[Campylobacteriosis outbreaks in the state of Hesse, Germany, 2005-2011: raw milk yet again]. / Campylobacteriose-Ausbrüche in Hessen, 2005-2011 - und immer wieder Rohmilch.

BACKGROUND: Campylobacter is the most frequently reported cause of acute infectious diarrhea in Germany. Campylobacter outbreaks are rare events. However, their investigation provides useful information on risks of infection and unused prevention potentials. METHODS: We analyzed the Hessian database for notifiable diseases for cases of campylobacteriosis reported from 2005 through 2011. For campylobacter outbreaks including five or more cases we prospectively obtained additional information from local public health authorities. RESULTS: From 2005 through 2011, 29,473 cases of campylobacteriosis were reported in Hesse, Germany (approx. 6 million inhabitants), yielding an annual incidence ranging from 53.4 to 81.4 cases per 100,000 inhabitants. Only 236 cases were part of 16 outbreaks with five or more cases. Among these, six outbreaks occurred among groups traveling outside Germany, four were associated with the consumption of raw milk. For eight outbreaks consumption of poultry was considered a probable or - based on the frequent consumption of poultry during group travel - possible vehicle of infection. Two of the raw-milk associated outbreaks were reported among two groups who visited the same farm within 18 days. Five of 14 members of several families and 77 of 117 students fell sick. The local public health authority was only informed when both groups had visited the farm. CONCLUSION: The reported outbreaks can be attributed to known risk factors for campylobacteriosis - consumption of raw milk and poultry and international travel. This underlines that prevention possibilities are insufficiently used. These include avoiding the consumption of unpasteurized milk and milk products, the hygienically correct handling of raw poultry and timely identification and notification of outbreaks to public health authorities.

Secondary transmissions during the outbreak of Shiga toxin-producing Escherichia coli O104 in Hesse, Germany, 2011.

During the recent outbreak of Shiga toxin-producing Escherichia coli (STEC) O104:H4 in Germany most cases notified in the State of Hesse (6 million inhabitants) were linked to satellite clusters or had travelled to the outbreak area in northern Germany. Intensified surveillance was introduced to rapidly identify cases not linked to known clusters or cases and thus to obtain timely information on possible further contaminated vehicles distributed in Hesse, as well to describe the risk of secondary transmission among known cases. As of 2 August 2011* [corrected], 56 cases of haemolytic uraemic syndrome (HUS) including two fatal cases, and 124 cases of STEC gastroenteritis meeting the national case definitions have been reported in Hesse. Among the 55 HUS and 81 STEC gastroenteritis cases thatmet the outbreak case definition, one HUS case and eight STEC gastroenteritis cases may have acquired their infection through secondary transmission. They include six possible transmissions within the family, two possible nosocomial and one possible laboratory transmission. Our results do not suggest an increased transmissibility of the outbreak strain compared to what is already known about E. coli O157 and other STEC serotypes.

The impact of different biocompatible coated cardiopulmonary bypass circuits on inflammatory response and oxidative stress.

This study was to compare the impact of different biocompatible coated circuits on inflammatory response and oxidative stress induced during cardiopulmonary bypass (CPB). Seventy-eight patients undergoing elective coronary artery bypass grafting (CABG) with CPB were randomly assigned to five groups with different biocompatible coated circuits: Trillium, Bioline, Phosphorylcholine, Polymethoxyethyl acrylate (PMEA), and the uncoated control group. Blood was drawn at three different time points: before CPB, 6 and 72 hours post CPB. Unlike the Trillium group, serum levels of TNF-alpha in the Bioline and Phosphorylcholine groups significantly increased only at 72 hours post CPB (p < 0.05). Serum levels of IL-6 significantly increased at 6 and 72 hours post CPB in all groups (p < 0.01). The Trillium group showed a significant increase of IL-10 compared to the control group at 72 hours post CPB (p < 0.05). Serum levels of NOx in the Phosphorylcholine group significantly decreased at 6 hours post CPB compared to baseline (p < 0.05). Both the Bioline and Phosphorylcholine groups showed statistical decreases in serum NOx levels compared with other groups at 6 hours post CPB (p < 0.05). A significant difference in NOx levels between the Bioline and the control group was also observed at 72 hours post CPB. Myeloperoxidase levels were significantly elevated at 6 and 72 hours post CPB in all groups (p < 0.05). Inflammatory response and oxidative stress are elevated during CABG with CPB. Heparin-coated and the Phosphorylcholine-coated circuits induce less inflammatory responses and oxidative stress compared to other circuits.

Cardiac injury markers in non-elite marathon runners.

An elevation of cardiac injury markers including creatinine kinase (CK), myoglobin (Myo) and cardiac troponin T (cTnT) has been observed in elite athletes following strenuous exercise. The mechanism and significance of this observation however have not been fully elucidated. The goals of this study were: 1) to determine whether these changes in biomarkers also occur in a large, heterogeneous group of non-elite athletes; and 2) to identify possible clinical or biochemical associations. We recruited 129 non-elite runners in 2006, 61 individuals who were taking part in the half (13.1 miles) marathon and 68 individuals participating in the full (26.2 miles) marathon. Demographic data and blood samples were collected for analysis of CK, Myo, cTnT, and Creatinine (Cr) levels within two hours of race start, at race completion, and 1-h post-race for both patient cohorts. In the 61 individuals (40 males, 40+/-12 yrs) completing the half marathon in a mean time of 150+/-20 min, 90.3%, 65.2%, and 30.6% of the subjects exhibited significant elevations in Myo, CK, and cTnT, respectively immediately post race and 100%, 74.9% and 45.9% in the same biomarkers one hour-post race. In the 68 individuals (44 males, 42+/-14 yrs) completing the full marathon in a mean time of 310+/-30 min, 95.3%, 70.2% and 35.7% exhibited significant elevations in Myo, CK and cTnT respectively immediately post race and 100%, 78.5% and 52.8% in the same biomarkers one hour-post race. The elevation in cTnT levels post-race were modestly associated with the time required to complete the race for the entire cohort of marathon runners. The serum levels of Cr, CK, and Myo post-race did not correlate however with age, sex, BMI, level of training, or prior marathon experience. Elevations of cardiac injury markers in non-elite athletes are extremely common following the completion of endurance events and correlate to the increased endurance time. Whether the increase in the levels of these enzymes represents true myocardial injury or a result of the release of cTnT from the myocytes requires further investigation.

Osmolality revisited--deriving and validating the best formula for calculated osmolality.

OBJECTIVE: To derive a formula that can be used (i) to calculate osmolality in normal patients as well as those that are hyperglycemic and intoxicated, and (ii) to predict the presence of unexplained compounds with the osmol gap calculation in the presence and absence of ethanol. DESIGN AND EXPERIMENTS: We performed in vitro experiments to determine the relationship of serum osmolality with sodium, potassium, urea, glucose, ethanol, methanol, and ethylene glycol. Several formulas were then tested for their validity in predicting osmolality in normal individuals. Finally, we assessed whether these formulas would allow us to calculate the osmolality gap (OG) that may be indicative of the presence of other osmotically active compounds. The OG calculation was done both in the presence and absence of ethanol. In this way, the OG should be able to detect compounds like methanol and ethylene glycol even in the presence of ethanol which is easily measured and is very often present in the above-named poisonings. RESULTS: Experimental results show that glucose, ethanol, methanol, and ethylene glycol need factors of 1.15, 1.20, 1.07, and 1.00, respectively, to accurately predict osmolality. The factors for glucose and ethanol were then validated in normal subjects as well as in a large patient database. The formulas below predicted osmolality very well in patients whether ethanol was present or not. All concentrations are expressed in mmol/L. The mean osmol gap for healthy subjects without ethanol present was 0.77 +/- 3.80 mosM/kg with the reference interval being -6.68 to 8.23 mosM/kg for formula 1 and -8.04 to 6.50 mosM/kg for formula 2. The mean osmol gap (OG) in patients who had ethanol present was 1.22 +/- 5.32 for formula 1 and -0.2 +/- 5.0 for formula 2. CONCLUSIONS: This study shows that factors of 1.20 and 1.15 have to be applied to ethanol and glucose to allow for accurate calculation of osmolality and osmolality gap. There were insufficient patient data to verify the factors for methanol and ethylene glycol.

Molecular dynamics simulations of the d(CCAACGTTGG)(2) decamer in crystal environment: comparison of atomic point-charge, extra-point, and polarizable force fields.

Molecular dynamics simulations of the DNA duplex d(CCAACGTTGG)(2) were used to study the relationship between DNA sequence and structure in a crystal environment. Three different force fields were used: a traditional description based on atomic point charges, a polarizable force field, and an "extra-point" force field (with additional charges on extranuclear sites). It is found that all the force fields reproduce fairly well the sequence-dependent features of the experimental structure. The polarizable force field, however, provides the most accurate representation of the crystal structure and the sequence-dependent effects observed in the experiment. These results point out to the need of the inclusion of polarization for accurate descriptions of DNA.

Tryptophan fluorescence monitors multiple conformational changes required for glutamine phosphoribosylpyrophosphate amidotransferase interdomain signaling and catalysis.

Single tryptophan residues were incorporated into each of three peptide segments that play key roles in the structural transition of ligand-free, inactive glutamine phosphoribosylpyrophosphate (PRPP) amidotransferase to the active enzyme-substrate complex. Intrinsic tryptophan fluorescence and fluorescence quenching were used to monitor changes in a phosphoribosyltransferase (PRTase) "flexible loop", a "glutamine loop", and a C-terminal helix. Steady state fluorescence changes resulting from substrate binding were used to calculate binding constants and to detect the structural rearrangements that coordinate reactions at active sites for glutamine hydrolysis and PRTase catalysis. Pre-steady state kinetics of enzyme.PRPP and enzyme.PRPP.glutamine complex formation were determined from stopped-flow fluorescence measurements. The kinetics of the formation of the enzyme.PRPP complex were consistent with a model with two or more steps in which rapid equilibrium binding of PRPP is followed by a slow enzyme isomerization. This isomerization is ascribed to the closing of the PRTase flexible loop and is likely the rate-limiting step in the reaction of PRPP with NH(3). The pre-steady state kinetics for binding glutamine to the binary enzyme. PRPP complex could also be fit to a model involving rapid equilibrium binding of glutamine followed by an enzyme isomerization step. The changes monitored by fluorescence account for the interconversions between "end state" structures determined previously by X-ray crystallography and define an intermediate enzyme.PRPP conformer.

Intravenous pamidronate attenuates bone density loss after acute spinal cord injury.

OBJECTIVE: To compare the effects of a 6-month treatment with intravenous pamidronate (30-mg infusion once per month) to conventional rehabilitation without pamidronate on bone density of the spine and leg bones and on the excretion rate of N-telopeptide, a urinary marker of bone catabolism, in acutely spinal cord injured patients. DESIGN: A nonrandomized control trial in which 24 spinal cord injured subjects entered the study within 6 weeks of their injury. Fourteen subjects received pamidronate; 10 did not. OUTCOME MEASURES: Bone density measurements by dual x-ray absorptiometry were performed before the initial treatment (within 6 weeks of the injury) and at 3, 6, and 12 months postinjury and was the primary efficacy parameter. Urine for N-telopeptide levels was the secondary efficacy parameter. RESULTS: After acute spinal cord injury, patients treated with intravenous pamidronate had significantly less bone density loss compared with those who did not receive pamidronate (parametric ANOVA, p<.02). Also, ambulatory subjects had significantly less bone density loss over the study period (p<.05) than nonambulatory subjects. In general, a high excretion level of the urinary bone-breakdown product N-telopeptide was found before intravenous pamidronate treatment, followed by a dramatic reduction in excretion after pamidronate treatment. Ambulatory subjects excreted significantly less N-telopeptide than motor-complete subjects at all time points. CONCLUSION: Intravenous pamidronate treatment and ambulatory ability in the first 6 months after an acute spinal cord injury prevents bone density loss.

Crystal structure of glutamine phosphoribosylpyrophosphate amidotransferase from Escherichia coli.

Crystal structures of glutamine phosphoribosylpyrophosphate (PRPP) amidotransferase from Escherichia coli have been determined to 2.0-A resolution in the absence of ligands, and to 2.5-A resolution with the feedback inhibitor AMP bound to the PRPP catalytic site. Glutamine PRPP amidotransferase (GPATase) employs separate catalytic domains to abstract nitrogen from the amide of glutamine and to transfer nitrogen to the acceptor substrate PRPP. The unliganded and AMP-bound structures, which are essentially identical, are interpreted as the inhibited form of the enzyme because the two active sites are disconnected and the PRPP active site is solvent exposed. The structures were compared with a previously reported 3.0-A structure of the homologous Bacillus subtilis enzyme (Smith JL et al., 1994, Science 264:1427-1433). The comparison indicates a pattern of conservation of peptide structures involved with catalysis and variability in enzyme regulatory functions. Control of glutaminase activity, communication between the active sites, and regulation by feedback inhibitors are addressed differently by E. coli and B. subtilis GPATases. The E. coli enzyme is a prototype for the metal-free GPATases, whereas the B. subtilis enzyme represents the metal-containing enzymes. The structure of the E. coli enzyme suggests that a common ancestor of the two enzyme subfamilies may have included an Fe-S cluster.

Coupled formation of an amidotransferase interdomain ammonia channel and a phosphoribosyltransferase active site.

Activation of gluatmine phosphoribosylpyrophosphate (RPPP) amidotransferase (GPATase) by binding of a PRPP substrate analog results in the formation of a 20 A channel connecting the active site for glutamine hydrolysis in one domain with the PRPP site in a second domain. This solvent-inaccessible channel permits transfer of the NH3 intermediate between the two active sites. Tunneling of NH3 may be a common mechanism for glutamine amidotransferase-catalyzed nitrogen transfer and for coordination of catalysis at two distinct active sites in complex enzymes. The 2.4 A crystal structure of the active conformer of GPATase also provides the first description of an intact active site for the phosphoribosyltransferase (PRTase) family of nucleotide synthesis and salvage enzymes. Chemical assistance to catalysis is provided primarily by the substrate and secondarily by the enzyme in the proposed structure-based mechanism. Different catalytic and inhibitory modes of divalent cation binding to the PRTase active site are revealed in the active conformer of the enzyme and in a feedback-inhibited GMP complex.

Transcutaneous bilirubinometry: its role in the assessment of neonatal jaundice.

OBJECTIVE: To review the literature on transcutaneous bilirubinometry so that its exact role in the prevention of kernicterus or bilirubin encephalopathy could be determined. DESIGN AND METHODS: Literature searches were done in Medline and Current Contents. RESULTS: It is estimated that about 50% of newborns have an episode of jaundice in the first few days of life. Six percent of newborns may develop hyperbilirubinemia (> 220 mumol/L), which can potentially cause bilirubin encephalopathy or kernicterus, a severe neonatal disease. In the past, serum bilirubin (SB) has been the preferred method of detecting hyperbilirubinemia in newborns. The ordering of SB in neonates is based on visual evaluation by either physicians or nursing staff. Skin puncture collection of blood exposes the neonate to trauma and risk of infection. A noninvasive device for predicting serum bilirubin levels in newborns diminishes the need to do skin punctures. One such device that has been very extensively studied is the Minolta AirShields Jaundice Meter. It is a portable light-weight instrument that uses reflectance measurements on the skin to determine the amount of yellow color present in the skin, namely transcutaneous bilirubin (TcB). Although the TcB measurements correlate well with serum bilirubin (SB) levels, they cannot accurately predict serum bilirubin because of error related to a variety of factors. CONCLUSIONS: TcB cannot be used directly to make decisions about transfusions or phototherapy in neonates. It is a good tool for screening neonates to determine when a laboratory measurement of serum bilirubin is needed. Such a practice requires careful selection of the decision level so that false-negative TcB values do not prevent appropriate serum bilirubin tests from being done.

Clinical impact of transcutaneous bilirubinometry as an adjunctive screen for hyperbilirubinemia.

OBJECTIVE: To determine what the clinical impact would be of implementing a jaundice meter for use in a busy neonatal service as an adjunctive screening tool for hyperbilirubinemia. DESIGN AND METHODS: Test utilization data was collected for a 6-month period to determine how neonatal bilirubin was utilized in this hospital. The jaundice meter was evaluated in a study population of healthy term infants. The performance characteristics of the meter and the test utilization data were used to predict the clinical impact a meter could have on screening for hyperbilirubinemia. RESULTS: Utilization data indicated that about 60% of all single bilirubin neonatal testing (i.e., bilirubin only ordered) was done by normal nurseries. A jaundice meter cutoff decision reading of 17 was shown to have a sensitivity of 100% and a specificity of 68% for hyperbilirubinemia (> 260 mumol/L) in a study population of healthy term infants. From this data, it was estimated that use of a jaundice meter could eliminate 43% of the single (i.e., not combined with other tests) bilirubin tests done on healthy term neonates with no prior exposure to phototherapy. This constitutes an overall 20% reduction in bilirubin testing in normal nurseries when testing done on babies exposed to phototherapy and combined bilirubin testing are taken into consideration. Additionally, it was shown that there would be an improvement of 9% in the prediction of hyperbilirubinemia without loss of 100% sensitivity. CONCLUSION: Use of a jaundice meter in normal nurseries as an adjunctive screening tool enhances patient care by reducing the overall blood procurement rate in normal nurseries by 20% and increasing screening efficiency for significant hyperbilirubinemia by 5%.

Structure and function of the glutamine phosphoribosylpyrophosphate amidotransferase glutamine site and communication with the phosphoribosylpyrophosphate site.

Glutamine phosphoribosylpyrophosphate (PRPP) amidotransferase from Escherichia coli exhibits a basal PRPP-independent glutaminase activity having a kcat/Km that is 0.3% of fully active enzyme. Binding of PRPP activates the enzyme by a structural change that lowers the Km for glutamine 100-fold and couples glutamine hydrolysis to synthesis of 5-phosphoribosylamine. By analysis of the x-ray structure of the glutamine site containing bound 6-diazo-5-oxonorleucine, a glutamine affinity analog, and by site-directed mutagenesis we have identified residues important for glutamine binding, catalysis, and coupling with PRPP. Tyr74 is a key residue in the coupling between the sites for glutamine in the NH2-terminal domain and PRPP in the COOH-terminal domain. Arg73 and Asp127 have roles in glutamine binding. The x-ray structure indicates that there are no amino acid side chains sufficiently close to Cys1 to participate as a proton acceptor in formation of the thiolate needed for nucleophilic attack on the carboxamide of glutamine, nor as a general acid for amide nitrogen transfer. Based on the x-ray model of the glutamine site and analysis of a mutant enzyme we propose that the free NH2 terminus of Cys1 functions as the proton acceptor and donor. The results indicate that the side chain of Asn101 and the backbone nitrogen of Gly102 function to stabilize a tetrahedral oxyanion resulting from attack of Cys1 on the glutamine carboxamide. Cys1, Arg73, Asn101, Gly102, and Asp127 are conserved in the NH2-terminal domain of a subfamily of amidotransferases that includes asparagine synthetase, glucosamine 6-phosphate synthase, and glutamate synthase, implying a common function in the four enzymes. Tyr74, on the other hand, is conserved only in glutamine PRPP amidotransferase sequences consistent with a specific role in interdomain coupling. The catalytic framework of key glutamine site residues supports the assignment of glutamine PRPP amidotransferase to a recently described Ntn (NH2-terminal nucleophile) hydrolase family of enzymes.

Falsely elevated enzyme-multiplied immunoassay serum valproic acid results in 12 patients.

A split sample study of 69 patients' serum for valproic acid analysis by enzyme-multiplied immunoassay technique (EMIT) and by gas-liquid chromatography revealed falsely elevated EMIT values in 12 patients. Of the 69 serum samples, 28 were also assayed by fluorescence polarization immunoassay and all results agreed with those of gas-liquid chromatography. Five of the 12 patients with elevated EMIT values were included in this subgroup. The unidentified interfering substance appears to be a heat-labile, high-molecular-weight compound, possibly a protein complex that can persist in blood for several months following discontinuation of the drug.

Detection of Brucella melitensis and Brucella abortus by DNA amplification.

Suitable reaction conditions and oligonucleotide primers were sought for the detection of Brucella melitensis and Brucella abortus by the polymerase chain reaction. Primers were chosen from within the coding sequence of a gene encoding a 31 kDa B. abortus antigen. The test was shown to be sensitive, and specificity was demonstrated using DNA derived from a panel of Gram-negative pathogens. There was no detectable difference between B. melitensis and B. abortus in the sensitivity of the reaction or in the size of the amplification product. The technique should be applicable in the diagnosis of brucellosis.

Excess serum osmolality gap after ingestion of methanol.

Two cases of methanol ingestion and one case of combined methanol and ethylene glycol ingestion are presented to illustrate the large differences that exist between the serum osmolality gap and the measured methanol (plus ethylene glycol) concentration(s) before treatment of these poisonings. After treatment with intravenous ethanol and hemodialysis was initiated, the differences disappeared in all three cases. We speculate that one or more metabolites with osmotic activity are formed in cases of methanol intoxication where no ethanol has also been consumed. The possible identity of these compounds is discussed.