Detection of Rickettsia spp. and Bartonella spp. in Ctenocephalides felis fleas from free-ranging crab-eating foxes (Cerdocyon thous).

Fleas are insects with a worldwide distribution that have been implicated in the transmission of several pathogens. The present study aimed to investigate the presence of Rickettsia spp. (Rickettsiales: Rickettsiaceae) and Bartonella spp. (Rhizobiales: Bartonellaceae) in fleas from free-ranging crab-eating foxes Cerdocyon thous (Linnaeus, 1766) (Carnivora: Canidae) from Rio Grande do Sul, southern Brazil. Fleas were collected manually from animals and used for the molecular detection of Rickettsia spp. and Bartonella spp. Twenty-nine C. thous were sampled in six municipalities. Four foxes were parasitized by 10 fleas, all of which were identified as Ctenocephalides felis (Bouché, 1935) (Siphonaptera: Pulicidae). DNA from Rickettsia felis Bouyer et al., 2001 and Rickettsia asembonensis Maina et al., 2016 were found in three and eight fleas, respectively. In four fleas, DNA of Bartonella sp. was identified. Phylogenetic analysis grouped Bartonella sp. together with other genotypes previously reported in C. felis worldwide. The scenario described in the present study highlights a Neotropical canid parasitized by the invasive cosmopolitan cat flea, which in turn, is carrying potentially invasive vector-borne microorganisms. These findings suggest that C. felis is adapted to wild hosts in wilderness areas in southern Brazil, hypothetically exposing the Neotropical fauna to unknown ecological and health disturbances.

Effect of growth hormone treatment on energy expenditure and its relation to first-year growth response in children.

PURPOSE: The effects of growth hormone (GH) treatment on linear growth and body composition have been studied extensively. Little is known about the GH effect on energy expenditure (EE). The aim of this study was to investigate the effects of GH treatment on EE in children, and to study whether the changes in EE can predict the height gain after 1 year. METHODS: Total EE (TEE), basal metabolic rate (BMR), and physical activity level (PAL) measurements before and after 6 weeks of GH treatment were performed in 18 prepubertal children (5 girls, 13 boys) born small for gestational age (n = 14) or with growth hormone deficiency (n = 4) who were eligible for GH treatment. TEE was measured with the doubly labelled water method, BMR was measured with an open-circuit ventilated hood system, PAL was assessed using an accelerometer for movement registration and calculated (PAL = TEE/BMR), activity related EE (AEE) was calculated [AEE = (0.9 × TEE) - BMR]. Height measurements at start and after 1 year of GH treatment were analysed. This is a 1-year longitudinal intervention study, without a control group for comparison. RESULTS: BMR and TEE increased significantly (resp. 5% and 7%). Physical activity (counts/day), PAL, and AEE did not change. 11 out of 13 patients (85%) with an increased TEE after 6 weeks of GH treatment had a good first-year growth response (∆height SDS > 0.5). CONCLUSIONS: GH treatment showed a positive effect on EE in prepubertal children after 6 weeks. No effect on physical activity was observed. The increase in TEE appeared to be valuable for the prediction of good first-year growth responders to GH treatment.

[Algorithm for the treatment of the haemophilic arthropathia of the upper ankle joint]. / Behandlungsalgorithmus der hämophilen Arthropathie des oberen Sprunggelenkes.

The upper ankle joint is one of the target-joints of the haemophilic patient. Therefore, the secondary arthritis of the upper ankle joint is one of the most frequent forms of haemophilic arthropathy. It is a secondary form of arthritis not only because of chronic synovitis and cartilage injury resulting from chronic recurrent intraarticular bleeds, but also due to the misalignment of the joint and abnormal joint stress. The consequences are manifest even in young patients and finally lead to upper ankle joint arthritis. In such clinical situations, the upper ankle joint-arthroplasty is a viable alternative to arthrodesis. After several years of bleeding of the upper ankle joint many patients with haemophilia suffer from symptomatic arthritis. Open joint cleansing considerably improves mobility in the upper ankle joint and alleviates the pain in the talonavicular joint. However, the recovered mobility of the arthritic upper ankle joint also activates arthritis, associated with severe pain. With no contraindication to upper ankle joint replacement, a cement-free prosthesis can be implanted. Three months after surgery, the patients are mobile, with good foot rolling properties without orthopaedic aids and without pain in the upper joint ankle. Concludion: In terms of biomechanics the upper ankle joint-arthroplasty is a superior alternative to arthrodesis in haemophilia patients. In order to minimize the complication rate, their treatment should be restricted to specially equipped interdisciplinary centers with adequately trained and experienced surgeons as well as haemostaseologists.

Tropomyosin-containing actin cables direct the Myo2p-dependent polarized delivery of secretory vesicles in budding yeast.

The actin cytoskeleton in budding yeast consists of cortical patches and cables, both of which polarize toward regions of cell growth. Tropomyosin localizes specifically to actin cables and not cortical patches. Upon shifting cells with conditionally defective tropomyosin to restrictive temperatures, actin cables disappear within 1 min and both the unconventional class V myosin Myo2p and the secretory vesicle-associated Rab GTPase Sec4p depolarize rapidly. Bud growth ceases and the mother cell grows isotropically. When returned to permissive temperatures, tropomyosin-containing cables reform within 1 min in polarized arrays. Cable reassembly permits rapid enrichment of Myo2p at the focus of nascent cables as well as the Myo2p- dependent recruitment of Sec4p and the exocyst protein Sec8p, and the initiation of bud emergence. With the loss of actin cables, cortical patches slowly assume an isotropic distribution within the cell and will repolarize only after restoration of cables. Therefore, actin cables respond to polarity cues independently of the overall distribution of cortical patches and are able to directly target the Myo2p-dependent delivery of secretory vesicles and polarization of growth.

The COOH-terminal domain of Myo2p, a yeast myosin V, has a direct role in secretory vesicle targeting.

MYO2 encodes a type V myosin heavy chain needed for the targeting of vacuoles and secretory vesicles to the growing bud of yeast. Here we describe new myo2 alleles containing conditional lethal mutations in the COOH-terminal tail domain. Within 5 min of shifting to the restrictive temperature, the polarized distribution of secretory vesicles is abolished without affecting the distribution of actin or the mutant Myo2p, showing that the tail has a direct role in vesicle targeting. We also show that the actin cable-dependent translocation of Myo2p to growth sites does not require secretory vesicle cargo. Although a fusion protein containing the Myo2p tail also concentrates at growth sites, this accumulation depends on the polarized delivery of secretory vesicles, implying that the Myo2p tail binds to secretory vesicles. Most of the new mutations alter a region of the Myo2p tail conserved with vertebrate myosin Vs but divergent from Myo4p, the myosin V involved in mRNA transport, and genetic data suggest that the tail interacts with Smy1p, a kinesin homologue, and Sec4p, a vesicle-associated Rab protein. The data support a model in which the Myo2p tail tethers secretory vesicles, and the motor transports them down polarized actin cables to the site of exocytosis.

[Upper ankle joint prostheses in haemophilia patients]. / OSG-Prothesen bei hämophilien Patienten.

UNLABELLED: Secondary arthrosis of the upper ankle joint (talocalcanean joint) is one of the most frequent forms of haemophilic arthropathy. It is a secondary form of arthrosis not only because of chronic synovitis and cartilage injury resulting from chronic recurrent intraarticular bleeds, but also due to the misalignment of the joint and abnormal joint stress. The consequences are manifest even in young patients and finally lead to subtalar joint ankylosis with the biomechanical disorder of foot drop (talipes equinus). In such clinical situations, implantation of a subtalar joint endoprosthesis is a viable alternative to arthrodesis. CASE REPORT: A man (age: 52 years), suffering from severe haemophilia A (residual FVIII activity < 1 %), no inhibitor formation. The patient has a history of several years of painful ankylosis of the right ankle joint and minor talipes equinus, and suffers from symptomatic talonavicular arthrosis. Open joint cleansing considerably improved mobility in the upper ankle joint and alleviated the pain in the talonavicular joint. However, the recovered mobility of the arthrotic upper ankle joint also activated the patient's arthrosis, associated with severe pain. As there was no contraindication to upper ankle joint replacement, a cement-free prosthesis was implanted. Three months after surgery, the patient was mobile, with good foot rolling properties without orthopedic aids, and without pain in his upper joint ankle. CONCLUSION: In terms of biomechanics an upper ankle joint endoprosthesis is a superior alternative to arthrodesis in haemophilia patients. In order to minimize the complication rate, the treatment of haemophilia patients should be restricted to specially equipped interdisciplinary treatment centers with adequately trained and experienced surgeons as well as haemostaseologists.

Evaluation of a follow-up program for mild traumatic brain injury in schoolchildren.

INTRODUCTION: Mild traumatic brain injury is a common condition in childhood. Although classified as mild, post-concussive symptoms may persist and interfere with daily activities. Because no established guidelines exist with respect to follow-up medical care for these children, there may be a delay in receiving appropriate care. We developed a follow up program to screen for persistent symptoms and if necessary, refer patients for further medical assistance. METHODS: From July 2010 until December 2013, eligible children aged 4-18 years who presented after sustaining a mild traumatic brain injury were included. All patients received a phone call after 6 weeks. After a period of 3 months, both their schoolteacher and parents were asked to complete in a questionnaire. The results were discussed monthly by a multidisciplinary team. RESULTS: A total of 305 children were enrolled in our follow-up program. Headache was the most common acute symptom upon presentation (63%). Overall, 19% of all patients had problems, either at 6 weeks or 3 months. 14% of these patients were referred for special care. Most common persistent post-concussive symptoms were headache (32%), cognitive problems (23%) and behavioural problems (16%). After a period of two years, a review of patient charts revealed that all of the problems were resolved. CONCLUSION: One fifth of the children exhibit post-concussive symptoms after mild traumatic brain injury. Education of patients and caregivers and a follow up visit if needed applied appropriate care at an early stage to minimise physical and mental problems.

Loss of heterozygosity and p53 gene mutations in breast cancer.

Immunopositivity for the p53 tumor suppressor gene product was evaluated in 133 breast cancers and compared to loss of heterozygosity (LOH) at various chromosomal loci. The validity of p53 immunopositivity as an indicator for p53 mutations was verified using two molecular assays of p53 mutations: single stranded conformational polymorphism (32 cases) and/or direct sequencing (14 cases). Immunopositivity was highly specific for mutations, since all of 15 strongly immunopositive tumors (> 10% of the cells are positive) and seven of nine cases with borderline immunopositivity had mutations by molecular analysis but were somewhat lower in sensitivity, p53 mutations being also detected in three of 23 (13%) immunonegative cases. LOH was measured at loci on the following chromosomes (1p,q; 2p; 3p; 7q; 11p,q; 13q; 16q; 17p; 18p,q; and 22q) by Southern blotting, polymerase chain reaction amplification of restriction fragment length polymorphisms, or repetitive cytidine and adenine stretches (CA repeats). There was no association between p53 mutations and one measure of genomic instability, namely, high incidence of overall LOH. In contrast, p53 mutations strongly associated with LOH at two specific loci, 3p24-26 (P < 0.001) and 7q31 (P < 0.05). There was no association between p53 mutations and LOH at 17p (site of the p53 gene), suggesting that breast cancers often have only one defective allele of the p53 gene.

A candidate tumor suppressor gene in human breast cancers.

We have isolated a candidate gene (designated Brush-1) located at 13q12-q13, proximal to the retinoblastoma gene (RB1). Brush-1 codes for a 4.7-kilobase mRNA expressed at high levels in normal breast epithelium but drastically reduced in 6 of 13 breast cancer cell lines. RB1 mRNA expression is at normal levels for 5 of these 6 lines suggesting a greater importance of Brush-1 for breast cancer. Four primary breast tumors which showed no loss of heterozygosity in the 13q13-q14 region demonstrated normal levels of mRNA for both Brush-1 and RB1. However, four additional primary tumors which displayed loss of heterozygosity for this region had markedly decreased levels of Brush-1 mRNA while maintaining the normal levels for RB1. This differential loss of Brush-1 mRNA expression for both primary tumors and breast cancer cell lines is the expected pattern for a breast tumor suppressor gene.

Prevalence and awareness of functional and structural foot abnormalities in children and adolescents with type 1 diabetes.

BACKGROUND: The ISPAD guideline 2011/2014 advises annual podiatric screening to detect foot complications and identify other possible risk factors such as functional and structural foot abnormalities. The aim of this study was to assess the prevalence and awareness of neurovascular, functional and structural foot abnormalities in children and adolescents with type 1 diabetes mellitus (T1DM). METHODS: All patients aged 0-18 years with T1DM were invited for a foot examination and structured questionnaire by a certified podiatrist. RESULTS: Three of the examined patients (n=128) showed signs of possible long-term complications. Structural and functional foot problems were observed in 71.9% (n=92). Of all patients, 38.3% (n=49) required further treatment by a podiatrist. CONCLUSIONS: Functional and structural foot abnormalities are a frequent problem in children with T1DM in the southern part of the Netherlands. These abnormalities are an additional risk factor for developing long-term foot complications. Education in and implementation of the guideline are necessary.

Characterization of the AdhSL regulatory mutation in Drosophila melanogaster.

We describe the characterization of a previously reported control mutation, AdhSL, in the alcohol dehydrogenase gene of Drosophila melanogaster, which results in decreased production of ADH molecules and subsequently lower ADH activity in adults. We find that the regulatory element modifies ADH mRNA levels and acts cis on both ADH protein and mRNA. It is not promoter specific but is developmentally specific to the adult stage. The AdhSL allele carries a 4.5-kb insert approximately 3 kb 5' to the distal promoter. This new insertion may be responsible for the regulatory phenotype of AdhSL.

Developmental regulation of murine mammary progesterone receptor gene expression.

Previously we have shown that in normal murine mammary glands progesterone receptor (PgR) levels are modulated as a function of development and differentiation such that lactating mammary glands do not contain detectable levels of PgR as measured by steroid binding. The objective of the present study was to determine whether the lack of steroid binding in lactating mammary glands was due to the absence of receptor protein and if so whether it was accompanied by an alternation in the pattern of PgR gene expression. Accordingly, we have performed an immunological analysis of murine mammary PgR isolated from different developmental states and have also examined these tissues for PgR gene expression. In mammary tissues from all developmental states other than lactation, immunoreactive PgR corresponding to both A and B forms of the protein were detected. Analysis for PgR mRNA revealed multiple species in mammary tissues and the relative order of abundance of the various transcripts and their sizes were approximately 6.9 greater than 8.7 greater than 3.5 greater than 2.7 greater than 4.2. The 6.9 and 8.7 kilobase transcripts accounted for between 70-80% of total mRNA. All five species of mRNA were detected in tissues from nulliparous mice which decreased dramatically during pregnancy, became undetectable during lactation, and were once again detectable in tissues from mice undergoing lactational involution. Experiments designed specifically to examine the effect of estradiol on mammary PgR mRNA revealed that in contrast to tissues from other developmental states, lactating mammary glands were unable to respond to estradiol with an increase in PgR mRNA. Based on these findings and the fact that estrogenic insensitivity of lactating mammary glands coexists with the presence of ER we propose that in this tissue there is an alteration in the estrogen dependent transcriptional regulation of PgR gene expression.

Involvement of thymosin beta 4 and endoproteinase Asp-N in the biosynthesis of the tetrapeptide AcSerAspLysPro a regulator of the hematopoietic system.

It is shown that AcSDKP a new regulator of the hematopoietic system can be generated from thymosin beta 4 by a one-step enzymatic cleavage in vitro and in vivo. AcSDKP and T beta 4 were both detected in bone marrow cells (BMC). Incubation of [3H]T beta 4 with either intact or lysed BMC led to the formation of [3H]AcSDKP whereas the labelled tetrapeptide was not degraded under these conditions. Model enzymatic degradation of T beta 4 carried out with bacterial enzymes suggests that a mammalian endoproteinase Asp-N might be involved in the formation of AcSDKP through the specific cleavage of the 4Pro-5 Asp peptide bond of T beta 4.

Fibrinogen heterogeneity in homozygous plasminogen deficiency type I: further evidence that plasmin is not involved in formation of LMW- and LMW'-fibrinogen.

Human plasma fibrinogen is heterogeneous in SDS-polyacrylamide gel electrophoresis and other methods for separation of proteins by molecular size. A high molecular weight fraction (HMW-fibrinogen, 340 kD) contributes approximately 50% of total fibrinogen antigen. Low molecular weight fibrinogen (LMW-fibrinogen, 300 kD) adds another 40%. The residual amount is LMW'-fibrinogen with a molecular weight of 280 kD, and a small amount of very high molecular weight fibrinogen (Fib420), the product of alternative splicing of the A alpha-chain genetic information, resulting in an extended A alpha-chain C-terminus. Fibrinogen was detected by specific immunostaining of nonreduced SDS-PAGE gel immunoblots with antibodies against fibrinopeptide A. Using densitometric scans of the immunoblots we found a ratio of HMW-, LMW- and LMW'-fibrinogen in a patient with homozygous plasminogen deficiency that was similar to the ratio found in immunoblots of plasma from healthy blood donors. Treatment with plasminogen concentrate resulted in a slight decrease of the proportion of HMW-fibrinogen, followed by an increase to 78%. The LMW'-fibrinogen band gained intensity initially, increasing to 11.9% of fibrinogen antigen 6 h after starting plasminogen infusion, but then dropped to levels below detection limit of the immunoblotting assay. LMW-fibrinogen remained constant during the initial 72 h of plasminogen treatment, then dropping to values in the range of 22-25% afterwards. The proportion of HMW-, LMW-, and LMW'-fibrinogen again reached the initial levels two weeks after starting treatment with plasminogen concentrate. We conclude that plasminogen is not involved in the limited proteolysis leading to formation of LMW-fibrinogen and LMW'-fibrinogen in the absence of a generalized fibrinolytic condition. Fibrinolytic activation may lead to the formation of fibrinogen degradation product X, which appears in a similar position as LMW'-fibrinogen in SDS-PAGE.

Visual pattern evoked responses and blink reflexes in assessment of MS diagnosis. A clinical study of 135 multiple sclerosis/pathol.

VEPs were measured after pattern reversal in 135 MS patients and 30 control subjects. Neurological findings were documented in a standard manner. An extensive ophthalmological examination of all subjects was part of the study. The latency of P2 was abnormally delayed in 82% of the "definite", in 60% of the "probable" and in 65% of the "possible" MS groups respectively. The VEP was more often delayed in relation to ophthalmological disturbances. Changes in the MS classification had to be made in more than 10% of the patients, due to delay of VEP latency. These were patients with a "spinal" form of MS, which is known to create diagnostic problems. Optically and electrically evoked blink reflexes were recorded in 107 MS patients. All patients with mesencephalic lesions had delayed responses of the optically evoked reflex. 74% of the patients with caudal brainstem lesions had delayed latencies of the components of the electrically evoked blink reflex. The blink reflex was delayed in 18 additional patients without brainstem signs. The possibility of delineating clinically silent brainstem lesions by investigating blink reflexes is discussed.

The ocular penetration of oral sparfloxacin in humans.

The penetration of sparfloxacin into the aqueous humor after oral administration was studied in 28 patients undergoing cataract surgery. Each patient received a single, oral dose of 400 mg of sparfloxacin. In eight other patients scheduled to undergo vitreal surgery, multiple daily oral doses were administered for a total amount of 1,000 mg. The aqueous levels were (mean +/- SEM) 0.127 +/- 0.036 microgram/ml to 0.404 +/- 0.159 microgram/ml from two to 24 hours after ingestion. In the vitreous, the mean drug level was 0.840 microgram/ml (range, 0.480 to 2.060 microgram/ml), from 4.3 to 8.0 hours after the most recent oral dose. Blood samples obtained at the same time as vitreous and aqueous taps were assayed by high-performance liquid chromatography. These data demonstrate that therapeutic levels of sparfloxacin may be achieved in noninflamed, noninfected eyes undergoing cataract or vitreous surgery.

Treatment of severe chronic bronchitis with ceftazidime.

The aim of this study was to determine whether ceftazidime is more effective than other antibiotics in the treatment of severe acute exacerbation of chronic bronchitis, particularly when other antibiotics have previously failed. Our investigations showed that ceftazidime is effective and well tolerated in patients with severe chronic bronchitis and purulent sputum. The effectiveness of ceftazidime was seen in a reduction of the volume of sputum produced daily and in a colour change from yellow-green to clear-white. The absence of an effect on parameters of lung function was not unexpected since our patients had long-lasting chronic bronchitis.

GC-MS analysis of hydrophobic root exudates of sorghum and implications on the parasitic plant Striga asiatica.

Striga asiatica is a parasitic angiosperm that responds to germination stimulants produced by host plants, including many grasses. GC-MS analyses of hydrophobic root exudates of sorghum revealed the root exudates to be composed of fatty acids, resorcinol, and a series of structurally related hydroquinones, three of which were previously unknown. High yields of resorcinol and the hydroquinone series were detected in sorghum. At least one of the hydroquinones induces germination in Striga, and the resorcinol is thought to stabilize the hydroquinones in the soil. The previously unknown series of hydroquinones offers insight into the possible biosynthesis of the components of the exudate and their possible importance in initiating Striga germination.