes were complex with numerous modal and structural anomalies. Numerical anomalies most frequently involved chromosome 11 and structural aberrations occurred most often in chromosomes 1, 11 and 14. The most consistent structural aberration was a 14q+ chromosome (10 patients) resulting from a t(11;14)(q13;q32) in 4 patients and a t(8;14)(q24;q32) in 1 patient. Sequential cytogenetic studies were performed in 15 patients. In 5 of 8 cases with a normal karyotype at diagnosis, chromosomal anomalies were detected when disease progressed. In concomitant cytogenetic/cytological studies it was found that in the majority of patients with normal karyotype the mitoses originated from contaminating normal bone marrow cells. Pathological karyotypes were detected more frequently in pretreated than in untreated patients, in patients with plasma cell leukaemia than in patients with multiple myeloma, in patients with stage III and dense bone marrow infiltration than in patients with stage I. Patients with abnormal karyotype, irrespective if pretreated or not, had a significantly shorter median survival than those with normal karyotype. These findings suggest that karyotype is an independent prognostic factor in multiple myeloma.

Total genomic DNA of 13 pseudomonads representing rRNA homology groups I-IV were screened for sequences homologous to four Pseudomonas aeruginosa alginate (alg) genes by Southern hybridization. Biotinylated probes for three structural genes (algA, algC and algD) and one regulatory gene (algR1) were prepared. Genomic DNA of strains representing group I (P. syringae pv. glycinea, P. viridiflava and P. corrugata) hybridized with all four gene probes. Hybridizing fragments were of differing sizes, indicating that evolutionary divergence among group I members has occurred. P. corrugata has not been reported to synthesize alginate. Genomic DNA from representatives of groups II-IV gave no or very weak hybridization with the probes except for algC. This study indicates that the ability to produce alginic acid as an exopolysaccharide among the pseudomonads is restricted to members of rRNA homology group I in agreement with earlier physiological studies.

Two hundred thirty-two nonfilamentous bacterial strains, including saprophytes, plant pathogens, and opportunistic plant and human pathogens, were screened for the ability to produce cutinases (cutin-degrading esterases). Initially, esterase activity of culture filtrates of strains grown in nutrient broth-yeast extract medium supplemented with 0.4% apple or tomato cutin was determined by a spectrophotometric assay utilizing the model substrate p-nitrophenyl butyrate. The culture filtrates of the 10 Pseudomonas aeruginosa strains tested exhibited the highest esterase activity, with values of >500 nmol/min/ml. Of these 10 strains, 3 (K799, 1499A, and DAR41352) demonstrated significant induction (10-fold or above) of esterase activity by addition of cutin to nutrient broth-yeast extract medium. The ability of culture filtrates of the three strains to cause release of apple cutin monomers was confirmed by a novel high-performance liquid chromatography technique. Monomer identification was confirmed by gas chromatography-mass spectroscopy analyses. Addition of the nonionic detergent n-octylglucoside stimulated cutinase activity of culture filtrates from strains K799 and DAR41352, but not that of filtrates from strain 1499A. Time course studies in nutrient broth-yeast extract medium supplemented with apple cutin indicated maximal levels of cutinase in the culture fluids after cultures entered stationary phase. Incubation temperatures below the optimal temperature for growth (37 degrees C) led to maximal production of cutinase.

In previous studies workers have shown that three yeast strains (strains US-7, 82, and 101) have biological control activity against various postharvest fungal pathogens of fruits and vegetables, including Penicillium rots of apples and citrus and Botrytis rot of apples. In these reports the researchers have described these strains as Debaryomyces hansenii (anamorph, Candida famata) or Candida sp. strains. In this study we performed additional physiological, DNA reassociation, and mannan characterization tests that clearly established a new taxonomic classification for these strains, Candida guilliermondii. We also propose amendment of the physiological test profile in the taxonomic description of C. guilliermondii.

The structure of an acidic exopolysaccharide of two strains of Pseudomonas marginalis, a bacterium which causes soft rots of various vegetables, has been determined to consist of a repeating unit of: ----4) beta-D-Manp-(1----3)alpha-D-Glcp-(1----4)alpha-L-Rhap-(1-. The glucose is pyruvated at O-4 and O-6 and the mannose is acetylated at either O-2 or O-3.

Chromosomal DNA from group I Pseudomonas species, Azotobacter vinelandii, Azomonas macrocytogens, Xanthomonas campestris, Serpens flexibilis, and three enteric bacteria was screened for sequences homologous to four Pseudomonas aeruginosa alginate (alg) genes (algA, pmm, algD, and algR1). All the group I Pseudomonas species tested (including alginate producers and nonproducers) contained sequences homologous to all the P. aeruginosa alg genes used as probes, with the exception of P. stutzeri, which lacked algD. Azotobacter vinelandii also contained sequences homologous to all the alg gene probes tested, while Azomonas macrocytogenes DNA showed homology to all but algD. X. campestris contained sequences homologous to pmm and algR1 but not to algA or algD. The helical bacterium S. flexibilis showed homology to the algR1 gene, suggesting that an environmentally responsive regulatory gene similar to algR1 exists in S. flexibilis. Escherichia coli showed homology to the algD and algR1 genes, while Salmonella typhimurium and Klebsiella pneumoniae failed to show homology with any of the P. aeruginosa alg genes. Since all the organisms tested are superfamily B procaryotes, these results suggest that within superfamily B, the alginate genes are distributed throughout the Pseudomonas group I-Azotobacter-Azomonas lineage, while only some alg genes have been retained in the Pseudomonas group V (Xanthomonas) and enteric lineages.

Eighteen plant-pathogenic and three non-plant-pathogenic pseudomonads were tested for the ability to produce alginic acid as an exopolysaccharide in vitro. Alginate production was demonstrated for 10 of 13 fluorescent plant-pathogenic pseudomonads tested with glucose or gluconate as the carbon source, but not for all 5 nonfluorescent plant pathogens and all 3 non-plant pathogens tested. With sucrose as the carbon source, some strains produced alginate while others produced both polyfructan (levan) and alginate. Alginates ranged from <1 to 28% guluronic acid, were acetylated, and had number-average molecular weights of 11.3 x 10 to 47.1 x 10. Polyfructans and alginates were not elicitors of the soybean phytoalexin glyceollin when applied to wounded cotyledon surfaces and did not induce prolonged water soaking of soybean leaf tissues. All or most pseudomonads in rRNA-DNA homology group I may be capable of synthesizing alginate as an exopolysaccharide.

The exopolysaccharide of Pseudomonas marginalis HT041B has been characterized as a 1,3-linked galactoglucan in which galactose and glucose are in the alpha- and beta-anomeric configurations, respectively. The polysaccharide is substituted with pyruvate at the 4 and 6 positions of galactose and with succinic acid at either the 2 or 4 position of glucose. This polysaccharide has been given the trivial name marginalan.