Occurrence and hygienic relevance of fungi in drinking water.

Fungi, above all filamentous fungi, can occur almost everywhere, even in water. They can grow in such a quantity in water that they can affect the health of the population or have negative effects on food production. There are several reports of fungal growth in water from different countries, but to our knowledge none from Austria so far. The aim of this study was to gain an overview of the spectrum of filamentous fungi and yeasts in drinking water systems. Thirty-eight water samples from drinking water and groundwater were analysed. Fungi were isolated by using membrane filtration and plating method with subsequent cultivation on agar plates. The different taxa of fungi were identified using routine techniques as well as molecular methods. Fungi were isolated in all water samples examined. The mean value for drinking water was 9.1 CFU per 100 ml and for groundwater 5400 CFU per 100 ml. Altogether 32 different taxa of fungi were found. The taxa which occurred most frequently were Cladosporium spp., Basidiomycetes and Penicillium spp. (74.6%, 56.4% and 48.7%, respectively). This study shows that drinking water can be a reservoir for fungi, among them opportunists, which can cause infections in immunosuppressed patients.

Isolation of Gordonia terrae from a patient with catheter-related bacteraemia.

A cornyeform bacterium was isolated from a blood culture from a 24-year-old man with familial hypertrophic non-obstructive cardiomyopathy, chronic abuse of anabolic steroids and prior admission to hospital because of clinical signs of sepsis. 16S rRNA gene analysis unambiguously identified Gordonia terrae.

Bioavailability and pharmacokinetics of Echinacea purpurea preparations and their interaction with the immune system.

Echinacea is a widely used herbal remedy for the prevention and treatment of the common cold. Recently, many new insights concerning the molecular mode of action of the main lipophilic constituents, the alkamides, have renewed interest in this plant. In order to compare the bioavailability of alkamides from liquid and tablet preparations of E. purpurea (Echinaforce) in humans and to study the effects on ex vivo stimulated blood cells, a randomized, single-dose, crossover study with 10 (8 test, 2 placebo) volunteers has been performed. They received either 4 ml of the standardized E. purpurea (Echinaforce) tincture or 12 E. purpurea (Echinaforce) tablets or placebo. Both doses contained the same amount (0.07 mg) of the major alkamides, dodeca-2E,4E,8Z, 10E/Z-tetraenoic acid isobutylamides. Liquid chromatography electrospray ionization ion-trap mass spectrometry was used to determine the content of alkamides in serum. It was found that the arithmetic mean C(max) of dodeca-2E,4E, 8Z,10E/Z-tetraenoic acid isobutylamides absorbed after oral application of the Echinaforce tincture appeared after 30 min (0.40 ng/ml serum). In comparison, the t(max) of tablets was 45 min with a C(max) of 0.12 ng/ml. An ex vivo stimulation of blood by LPS was carried out to measure the influence of E. purpurea on the innate and adaptive immune system. Both E. purpurea preparations led to the same effects on the immune system according to the concentration of pro-inflammatory cytokines TNF-alpha and IL-8. 23 hours after oral application a significant down-regulation of TNF-alpha and IL-8 in LPS pre-stimulated whole blood was found. However, no significant changes in the concentration of IL-6 were observed. Although a quarter of the dodeca-2E,4E,8Z, 10E/Z-tetraenoic acid isobutylamides was absorbed from the tablets, the study shows that the formulations trigger the same effects on the measured immune parameters.

Cytomegalovirus diagnosis in renal and bone marrow transplant recipients: the impact of molecular assays.

BACKGROUND: Cytomegalovirus (CMV) infections are a major threat in transplant recipients. In recent years, new assays for routine CMV diagnosis, based on molecular techniques, have become available. OBJECTIVE: The impact of molecular assays for CMV diagnosis in transplant recipient was evaluated. STUDY DESIGN: A total of 51 transplant recipients were screened for CMV infection. Serological (AxSYM CMV IgG and recombinant CMV IgM assays), antigenemia, CMV DNA (qualitative in house PCR and the quantitative COBAS AMPLICOR CMV MONITOR Test), and CMV mRNA (NucliSens CMV pp67 Test) tests were compared. RESULTS: In 11/20 bone marrow transplant (BMT) recipients and 10/31 renal transplant (RTX) recipients there was no evidence of active CMV infection. Ten RTX recipients and one BMT recipient were antigenemia positive, 21 RTX and seven BMT recipients were PCR positive (qualitative CMV PCR). There were more BMT recipients CMV DNA positive in serum (7/21) than antigenemia positive (1/21). CMV mRNA was found positive in two BMT recipients (one case with no other evidence of CMV infection, the other one CMV DNA positive and antigenemia negative). The only antigenemia positive BMT recipient was found negative for CMV mRNA, but positive in all other tests. Eight RTX recipients were found positive for CMV mRNA. Six of them were also antigenemia positive and five of those were also found positive for CMV IgM. One CMV mRNA positive RTX recipient was CMV IgM positive but antigenemia negative and the other one CMV mRNA positive RTX recipient was found negative in all other tests. Two antigenemia positive RTX recipients were found negative for mRNA and CMV IgM. CONCLUSION: Antigenemia was found to be a good screening test for CMV infection in RTX recipients. In BMT recipients, tests based on molecular techniques appeared to be superior compared to antigenemia.

Sialic acid as a marker for differentiation between benign and malignant intracranial tumors.

One of the characteristics of malignant tumor cells is the production of glycocompounds with a high content of N-acetylneuraminic acid (sialic acid). Total sialic acid concentration was determined in the serum of 136 patients with an intracranial tumor. The concentration was determined enzymatically and using HPLC. Both methods had a relatively high specificity (90%), provided an inflammatory process due to an infection could be excluded. Sensitivity was 72.6%; the cut-off level of sialic acid concentration was determined to be 2.75 mumol/ml. There was a significant difference in the average sialic acid concentration of benign and malignant tumors. The test was least reliable in determining whether the growth of an astrocytoma is malignant.

Distribution of aflatoxin B1 in tissues of mink (Mustela vison).

Seven female mink (Mustela vison) were injected intraperitoneally with a single dose of 100 mug aflatoxin B1 (14 C-label and unabeled). They were sacrificed 1, 2, 4, and 24 h after dosing. Liver, instestines, stomach, lung, kidney, brain, pancrease, spleen, urinary bladder, uterus, and bile were removed and examined for the retained radioactivity. 1 h after dosing, intestines and their contents retained the largest amount of 14C-radioacivity (18.9% of the amount that was administered) which was followed by liver (13.2%) and the bile (10.8%). At this time all other tissues retained less than 1% of the administered radioactivity. Generally, the amount of radioactivity retained in all tissues declined with time. Only 1.2 and 0.6% of the administered radioactivity was found in testines and bile, respectively, 24 h after dosing; however, the liver still contained 6.6% of the initial radioactivity. Examination of subcellular fractions of liver revealed that at all time intervals most of the radioactivity was associated with the micrososmal supernatant fluid.

"New" food-borne pathogens of public health significance.

Recent work by epidemiologists and microbiologists has uncovered several hitherto unrecognized food-borne bacterial pathogens of public health significance. One of these, Listeria monocytogenes, has attracted considerable attention because of two major cheese-related outbreaks of listeriosis that were characterized by cases of meningitis, abortion, and perinatal septicemia. Thus far, L. monocytogenes has been responsible for well over 300 reported cases of food-borne listeriosis, including about 100 deaths, and has cost the dairy industry alone more than 66 million dollars as a result of product recalls. The ability of L. monocytogenes to grow at refrigeration temperatures, coupled with appearance of the pathogen in raw and processed meats, as well as poultry, vegetables, and seafood, makes this bacterium a serious threat to susceptible consumers and to the entire food industry. Yersinia enterocolitica, another psychrotrophic food-borne pathogen of recent concern, was linked to several outbreaks of yersiniosis associated with consumption of both raw and pasteurized milk, as well as contaminated water. Food-borne infections involving Y. enterocolitica typically result in enterocolitis, which may be mistaken for acute appendicitis. Unfortunately, inadvertent removal of healthy appendixes from victims of food-borne yersiniosis is all too common. Although known for many years, Campylobacter jejuni has only recently been recognized as a food-borne pathogen and a leading cause of gastroenteritis in the United States. Notable outbreaks of campylobacteriosis linked to consumption of raw milk, cake icing, eggs, poultry, and beef have underscored the need for thorough cooking and proper handling of raw products.(ABSTRACT TRUNCATED AT 250 WORDS)

A cohort study with children living in an air-polluted region--a model for public health.

Regions with heavy industry are in many ways regions of crisis. The health of the population is primarily affected by the different air pollutants. Dust, with all its organic (dioxins and furans) and inorganic (heavy metals) contents, makes up the greatest part of the air-borne pollutants. The influence on health of environmental pollution was ascertained through the determination of different parameters (functional methods and determination of physiological parameters). This influence could be observed in children over a period of 8 years with regular investigations (e.g. determination of pulmonary function by spirometry and immunological parameters). Besides this exogenous load the persons are exposed to other environmental stresses-shift work, unemployment, alcoholism and divorce-which have a particular influence on the attitude and the upbringing of the children. Sixty per cent of the children in this polluted region ate no breakfast in the morning. Consequently it could be shown that the blood sugar in 70% of the children was below 70 mg/dl. Additionally, a relatively high amount of COHb (2.5% to 3%), and an increased concentration of serum IgE (47% of children with a concentration over 100 IU/ml), could be detected. Through a change in the environmental awareness of the children and their consequent influence, an effort should be made to achieve a positive effect on the health of the whole population. The children were given a chance to participate in various sports for the whole day during a week in the mountains at 1200 m. The teachers exercised with the children for at least 8 h per day. Besides gymnastics the program consisted of downhill and cross-country skiing. In addition, the children were offered a balanced and natural diet and they were instructed accordingly. This week of activity led to a clear reduction of the concentration of COHb, but to a far less clear improvement in the concentration of blood sugar and the pulmonary function.

Analysis of hsp70 mRNA levels in HepG2 cells exposed to various metals differing in toxicity.

Human hepatoma cells (HepG2) were exposed to several heavy metal salts and the induction of heat shock protein 70 (hsp70) mRNA was analysed by the reverse transcriptase-polymerase chain reaction (RT-PCR). Metals were added to the cell medium at concentrations ranging from 0.1 to 100 microM and incubation was continued for 4 h. In addition we analysed the time dependence of hsp70 induction by adding each metal at a certain concentration followed by an incubation for 0.5 to 24 h. CdCl2, NaAsO2, AgNO3 could be classified as very strong inducers (20-, 13- and 10-fold above control level) and they reached their maximum level of induction at 1-10 microM after 2 h. CuCl2, MnCl2, Pb(NO3)2, TlNO3, CoCl2 and NiCl2 were also strong inducing agents, giving a 4-6 fold induction at 10-100 microM after 4-8 h. ZnSO4, Hg(NO3)2 and AlCl3 were only weak inducers (1.5-2 fold at 50-100 microM after 4-8 h) of hsp70 mRNA. Cytotoxic effects (measured by release of lactate dehydrogenase) could only be detected for 100 microM Hg2+ after 4 h and when the cells were incubated with 5 microM Cd2+ for more than 8 h. We also tested a few combinations of these heavy metal salts for their hsp70-inducing ability. Zn2+ and Mn2+ were able to diminish Cd2+ induced hsp70 mRNA levels by 65%. Ag+ mediated induction was reduced by 40% when combined with Cu2+, whereas Hg2+ increased induction by Ag+ about 3-fold and led to a dramatic decrease in cell viability. In our study we were able to demonstrate that the analysis of hsp70 mRNA levels in chemically stressed HepG2 cells by RT-PCR can be a valuable tool for studying mechanisms of toxicity associated with elevated expression of hsp70.

Total serum IgE concentration of children from air-polluted regions.

Distributions of IgE levels were determined from 312 children (divided into three age groups), living in small Austrian towns with different levels of air pollution. The spread of IgE concentration was extremely wide; the mean value was 124 kU/I with a standard deviation of 240. An increase of concentration was found in the group aged 6 to 11 years. No significant difference was found according to sex and the higher rates of IgE. The reference rates were then used to compare children living in an industrial region with heavy metal production. A significant difference was found in the lower IgE concentrations (25th and 50th percentiles).

Borrelia burgdorferi: another cause of foodborne illness?

Borrelia burgdorferi was identified as the etiological agent of Lyme disease in 1982. This Gram-negative spirochete is classified in the order Spirochaetales and the family Spirochaetaceae. The pathogen is fastidious, microaerophilic, mesophilic and metabolises glucose through the Embden-Meyerhof pathway. A generation time of 11 to 12 h at 37 degrees C in Barbour-Stoenner-Kelly medium has been reported. Lyme disease, named after Lyme in Connecticut, is distributed globally. It is the most commonly reported vector-borne disease in the United States, where the incidence is highest in the eastern and midwestern states. Since establishment of national surveillance in 1982, there has been a nine-fold increase in the number of cases reported to the U.S. Centers for Disease Control. The deer tick of the genus Ixodes is the primary vector of Lyme borreliosis. The tick may become infected with B. burgdorferi, by feeding on an infected host, at any point in its 2-year life cycle which involves larval, nymphal and adult stages. The infection rate in deer ticks may be as high as 40% in endemic areas. The primary vertebrate reservoirs for Ixodes are the white-footed mouse (Peromyscus leucopus) and the white-tailed deer (Odocopileus virginianus). Dairy cattle and other food animals can be infected with B. burgdorferi and hence some raw foods of animal origin might be contaminated with the pathogen. Recent findings indicate that the pathogen may be transmitted orally to laboratory animals, without an arthropod vector. Thus, the possibility exists that Lyme disease can be a food infection. In humans, the symptoms of Lyme disease, which manifest themselves days to years after the onset of infection, may involve the skin, cardiac, nervous and/or muscular systems, and so misdiagnosis can occur.

Sites of action by propionate on Listeria monocytogenes.

Exposure of Listeria monocytogenes to a solution of sodium propionate (8% w/v) for 60 min caused 87% of the population to be injured. Injury was evidenced by inability of the bacterium to tolerate 6% sodium chloride in tryptose agar as compared to ability to grow on tryptose agar with no added salt. Injured cells were allowed to repair in tryptose broth and the repair process was studied by addition to tryptose broth of sublethal amounts of metabolic or synthetic inhibitors. Repair of injured cells did not require electron transport or synthesis of cell wall components, mRNA or protein. No changes which may have occurred in the cell membrane of injured cells, allowed leakage of proteins or nucleotides into the medium. Exogenous cation salts enhanced the rate of recovery of injured cells. The specific activity of lactic dehydrogenase was reduced in propionate-injured L. monocytogenes.

Behavior of Listeria monocytogenes in the presence of sodium propionate.

Survival or growth of Listeria monocytogenes in Tryptose Broth supplemented with 0, 0.05, 0.1, 0.15, 0.2, 0.25 or 0.3% sodium propionate was determined when the pH of the medium was 5.0 or 5.6 and incubation was at 4, 13, 21 and 35 degrees C. The pathogen grew in all controls, propionate-free broth, except at 4 degrees C and pH 5.0. At pH 5.6 and 4, 13, 21 and 35 degrees C the bacterium grew in the presence of all propionate concentrations used in this study. The higher concentrations permitted only minimal growth with smallest ultimate populations and longest generation times. Reducing the pH to 5.0 served to minimize growth further at 13, 21 and 35 degrees C than that observed at the same temperatures but at pH 5.6. The extent of growth was directly proportional to the propionate concentrations; at high concentrations, propionate caused a gradual decrease in populations and/or prolonged the lag phase. At 35 degrees C, a concentration of 0.25% did not allow growth, whereas 0.3% caused inactivation of the pathogen after 80 h of incubation. At 4 degrees C and pH 5.0, all concentrations of sodium propionate caused a gradual decrease in populations during the incubation period.

Evaluation of 1,3-pentadiene for mutagenicity by the Salmonella/mammalian microsome assay.

1,3-Pentadiene, a food contaminant produced by some molds when they metabolize sorbic acid, was tested for mutagenicity, using variations of the Salmonella/mammalian microsome assay. The chemical was incorporated into the test system (with and without S9 mix) by 3 methods: (a) the standard plate incorporation assay, (b) a liquid preincubation procedure and (c) exposure of test bacteria in the soft agar overlay to gaseous 1,3-pentadiene. The chemical was extremely toxic to the test bacteria with amounts as low as 2.0 microgram/plate causing cell death. However, none of the nonlethal concentrations tested by any of the methods was mutagenic to Salmonella typhimurium strains TA97, TA98, TA100, TA1535, TA1537 or TA1538.

Diarrhea in returning Austrian tourists: epidemiology, etiology, and cost-analyses.

BACKGROUND: Between 1995 and 1997, stool samples of 322 Austrian tourists returning from abroad with diarrhea were examined for bacteria, parasites and viruses. METHODS: Epidemiologic data were collected from information furnished by physicians and hospitals and from questionnaires. Moreover, testing expenses and additional cost for treated cases were evaluated. RESULTS: In 97 of 322 patients examined (30%), one or more pathogens were detected in the stool. Bacteria were found in 38 patients (39%), parasites in 33 patients (34%) and viruses in 26 patients (27%). In 6 patients, mixed infections with parasites and viruses were detected and in 5 patients with bacteria and viruses. Among bacteria, Campylobacter jejuni was most frequent; among parasitic infections, Giardia lamblia. Significant correlations were established between the country of destination, age, travel style and length of stay. Forty-four percent of all patients visited Asia (including Turkey), 27% Africa, 18% Latin America, and only 10% southern Europe. The group between 20 and 29 years of age was most frequently affected (p<.001), the group between 0 and 19 years of age least. Fifty-seven percent stayed in a hotel without frequent changes of location; 43% undertook a trekking trip; and of those, 75% belonged to the group aged between 20 and 39. In terms of the correlation between travel style and pathogen, it was found that 74% of patients with bacterial infections stayed in a hotel (avg. 57.9%; p<.05) whereas 64% of all patients with parasitic infections undertook a trekking trip (avg. 42%; p<.001). Thirty-six percent of all patients with parasitic infections spent their vacation in India (avg. 13%; p<.001). The length of stay of patients with bacterial infections was shorter than average (72% spent between 1 and 2 weeks abroad, avg. 49.8%). Patients with parasitic infections spent significantly more time abroad than average (42% more than 2 months; avg. 17.7%; p<.001). Average cost of specific antimicrobial therapy was U.S.$31 whereas the average cost of identifying a patient needing such treatment was almost U.S.$580. CONCLUSION: Optimal detection rate and cost reduction for the diagnosis require precise history, adequate collection of samples using adequate transport media, and rapid transfer to the laboratory.

Antibiotic resistance of E. coli in sewage and sludge.

The aim of the study is the evaluation of resistance patterns of E. coli in wastewater treatment plants without an evaluation of basic antibiotic resistance mechanisms. Investigations have been done in sewage, sludge and receiving waters from three different sewage treatment plants in southern Austria. A total of 767 E. coli isolates were tested regarding their resistance to 24 different antibiotics. The highest resistance rates were found in E. coli strains of a sewage treatment plant which treats not only municipal sewage but also sewage from a hospital. Among the antimicrobial agents tested, the highest resistance rates in the penicillin group were found for Ampicillin (AM) (up to 18%) and Piperacillin (PIP) (up to 12%); in the cephalosporin group for Cefalothin (CF) (up to 35%) and Cefuroxime-Axetil (CXMAX) (up to 11%); in the group of quinolones for Nalidixic acid (NA) (up to 15%); and for Trimethoprime/Sulfamethoxazole (SXT) (up to 13%) and for Tetracycline (TE) (57%). Median values for E. coli in the inflow (crude sewage) of the plants were between 2.0 x 10(4) and 6.1 x 10(4)CFU/ml (Coli ID-agar, BioMerieux 42017) but showed a 200-fold reduction in all three plants in the effluent. Nevertheless, more than 10(2)CFU E. coli/ml reached the receiving water and thus sewage treatment processes contribute to the dissemination of resistant bacteria in the environment.

Improved method for the fluorimetric detection of beta-D-galactosidase in water.

A very convenient method to quantify coliform bacteria in water can probably be designed via the determination of the activity of the enzyme beta-D-galactosidase, whose natural occurrence is, apart from less frequently occurring aeromonads mainly restricted to this type of microorganisms. 4-methylumbelliferyl-beta-D-galactoside is used as substrate, which is hydrolyzed during the enzymatic reaction; the released 4-methylumbelliferone can be quantified fluorimetrically. In the present study the influence of various physical and chemical parameters on the determination is investigated and the experimental conditions are optimized. Most important entities are the pH value during hydrolysis, the presence of nutrients and co-factors in the sample, and the modification of the substrate. Statistical evaluation of the results obtained by changing single or multiple parameters reflects clearly their positive or negative influence on the enzyme activity. Thus, deliberate addition of surfactants, specific nutrients, salts and co-enzymes results in a significantly increased activity of beta-D-galactosidase towards the substrate, which can be advantageously exploited to increase the sensitivity of the analytical method together with a decrease of the detection limit. The influence of the parameters and the optimized conditions of the improved analytical methods are presented.

Experimental acute aflatoxicosis in mink (Mustela vison).

Susceptibility of mink to intoxication by aflatoxins was studied by giving mink single doses (300, 600, and 900 mug of aflatoxins B1 and G1 (40:60)/kg of body weight) and observing them for 8 weeks. One, 2, and 4 of 5 mink in each group died within 4 days after ingesting the small, medium, and large doses, respectively. Enlarged liver with pale yellow to yellowish pink spots was the most consistent lesion observed in mink that died of acute aflatoxicosis. Some lobules of liver also appeared hemorrhagic and fragile, whereas others showed fatty metamorphosis. Histopathologic examination of the liver showed different degrees of fat infiltration, bile duct proliferation, and necrosis of liver cells. Hematologically, mink that survived were not appreciably different from control mink in terms of hemoglobin content, packed cell volume, and white blood cell count. Liver specimen from aflatoxin-treated mink contained more fat, was larger, and contained less protein, RNA and DNA than did liver from control mink. Aflatoxin residue (only B1 ) was recovered from liver of 6 mink and were between 0.1 and 7.6 mug of the original dose.

Mortality and some biochemical changes in mink (Mustela vison) given sublethal doses of aflatoxin each day.

Two feeding trials were done to study the susceptibility of mink (Mustela vison) to multiple sublethal doses of aflatoxins. In the 1st trial, twenty 3-month-old male mink were divided equally among groups. Each mink in groups 1, 2, 3, and 4 was given a meatball daily that contained 15, 30, 45, or 0 mug of aflatoxins (B1:G1, 40:60), respectively. All mink in group 3 died between the 25th and the 30th days of the feeding trial. Each mink had ingested 1,035 to 1,480 mug of aflatoxins. Four of the mink in group 2 died almost as soon as did mink in group 3. Four mink in group 1 died between 40 and 59 days after the start of the feeding trial. Generally, a marked increase in plasma cholesterol and alkaline phosphatase activity appeared before mink died. The liver from animals that died of aflatoxicosis showed prominent pathologic changes which included hemorrhages and appearance of pink yellow spots. Histopathologic examination of liver from dead mink revealed fatty infiltration, bile duct proliferation, bile stasis, pseudotubular formation, congestion, and fibrosis. The feeding trial was repeated with 20 mink (8 males and 12 females) that were 1.5 to 2 years old. In this instance, 0, 20, 40, and 60 mug of aflatoxins were administered each day. All treated animals, except 1, were dead within 37 days after the experiment started. The survivor was given the lowest dosage of toxins and died after 52 days by which time 960 mug of aflatoxins were consumed. Plasma cholesterol content and alkaline phosphatase activity generally were similar to those observed in younger mink of the 1st feeding trial.

Growth of mink (Mustela vison) kits when nursing dams given aflatoxin G1.

During lactation, 5 female mink (Mustela vison) were given 50 mug of aflatoxin B1 orally twice, 2 weeks apart. Another group of 5 lactating mink were used as controls, Milk was collected 24 hours after each treatment with toxin. Aflatoxin B1 and M1 could not be detected in the milk. There was no appreciable difference in body weight among kits that nursed treated mink and those that nursed untreated mink.