Typing of Campylobacter jejuni isolates from dogs by use of multilocus sequence typing and pulsed-field gel electrophoresis.

Campylobacter is a major cause of human gastroenteritis worldwide. Risk of Campylobacter infection in humans has been associated with many sources, including dogs. This study aimed to investigate whether C. jejuni carried by dogs could potentially be a zoonotic risk for humans and if there were common sources of C. jejuni infection for both humans and dogs. Multilocus sequence typing (MLST) together with macrorestriction analysis of genomic DNA using SmaI and pulsed-field gel electrophoresis (PFGE) were both used to analyze 33 C. jejuni isolates obtained from various dog populations, including those visiting veterinary practices and from different types of kennels. MLST data suggested that there was a large amount of genetic diversity between dog isolates and that the majority of sequence types found in isolates from these dogs were the same as those found in isolates from humans. The main exception was ST-2772, which was isolated from four samples and could not be assigned to a clonal complex. The most commonly identified clonal complex was ST-45 (11 isolates), followed by ST-21 (4 isolates), ST-508 (4 isolates), and ST-403 (3 isolates). The profiles obtained by macrorestriction PFGE were largely in concordance with the MLST results, with a similar amount of genetic diversity found. The diversity of sequence types found within dogs suggests they are exposed to various sources of C. jejuni infection. The similarity of these sequence types to C. jejuni isolates from humans suggests there may be common sources of infection for both dogs and humans. Although only a small number of household dogs may carry C. jejuni, infected dogs should still be considered a potential zoonotic risk to humans, particularly if the dogs originate from kennelled or hunt kennel dog populations, where the prevalence may be higher.

Risk factors for the carriage of Campylobacter upsaliensis by dogs in a community in Cheshire.

Samples of faeces were taken from 183 healthy pet dogs in a census-based, cross-sectional study in Cheshire; culture methods were used to detect any Campylobacter species and a direct PCR was used to detect Campylobacter upsaliensis. Forty-six of the dogs were positive for C upsaliensis by either culture or direct PCR, giving a prevalence of 25.1 per cent (95 per cent confidence interval [CI] 19.0 to 32.1 per cent). One sample was positive by culture for Campylobacter jejuni (95 per cent CI 0.0 to 3.0 per cent) and one for Campylobacter lari. Multivariable logistic regression identified risk factors for the carriage of C upsaliensis by a dog as: living with another dog that also carried C upsaliensis; being small rather than medium-sized; being less than three years old; living in a household that kept fish; being fed commercial dog treats; and being fed human food titbits, particularly in the dog's bowl.

An asymmetric structure of the Bacillus subtilis replication terminator protein in complex with DNA.

In Bacillus subtilis, the termination of DNA replication via polar fork arrest is effected by a specific protein:DNA complex formed between the replication terminator protein (RTP) and DNA terminator sites. We report the crystal structure of a replication terminator protein homologue (RTP.C110S) of B. subtilis in complex with the high affinity component of one of its cognate DNA termination sites, known as the TerI B-site, refined at 2.5 A resolution. The 21 bp RTP:DNA complex displays marked structural asymmetry in both the homodimeric protein and the DNA. This is in contrast to the previously reported complex formed with a symmetrical TerI B-site homologue. The induced asymmetry is consistent with the complex's solution properties as determined using NMR spectroscopy. Concomitant with this asymmetry is variation in the protein:DNA binding pattern for each of the subunits of the RTP homodimer. It is proposed that the asymmetric "wing" positions, as well as other asymmetrical features of the RTP:DNA complex, are critical for the cooperative binding that underlies the mechanism of polar fork arrest at the complete terminator site.

Structure and RNA binding of the third KH domain of poly(C)-binding protein 1.

Poly(C)-binding proteins (CPs) are important regulators of mRNA stability and translational regulation. They recognize C-rich RNA through their triple KH (hn RNP K homology) domain structures and are thought to carry out their function though direct protection of mRNA sites as well as through interactions with other RNA-binding proteins. We report the crystallographically derived structure of the third domain of alphaCP1 to 2.1 A resolution. alphaCP1-KH3 assumes a classical type I KH domain fold with a triple-stranded beta-sheet held against a three-helix cluster in a betaalphaalphabetabetaalpha configuration. Its binding affinity to an RNA sequence from the 3'-untranslated region (3'-UTR) of androgen receptor mRNA was determined using surface plasmon resonance, giving a K(d) of 4.37 microM, which is indicative of intermediate binding. A model of alphaCP1-KH3 with poly(C)-RNA was generated by homology to a recently reported RNA-bound KH domain structure and suggests the molecular basis for oligonucleotide binding and poly(C)-RNA specificity.

Lethal outbreak of disease associated with feline calicivirus infection in cats.

Recently, in the USA, virulent mutants of feline calicivirus (FCV) have been identified as the cause of a severe and acute virulent systemic disease, characterised by jaundice, oedema and high mortality in groups of cats. This severe manifestation of FCV disease has so far only been reported in the USA. However, in 2003, an outbreak of disease affected a household of four adult cats and an adult cat from a neighbouring household in the UK. Three of the adult cats in the household and the neighbouring cat developed clinical signs including pyrexia (39.5 to 40.5 degrees C), lameness, voice loss, inappetence and jaundice. One cat was euthanased in extremis, two died and one recovered. A postmortem examination of one of the cats revealed focal cellulitis around the right hock and right elbow joints. The principal finding of histopathological examinations of selected organs from two of the cats was disseminated hepatocellular necrosis with mild inflammatory infiltration. Immunohistology identified FCV antigen in parenchymal and Kupffer cells in the liver of both animals and in alveolar macrophages of one of them. In addition, calicivirus-like particles were observed by electron microscopy within the hepatocytes of one cat. FCV was isolated from two of the dead cats and from the two surviving cats. Sequence analysis showed that they were all infected with the same strain of virus, but that it was different from strains of FCV associated with the virulent systemic disease in cats in the USA. The outbreak was successfully controlled by quarantine in the owner's house.

Syncope in children and adolescents.

OBJECTIVES: The objectives of this study were to 1) define the incidence of syncope coming to medical attention among children and adolescents, 2) determine the outcome of syncope in these patients, and 3) determine changes over time in the evaluation and charges for evaluating this problem. BACKGROUND: Syncope occurs commonly in children and adolescents. However, the mid- and long-term outcome of children and adolescents who experience syncope is unknown. METHODS: Utilizing the Rochester Epidemiology Project, we determined the incidence, outcome and charges for medical evaluation for patients seeking medical attention for syncope during an early 5-year period (1950 to 1954) and a more recent 5-year period (1987 to 1991). RESULTS: The incidence of syncope coming to medical attention was 71.9 and 125.8/100,000 population for the early and more recent cohort, respectively. The incidence was higher for female than for male patients. The incidence peaked in 15- to 19-year old patients. Acute illness and noxious stimuli were associated with 24% and 23% of the episodes, respectively. Although long-term survival was not different from that of the general population, one child died suddenly, and another had hereditary prolonged QT interval syndrome. These were two of only six patients who had exertional syncope. Total charges for evaluation of syncope were similar in the two time periods. However, charges for testing procedures were greater for the more recent cohort. CONCLUSIONS: In general, syncope in children and adolescents is a benign event. Syncope occurring during exercise may identify patients with a potentially fatal condition. Detailed evaluation should be considered for patients who have syncope during exercise or who have a family history of syncope, sudden death, myocardial disease or arrhythmias. It may be prudent to obtain an electrocardiogram for all patients who seek medical attention for syncope.

Pacemaker treatment of sick sinus syndrome in children.

The sick sinus syndrome is being recognized with increasing frequency in children. Although it is sometimes benign, it can be serious or have fatal consequences. Fifty-one patients (mean age 10.5 years) underwent permanent cardiac pacing for sick sinus syndrome. Twenty patients had epicardial ventricular pacing and 12 had an epicardial atrial implant. Seven had endocardial atrial pacing, six epicardial atrioventricular (AV) sequential pacing, four epicardial universal pacing and two endocardial universal pacing. Of the 49 symptomatic patients, 45 had relief of symptoms. Eleven of 18 patients with associated tachyarrhythmias had amelioration of their tachycardia. There were no early but two late deaths unrelated to the pacemakers. Seven patients during a mean follow-up period of 26 months required reoperation for pacing lead or sensing problems. Permanent pacing for sick sinus syndrome in children is a safe and symptomatically effective procedure.

Amiodarone treatment of critical arrhythmias in children and young adults.

The majority of sudden cardiac deaths in children occur in patients with prior arrhythmias and an abnormal heart. Amiodarone was given to 39 young patients (35 with an abnormal heart) with arrhythmias unresponsive to conventional treatment. Their age ranged from 6 weeks to 30 years with nine patients younger than 2 years of age. Atrial flutter was present in 16 patients, ventricular tachycardia in 14 patients and supraventricular tachycardia in 9 patients. The most common diagnosis (14 patients) was postoperative repair of congenital heart disease. The dose ranged from 2.5 to 21.6 mg/kg per day (mean 8.2). Elimination of arrhythmia (on 24 hour electrocardiography) occurred in 15 of 16 patients with atrial flutter, 11 of 14 with ventricular tachycardia and 5 of 9 with supraventricular tachycardia. Symptomatic side effects were: rash (three patients), headache (two patients), nausea (one patient) and peripheral neuropathy (one patient); seven patients had asymptomatic corneal microdeposits which normalized in all after the drug was discontinued. No side effects occurred in patients younger than 10 years of age. The following changed with treatment (p less than 0.05): heart rate decreased (three patients with atrial flutter and sick sinus syndrome required pacemaker implantation for bradycardia) and QTc increased; thyroxine (T4) and serum reverse triiodothyronine (T3) increased. During follow-up study (range 6 months to 3 years), 21 of the 39 patients continued to take amiodarone with complete control of arrhythmias, 9 were no longer taking the drug and 9 died (7 nonsudden and 2 sudden deaths). Amiodarone is an extremely effective treatment for infants and children with tachyarrhythmias resistant to conventional treatment.(ABSTRACT TRUNCATED AT 250 WORDS)

Atrial flutter in the young: a collaborative study of 380 cases.

As children with cardiac disease grow older, atrial flutter becomes more prevalent. A collaborative study was performed in 19 institutions to determine the clinical characteristics of these children and the factors affecting prognosis. There were 380 patients with one or more electrocardiographically documented episodes of atrial flutter that first occurred between ages 1 and 25 years (mean age at onset 10.3). Episodes of flutter continued to occur for a mean of 2.5 years after the onset. Of the 380 patients, 60% had repaired congenital heart disease, 13% palliated congenital heart disease, 8% unoperated congenital heart disease, 8% an otherwise normal heart, 6% cardiomyopathy, 4% rheumatic heart disease and 2% other lesions. Overall, drugs were effective in eliminating atrial flutter in 58% of patients; specifically, amiodarone and digoxin plus quinidine were effective in 53%, digoxin alone in 44% and propranolol in 21%. Amiodarone was effective in seven (78%) of nine patients. Corrective surgery was performed after the onset of atrial flutter in 66 patients; in 52% the atrial flutter was easier to control or it resolved and in only 4% it was worse. At follow-up (mean 6.5 years), 83% of the patients were alive (49% without atrial flutter and 34% with atrial flutter) and 17% died (10% suddenly, 6% of nonsudden cardiac cause and 1% of noncardiac cause). Cardiac death occurred in 20% of those for whom an effective drug could not be found to eliminate atrial flutter compared with 5% of those who were treated with an effective drug (p less than 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)

Improved early morbidity and mortality after Fontan operation: the Mayo Clinic experience, 1987 to 1992.

OBJECTIVES: This study sought to evaluate changes in early morbidity and mortality as well as predictors of outcome in our most recent 339 patients undergoing modified Fontan operations. BACKGROUND: The Fontan operation is the preferred definitive palliation for patients with functional single ventricles. Previously reported early mortality rates after Fontan operation have been substantial. METHODS: Records of 339 consecutive patients who had a Fontan operation at the Mayo Clinic between 1987 and 1992 (recent cohort) were reviewed. This cohort was compared with the previous 500 patients who had Fontan operations performed between 1973 and 1986 (early cohort). RESULTS: Recently, overall early mortality after Fontan has decreased significantly compared with that for the early cohort (from 16% to 9%, p = 0.002). This decline occurred despite increased anatomic complexity of patients. Short-term posthospital survival has also improved significantly in recent patients. One-year survival improved to 88% from 79%, and 5-year survival to 81% from 73% (p = 0.006). Patients with common atrioventricular valves and those who took daily preoperative diuretic medication or had either postoperative renal failure or elevated postbypass right atrial pressure were at increased risk for early mortality. Young age was not found to be a risk factor for early mortality. Early mortality for patients with heterotaxia decreased dramatically: recent 30-day mortality was 15% compared with 41% in the early heterotaxy cohort. CONCLUSIONS: Many factors may have contributed to decreased early mortality after Fontan. Improved patient selection, younger age at time of operation, refinements in surgical techniques and postoperative management may all have had important roles. Proposed technical modifications of the Fontan operation must be evaluated in light of these improved results.

In vitro assessment of oral lipid based formulations.

In recent years there has been an increase in interest in the utility of lipid based delivery systems, at least in part as a result of the effective development of lipid based products such as Sandimmun Neoral (cyclosporin), Norvir (ritonavir) and Fortovase (saquinavir). The development pathway for lipid based formulations, however, is still largely empirical, and in vitro models that are predictive of oral bioavailability enhancement are lacking. The use of modified dissolution media, reflecting the bile salt and phospholipid levels in the intestine, has met with some success in terms of the ability to predict the bioavailability of poorly water soluble drugs and the potential bioavailability enhancing effects of food. These approaches, however, do not have the flexibility or complexity to deal with the interactions inherent in the digestion, dispersion and solubilisation of a lipid based formulation and the coincident dissolution profile of a co-administered drug. In this review, the utility of modified dissolution media to predict the impact of food on the absorption of poorly water soluble, lipophilic drugs, is explored. These dissolution based systems are subsequently contrasted with the use of lipid digestion models which have found increasing application in assessment of the interaction of digestible dose forms with the gastrointestinal milieu.

Intestinal lymphatic drug transport: an update.

The trend towards identification of poorly water-soluble and highly lipophilic candidate drug molecules has led to an increase in interest in intestinal lymphatic drug transport. In this article we provide a brief background to the mechanism of access of drugs to the intestinal lymph and the role of lipid digestion and absorption in the stimulation of lymphatic transport. The ability of different lipid types to stimulate lymphatic drug transport, is addressed, concentrating specifically on the impact of the class, chain length and degree of unsaturation of co-administered lipids. Comment is also made as to the relevance of dosing different lipid volumes to the rat and the possible complications this may provide when trying to assess the likely extent of intestinal lymphatic transport. Recent studies are described in which the extent of lymphatic transport of a highly lipophilic antimalarial, halofantrine, was investigated after post-prandial administration to greyhound dogs. Finally the possible future directions for studies of intestinal lymphatic transport are discussed, including the use of cell culture models and genetically modified animals.

Lymphatic transport of proteins after s.c. injection: implications of animal model selection.

Subcutaneous (s.c.) administration continues to be the main route for the delivery of protein drugs due to their poor bioavailability by most non-parenteral routes. While small drug molecules are rapidly and extensively absorbed after s.c. injection, the systemic bioavailability of protein drugs is often incomplete and variable. Given the widespread use of the s.c. route for protein drugs, surprisingly little is known about the factors that govern the rate and extent of protein absorption from the interstitial space and the role of the lymphatic system in the transport of these molecules to the systemic circulation. The few studies that have directly addressed the role of lymphatic transport in protein bioavailability are complicated by the use of methods and models that vary widely. In this review we will evaluate the available literature describing the lymphatic transport of proteins after s.c. injection and more specifically, address the impact of experimental variation (e.g. site of cannulation, animal model, anesthesia) on the interpretation of the data obtained. We will also describe in some detail the sheep model currently in use in our laboratory, which allows both estimation of the extent of uptake of protein drugs into the lymphatics draining the injection site, and quantification of the contribution of lymphatic transport to the absolute bioavailability.

Animal models for the study of intestinal lymphatic drug transport.

Drug transport via the intestinal lymphatic system has been shown to contribute to the absorption of a number of orally administered highly lipophilic drugs. In order to investigate this phenomenon and assist in the development of improved oral formulations, the use of appropriate animal models is required. This paper reviews the use of various animal models for this purpose, and describes in detail the conscious rat and dog models used in our laboratory. The advantages and disadvantages of both small and large animal models are explored, as well as the factors which have been found to influence the outcome of intestinal lymphatic drug transport studies with these models.

The polyoxyethylene/polyoxypropylene block co-polymer poloxamer-407 selectively redirects intravenously injected microspheres to sinusoidal endothelial cells of rabbit bone marrow.

Small colloidal particulates (150 nm and below, in diameter) can be redirected specifically to the rabbit bone marrow following intravenous administration by coating their surface with the block co-polymer poloxamer-407, a non-ionic surfactant. The coated colloids are sequestered by the sinusoidal endothelial cells of the bone marrow and are accumulated in dense bodies within these cells. The uptake of poloxamer-407-coated colloids by marrow endothelial cells suggests that the steric repulsive barrier, imposed by the polyoxyethylene segment of the polymer, to particle-cell interaction can apparently be overcome by a specific interaction mechanism(s) with the cell surface. Such a dramatic uptake cannot be achieved with other block co-polymers of similar structure to poloxamer-407. The application of the current model for the site-specific targeting of drug carriers to bone marrow and the prevention of the adherence of metastases of tumours which selectively colonize the bone marrow endothelium is discussed.

fas-fas-ligand antigen expression and its relationship to increased apoptosis in acute renal transplant rejection.

BACKGROUND: To examine the role of fas-fas-ligand interaction and apoptosis in acute transplant rejection. METHODS: Pre- and posttransplant renal allograft biopsies were stained by in situ 3-end labeling of DNA for detection of apoptotic cells (TUNEL) and immunohistochemistry techniques were used for demonstration of fas and fas-ligand antigen expression. RESULTS: Posttransplantation apoptosis was significantly increased in acute rejection and acute tubular necrosis (P<0.0001) compared to preimplantation biopsies and biopsies taken from grafts showing dysfunction not attributed to rejection. Fas and fas-ligand expression was demonstrated predominantly in the tubular epithelium. In preimplant biopsies fas was expressed in 11% (4/37) of cases; posttransplantation expression increased to: 44% (8/18) acute rejection, 63% (5/8) acute tubular necrosis, and 38% (5/13) dysfunction without evidence of rejection. Fas-ligand was expressed by 30% (11/37) of preimplant biopsies, posttransplantation expression was reduced in all groups: 17% (3/18) acute rejection, 13% (1/8) acute tubular necrosis, delayed xenograft rejection and 15% (2/13) dysfunction without evidence of rejection. A correlation with fas-1 expression preimplantation and a subsequent absence of acute rejection post transplant was noted (P<0.001). CONCLUSIONS: Apoptosis is a feature of acute rejection and acute tubular necrosis. Fas expression is uncommon preimplantation and increases non-specifically post transplant. Fas-1 was expressed by a third of preimplantation biopsies and expression was lost non-specifically post transplant. The expression of fas-ligand preimplantation correlated with an absence of acute rejection episodes posttransplant, suggesting some degree of immune privilege. These data suggest that the fas-fas-1 mediated pathway does not play a specific role in apoptosis during acute rejection. We were unable to find any evidence that the fas-fas-1-mediated pathway has a role in the increased apoptosis seen during acute rejection.

Accelerated idioventricular rhythm: a benign arrhythmia in childhood.

OBJECTIVE: To determine whether accelerated idioventricular rhythm (AIVR) is benign in pediatric patients. METHODS: The records were reviewed of all patients younger than age 15 years who had been diagnosed with a ventricular arrhythmia between 1976 and 1991. RESULTS: AIVR was diagnosed in 12 patients, ages 1 day to 15 years (mean, 8.9 years). In 3 patients the arrhythmia was discovered on ambulatory electrocardiographic monitoring after presentation with syncope or presyncopal symptoms. One patient had palpitations. The remaining diagnoses were made during routine examinations or at postoperative follow-ups for congenital heart disease. The AIVR rate ranged from 90 to 150 beats per minute. The rate was within 10 beats per minute of the preceding sinus rate in 11 patients. Echocardiograms were normal in 8 patients. Two patients had double-outlet right ventricles; 1 had repair of the tetralogy of Fallot; and 1 had abnormal ventricular septal motion. Seven patients were taking antiarrhythmia medication without control of the AIVR at presentation. Patients were followed for a mean of 68.4 months (range, 31 to 191 months). All patients were alive and asymptomatic, with normal activity at the last follow-ups. CONCLUSIONS: Complete resolution of AIVR may not occur. However, AIVR seems to be a benign arrhythmia. Treatment was not effective in controlling the arrhythmia and is likely unnecessary.

Verapamil: an effective calcium blocking agent for pediatric patients.

Verapamil is a slow-channel calcium-blocking agent that has been released recently by the Food and Drug Administration for treatment of cardiac dysrhythmias in all age groups. Its primary action is to slow conduction in the atrioventricular node, thereby abolishing those types of supraventricular tachycardia using the atrioventricular node as a part of the reentry circuit or slowing the ventricular rate in someone with atrial flutter. Other investigations have shown that it is likely to relieve left ventricular outflow obstruction in patients with hypertrophic obstructive cardiomyopathy. Because of its potential widespread usefulness in the pediatric population, all pediatricians should be more aware of how it is used and the potential hazards. The methods of administration and treatment of overdoses as well as indications for usage, contraindications, and adverse reactions will be explained.

Tilt-table testing and oral metoprolol therapy in young patients with unexplained syncope.

OBJECTIVE: Tilt-table testing is useful for investigating unexplained syncope in pediatric patients. No data are available on the use of intravenous metoprolol during testing to identify children who might respond to subsequent oral beta-adrenergic blockade or on the efficacy and safety of such oral therapy. DESIGN: To provide these data, we obtained follow-up information on 27 consecutive pediatric patients who had tilt-table testing for unexplained syncope. RESULTS: Nineteen patients (70%) had positive test results with or without isoproterenol infusion. All these patients had negative test results after intravenous infusion of metoprolol and subsequently were treated with oral metoprolol. Taking oral metoprolol therapy alone, 9 (47%) of the 19 patients were asymptomatic, 8 (42%) reported a decreased frequency of syncopal episodes, and 2 (11%) had unchanged or more frequent episodes. The mean dosage of metoprolol required to prevent symptoms was 1.5 mg/kg per day. Mild side effects were reported by 6 (32%) of the 19 patients. Six patients (32%) required additional medications. CONCLUSIONS: We conclude that metoprolol is safe and effective for the treatment of most cases of neurocardiogenic syncope in children and that this response cannot be predicted accurately by the use of intravenous metoprolol during testing.

Exercise testing in children with Wolff-Parkinson-White syndrome.

Exercise testing using a modified Bruce treadmill protocol was performed by 17 children with Wolff-Parkinson-White (WPW) syndrome. All had intracardiac electrophysiology studies as well. Endurance time, heart rate and blood pressure were normal during exercise. Ventricular premature complexes were seen with exercise in 2 patients and supraventricular tachycardia with exercise testing was seen in 2. Disappearance of the delta wave with exercise correlated with a long anterograde effective refractory period of the Kent bundle (360 to 390 ms). Children with partial normalization of the QRS during exercise had a longer anterograde effective refractory period of the Kent bundle than those in whom preexcitation persisted. In 1 patient, disappearance of the delta wave with exercise confirmed the diagnosis of WPW syndrome. Preexcitation was seen only after exercise in 1 patient. Exercise testing is of value in the evaluation of children with WPW syndrome; children with WPW syndrome who have total normalization of the QRS interval during exercise and few or no symptoms of tachycardia do not require electrophysiologic study.