Isolation and characterization of a neprilysin-like protein from Venturia canescens virus-like particles

Maternal protein secretions from endoparasitoid wasps are evolutionary adaptations to regulate host physiology as part of an extended wasp phenotype. Virus-like particles (VLPs) produced in the calyx region of Venturia canescens wasps are involved in immune evasion of the developing parasitoid inside the host. In contrast to polydnaviruses (PDVs), VcVLPs are devoid of any nucleic acids. To understand the role of these particles in the regulation of host physiology and phylogenetic relationship between VLPs and PDVs, it is essential to identify particle proteins. In this paper, we describe the isolation and molecular cloning of a neprilysin-like gene (VcNEP) coding for a 94 kDa VcVLP protein and discuss its possible role in host regulation.

Isolation and characterization of a novel venom protein from an endoparasitoid, Cotesia rubecula (Hym : Braconidae)

Insects are important vectors of diseases with remarkable immune defense capabilities. Hymenopteran endoparasitoids are adapted to overcome the host defense system and, therefore, are useful sources of immune-suppressing proteins. Not much is known about venom proteins in endoparasitoids, especially those that have a functional relationship with polydnaviruses (PDVs). Here, we describe the isolation and characterization of a small venom protein (Vn4.6) from an endoparositoid, Cotesia rubecula, which interferes with the activation of the host hemolymph prophenoloxidose. The coding region for Vn4.6 is located upstream in the opposite direction of a gene coding for a C rubecula PDV-protein (Crp32). Arch. Insect Biochem. Physiol. 53:92-100, 2003. (C) 2003 Wiley-Liss, Inc.

GABAergic REGULATION OF AUDITORY SENSORY GATING IN LOW- AND HIGH-ANXIETY RATS SUBMITTED TO A FEAR CONDITIONING PROCEDURE

The inferior colliculus (IC) is primarily involved in the processing of acoustic stimuli, being in a position to send auditory information to motor centers that participate in behaviors such as prey catching and predators` avoidance The role of the central nucleus of the IC (CIC) on fear and anxiety has been suggested on the basis that rats are able to engage in tasks to decrease the aversiveness of CIC stimulation, increased Fos immunolabeling during diverse aversive states and increased CIC auditory evoked potentials (AEP) induced by conditioned fear stimuli Additionally it was shown that brainstem AEP, represented by wave V, for which the main generator is the IC, is increased during experimentally induced anxiety Rats segregated according to their low or high emotional reactivity have been used as an important tool in the study of fear and anxiety The IC contains a high density of GABA receptors Since the efficacy of an anxiolytic compound is a function of the animal`s anxiety level, it is possible that GABA-benzodiazepine (Bzp) agents affect LA and HA animals differently In this study we investigated the GABA-Bzp influence on the modulation of AEP in rats with low (LA) or high-anxiety (HA) levels, as assessed by the elevated plus maze test (EPM) GABA-Bzp modulation on the unconditioned AEP response was analyzed by using intra CIC injections (0 2 mu l) of the GABA-Bzp agonists muscimol (121 ng) and diazepam (30 mu g) or the GABA inhibitors bicuculline (10 ng) and semicarbazide (7 mu g) In a second experiment, we evaluate the effects of contextual aversive conditioning on AEP using foot shocks as unconditioned stimuli On the unconditioned fear paradigm GABA inhibition in creased AEP in LA rats and decreases this measure in HA counterparts Muscimol was effective in reducing AEP in both LA and HA rats Contextual fear stimuli increased the magnitude of AEP In spite of no effect obtained with diazepam in LA rats the drug inhibited AEP in HA animals The specificity of the regulatory mechanisms mediated by GABA Bzp for the ascending neurocircuits responsible for the acquisition of aversive information in LA and HA animals shed light on the processing of sensory information underlying the generation of defensive reactions (C) 2010 IBRO Published by Elsevier Ltd All rights reserved

Isolation of dopamine D-2 receptor (D2R) promoters in Mugil cephalus

This paper reports the isolation of two putative D2R promoters from grey mullet, one 5' flanking and the other an intronic sequence immediately upstream of the first coding exon. Promoter activity of the intronic sequence was confirmed in vitro through functional analysis using luciferase as reporter gene. The functional characteristics of the region flanking the 5'-UTR is currently under investigation.

Isolation and molecular identification of planctomycete bacteria from postlarvae of the giant tiger prawn, Penaeus monodon

Bacteria phenotypically resembling members of the phylogenetically distinct planctomycete group of the domain Bacteria were isolated from postlarvae of the giant tiger prawn, Penaeus monodon. A selective medium designed in the light of planctomycete antibiotic resistance characteristics was used for this isolation. Planctomycetes were isolated from both healthy and monodon baculovirus-infected prawn postlarvae, The predominant colony type recovered from postlarvae regardless of viral infection status was nonpigmented. Other, less commonly observed types were pink or orange pigmented, A planctomycete-specific 16S rRNA-directed probe was designed and used to screen the isolates for their identity as planctomycetes prior to molecular phylogenetic characterization. 16S rRNA genes from nine prawn isolates together with two planctomycete reference strains (Planctomyces brasiliensis and Gemmata obscuriglobus) were sequenced and compared with reference sequences from the planctomycetes and other members of the domain Bacteria, Phylogenetic analyses and sequence signatures of the 16S rRNA genes demonstrated that the prawn isolates were members of the planctomycete group, Five representatives of the predominant nonpigmented colony type were members of the Pirellula group within the planctomycetes, as were three pink-pigmented colony type representatives. Homology values and tree topology indicated that representatives of the nonpigmented and pink-pigmented colony types formed two discrete clusters within the Pirellula group, not identical to any known Pirellula species, A sole representative of the orange colony type was a member of the Planctomyces group, virtually identical in 16S rDNA sequence to P. brasiliensis, and exhibited distinctive morphology.

Laparoscopic extended cardiomyotomy in children: an effective procedure for the treatment of esophageal achalasia

Purpose: Achalasia of the esophagus is characterized by aperistalsis and incomplete relaxation of the lower esophageal sphincter in response to swallowing. The objective of the present study is to present the experience of a modified Heller myotomy via a laparoscopic approach for the treatment of children who had this condition. Methods: A retrospective review of medical records of all patients who underwent this procedure from 2000 to 2009 was performed. The procedure consisted of an extended esophagomyotomy beginning on the lower part of the lower esophageal sphincter and continuing 5 to 6 cm above on the lower third of the esophagus, and then extended 3 to 4 cm below to the stomach, associated with an anterior 180-degree hemi-fundoplication according to Dor`s technique. Results: Fifteen patients were included in the study. There were 8 female and 7 male patients. Mean operating time was 190 minutes with no intraoperative complications and 1 conversion to open surgery because of difficulty in dissecting an inflamed distal esophagus. In a mean follow-up period of 32.3 months, 2 patients had recurrence of mild dysphagia that disappeared spontaneously, and 1 required a single botulinum toxin injection with complete resolution of symptoms. Conclusion: We conclude that the laparoscopic extended Heller myotomy with Dor fundoplication is a safe and effective method for the treatment for achalasia in the pediatric population even in advanced cases. (C) 2010 Elsevier Inc. All rights reserved.

Transanal endorectal pull-through in children with Hirschsprung`s disease-technical refinements and comparison of results with the Duhamel procedure

Purpose: Transanal endorectal pull-through (TEPT) has drastically changed the treatment of Hirschsprung`s disease (HD). A short follow-up of children Submitted to TEPT reveals results that are similar to the classic transabdominal pull-through procedures. However, few reports compare the late results of TEPT with transabdominal pull-through procedures with respect to complication rates and the fecal continence. The aims of the present work are to describe some technical refinements that we introduced in the procedure and to compare the short and long-term outcome of TEPT with the outcomes of a group of patients with HD who previously underwent the Duhamel procedure. Methods: Thirty-five patients who underwent TEPT were prospectively studied and compared to a group of 29 patients who were treated with colostomy followed by a classical Duhamel pull-through. The main modifications introduced in the TEPT group were no preoperative colon preparation, operation conducted under general anesthesia in addition to regional sacral anesthesia, use of only one purse-string Suture in the rectal mucosa before transanal submucosal dissection, and no use of retractors and electrocautery during file submucosal dissection. Results: The most frequent early complications of TEPT group were perineal dermatitis (22.8%) and anastomotic strictures (8.6%). The comparison with patients who underwent Duhamel procedure revealed no difference in the incidence of preoperative enterocolitis, the patients of the TEPT group were younger at the time of diagnosis and of surgery, they had shorter operating times, and they began oral feeding more quickly after the operation. The incidence of wound infection was lower in the TEPT group. Moreover, the TEPT and Duhamel groups showed no difference in the incidences of mortality, postoperative partial continence, and total incontinence. Although the incidences of complete continence and postoperative enterocolitis were not different, a tendency to the increased incidence in the TEPT group was observed. Conclusions: This study further supports the technical advantages, the simplicity, and the decreased incidence of complications of a primary TEPT procedure when compared to a classical form of pull-through. Sonic technical refinements are described, and no preoperative colon preparation was necessary for the patients studied here. The results show that the long-term outcomes of the modified TEPT procedure are generally better than those obtained with classical approaches. Published by Elsevier Inc.

Isolation of Lautropia mirabilis from sputa of a cystic fibrosis patient

An aggregate-forming coccus, isolated twice as the predominant microorganism in sputa from a cystic fibrosis patient on consecutive days, was shown to belong to the species Lautropia mirabilis on the bases of similarities of 16S rRNA gene sequences and phenotype. These isolates of L. mirabilis appear to be the first reported from a patient with cystic fibrosis and outside of Denmark.

Modeling the optical constants of solids using acceptance-probability-controlled simulated annealing with an adaptive move generation procedure

The acceptance-probability-controlled simulated annealing with an adaptive move generation procedure, an optimization technique derived from the simulated annealing algorithm, is presented. The adaptive move generation procedure was compared against the random move generation procedure on seven multiminima test functions, as well as on the synthetic data, resembling the optical constants of a metal. In all cases the algorithm proved to have faster convergence and superior escaping from local minima. This algorithm was then applied to fit the model dielectric function to data for platinum and aluminum.

Outpatient Radioiodine Therapy for Thyroid Cancer A Safe Nuclear Medicine Procedure

Purposes: To evaluate the dosimetric effect of outpatient radioiodine therapy for thyroid cancer in members of a patient`s family and their living environment, when using iodine-131 doses reaching 7.4 GBq. The following parameters were thus defined: (a) whole-body radiation doses to caregivers, (b) the production of contaminated solid waste, and (c) radiation potential and surface contamination within patients` living quarters. Methods: In total, 100 patients were treated on an outpatient basis, taking into consideration their acceptable living conditions, interests, and willingness to comply with medical and radiation safety guidelines. Both the caregivers and the radiation dose potentiality inside patients` residences were monitored by using thermoluminescent dosimeters. Surface contamination and contaminated solid wastes were identified and measured with a Geiger-Muller detector. Results: A total of 90 monitored individuals received a mean dose of 0.27 (+/- 0.28) mSv, and the maximum dose registered was 1.6 mSv. The mean value for the potential dose within all living quarters was 0.31(+/- 0.34) mSv, and the mean value per monitored surface was 5.58 Bq/cm(2) for all the 1659 points measured. The overall production of contaminated solid wastes was at a low level, being about 3 times less than the exemption level indicated by the International Atomic Energy Agency. Conclusions: This study indicates that the treatment of thyroid cancer by applying radioiodine activities up to 7.4 GBq, on an outpatient basis, is a safe procedure, especially when supervised by qualified professionals. This alternative therapy should be a topic for careful discussion considering the high potential for reducing costs in healthcare and improving patient acceptance.

Application of a wrist dosimeter prototype for radiation monitoring ((153)Sm) during a therapeutic procedure simulation

Gamma and beta radiation emitting radiopharmaceuticals are handled in nuclear medicine services, and in many cases there is only individual monitoring of gamma radiation. In this paper, the results obtained using a wrist dosimeter prototype (CaSO(4):Dy + Teflon pellets) show that the doses for workers occupationally exposed to beta radiation from (153)Sm are not negligible. It is important that this dose is evaluated, and it has to be taken into consideration in the individual monitoring system.

Enolase from Paracoccidioides brasiliensis: isolation and identification as a fibronectin-binding protein

Paracoccidioides brasiliensis yeast cells can enter mammalian cells and may manipulate the host cell environment to favour their own growth and survival. Moreover, fibronectin and several other host extracellular matrix proteins are recognized by various components of the yeast cell extracts. The present study was designed to isolate and characterize a fibronectin-binding protein from P. brasiliensis. We also compared P. brasiliensis strain 18, tested before (Pb18a) and after (Pb18b) animal passage, in relation to its adhesion and invasion processes. Extracts from both samples, when cultured on blood agar solid medium, showed higher levels of protein expression than when the same samples were cultured on Fava-Netto solid medium, as demonstrated by two-dimensional electrophoresis and SDS-PAGE. Also, both Pb18a and Pb18b exhibited stronger adhesion to A549 epithelial cells when cultured on blood agar medium than when cultured on Fava-Netto medium. Ligand affinity binding assays revealed a protein of 54 kDa and pl 5.6 in P. brasiliensis cell-free extracts with the properties of a fibronectin-binding adhesin, which was characterized by tryptic digestion and mass spectroscopy as a homologue of enolase from P. brasiliensis. Antibody raised against this 54 kDa protein abolished 80 % of P. brasiliensis adhesion to A549 epithelial cells. Our results demonstrate that P. brasiliensis produces a fibronectin-binding adhesin, irrespective of the culture medium, and that this activity can be inhibited by a specific antibody and is involved in the adhesion of the fungus to pulmonary epithelial cells.

SUITABILITY OF A RAPID DNA ISOLATION AND AMPLIFICATION FOR DETECTION OF TRYPANOSOMA CRUZI IN TRIATOMA INFESTANS DRY FECAL SPOTS COLLECTED ON FILTER PAPER

A rapid DNA extraction was used for T. cruzi detection in triatomines dry fecal spots collected on filter paper and analyzed by PCR. Fifty T infestans were fed on experimentally infected Balb/C mice with high T. cruzi parasitemia and divided into five groups of len triatomines, and 100 triatomines were infected with lower parasitemia and divided into five groups of 20 triatomines, One dry fecal spot was analyzed per group on days 1, 2, 3, 4 and 5 post feeding. Amplification targeted T. cruzi TCZ sequence and resulted positive from day 4 after bugs feeding in the two models (high and lower parasitemia). The rapid DNA isolation and PCR proposed are suitable for detection of T. cruzi DNA in in filter paper and should be considered in field research.

Isolation and Genotyping of Rubella Virus From a Case of Congenital Infection in Brazil

The incidence of CRS and CRI has decreased markedly worldwide with the implementation of efficient vaccination programs. We report a congenital rubella case with fetal death occurred at 29th week of gestation. RV was confirmed in placenta. The results of phylogenetic analysis showed that the RVs/Sao-Paulo01.- BRA/08.CRI belongs to the genotype 2B of RV. J. Med. Virol. 83:2048-2050, 2011. (C) 2011 Wiley-Liss, Inc.