Sensibilidade toxicológica e especificidade do teste de microfixação de complemento na detecção de toxinas botulínicas C e D em meio de cultura e fígado de camundongos

No presente estudo pretendeu-se verificar a sensibilidade toxicológica e especificidade do Teste de Microfixação de Complemento (MCF) na detecção de toxinas botulínicas C e D no sobrenadante de cultivos bacterianos e em fígados de camundongos inoculados com doses letais e subletais. As toxinas foram produzidas em meio de cultura Hemoline, tituladas através da determinação da DL50 pelo Bioensaio em Camundongo e diluídas nas concentrações de 10, 1, 0,1, 0,01 e 0,001 DL50. Desta forma, foram utilizadas em dois modelos experimentais, onde foi determinada a sensibilidade toxicológica do MCF no sobrenadante do meio de cultura com as diluições descritas acima e ainda em extratos hepáticos de camundongos com peso corporal de 20g, inoculados com as mesmas diluições. A tentativa de evidenciação das toxinas botulínicas nos extratos hepáticos de camundongos foi realizada através da sua extração após a morte pela administração das doses letais e ainda pelo sacrifício dos animais inoculados com doses subletais, em intervalos de 5 dias. Os resultados evidenciaram uma sensibilidade toxicológica para o MCF de 100% para os dois tipos de toxinas ao nível de 0,01 DL50, quando testados os sobrenadantes de meio de cultura, portanto 100 vezes superior ao Bioensaio em Camundongo. A sensibilidade toxicológica do MCF, quando examinados extratos hepáticos de camundongos inoculados com 1 e 10 DL50 de toxinas botulínicas C e D, foi inferior, com valores de 100, 80, 89 e 72%, respectivamente. Pelo teste foi possível detectar toxinas botulínicas tipos C e D nos extratos hepáticos de camundongos inoculados com doses subletais até 15 dias após a sua inoculação. A especificidade do MCF foi de 88 e 92%, quando testados extratos hepáticos de camundongos sadios, e confrontados com as antitoxinas C e D; e 100% no sobrenadante do meio de cultura. Os resultados apontam para uma possível utilização do teste como importante instrumento de pesquisa e ainda na eventual substituição dos testes in vivo pelas suas implicações éticas e limitações práticas.

Neonatal immune response of Brazilian beef cattle to vaccination with Clostridium botulinum toxoids types C and D by indirect ELISA

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Niemann-Pick disease type C symptomatology: an expert-based clinical description

Niemann-Pick disease type C (NP-C) is a rare, progressive, irreversible disease leading to disabling neurological manifestations and premature death. The estimated disease incidence is 1:120,000 live births, but this likely represents an underestimate, as the disease may be under-diagnosed due to its highly heterogeneous presentation. NP-C is characterised by visceral, neurological and psychiatric manifestations that are not specific to the disease and that can be found in other conditions. The aim of this review is to provide non-specialists with an expert-based, detailed description of NP-C signs and symptoms, including how they present in patients and how they can be assessed. Early disease detection should rely on seeking a combination of signs and symptoms, rather than isolated findings. Examples of combinations which are strongly suggestive of NP-C include: splenomegaly and vertical supranuclear gaze palsy (VSGP); splenomegaly and clumsiness; splenomegaly and schizophrenia-like psychosis; psychotic symptoms and cognitive decline; and ataxia with dystonia, dysarthria/dysphagia and cognitive decline. VSGP is a hallmark of NP-C and becomes highly specific of the disease when it occurs in combination with other manifestations (e.g. splenomegaly, ataxia). In young infants (<2 years), abnormal saccades may first manifest as slowing and shortening of upward saccades, long before gaze palsy onset. While visceral manifestations tend to predominate during the perinatal and infantile period (2 months–6 years of age), neurological and psychiatric involvement is more prominent during the juvenile/adult period (>6 years of age). Psychosis in NP-C is atypical and variably responsive to treatment. Progressive cognitive decline, which always occurs in patients with NP-C, manifests as memory and executive impairment in juvenile/adult patients. Disease prognosis mainly correlates with the age at onset of the neurological signs, with early-onset forms progressing faster. Therefore, a detailed and descriptive picture of NP-C signs and symptoms may help improve disease detection and early diagnosis, so that therapy with miglustat (Zavesca®), the only available treatment approved to date, can be started as soon as neurological symptoms appear, in order to slow disease progression.

The updated European Consensus 2009 on the use of Botulinum toxin for children with cerebral palsy

An interdisciplinary European group of clinical experts in the field of movement disorders and experienced Botulinum toxin users has updated the consensus for the use of Botulinum toxin in the treatment of children with cerebral palsy (CP). A problem-orientated approach was used focussing on both published and practice-based evidence. In part I of the consensus the authors have tabulated the supporting evidence to produce a concise but comprehensive information base, pooling data and experience from 36 institutions in 9 European countries which involves more than 10,000 patients and over 45,000 treatment sessions during a period of more than 280 treatment years. In part II of the consensus the Gross Motor Function Measure (GMFM) and Gross Motor Function Classification System (GMFCS) based Motor Development Curves have been expanded to provide a graphical framework on how to treat the motor disorders in children with CP. This graph is named "CP(Graph) Treatment Modalities - Gross Motor Function" and is intended to facilitate communication between parents, therapists and medical doctors concerning (1) achievable motor function, (2) realistic goal-setting and (3) treatment perspectives for children with CP. The updated European consensus 2009 summarises the current understanding regarding an integrated, multidisciplinary treatment approach using Botulinum toxin for the treatment of children with CP.

In the human urothelium and suburothelium, intradetrusor botulinum neurotoxin type A does not induce apoptosis: preliminary results

Intradetrusor injections of botulinum neurotoxin type A (BoNTA) are emerging as the preferred second-line treatment for neurogenic and idiopathic overactive bladder (OAB). In animal experiments, intradetrusor BoNTA injections have been shown to cause apoptosis in the bladder urothelium and suburothelium but not the detrusor.

Complete continence after botulinum neurotoxin type A injections for refractory idiopathic detrusor overactivity incontinence: patient-reported outcome at 4 weeks

Objective improvement following intradetrusor injections of botulinum neurotoxin type A (BoNTA) is well documented. Although patient-related outcome measures are highly recommended for monitoring overactive bladder symptoms, no study before has dealt with the question of patient-reported complete continence after BoNTA treatment using validated questionnaires.

Eye movement and diffusion tensor imaging analysis of treatment effects in a Niemann-Pick Type C patient

New treatment options for Niemann-Pick Type C (NPC) have recently become available. To assess the efficiency and efficacy of these new treatment markers for disease status and progression are needed. Both the diagnosis and the monitoring of disease progression are challenging and mostly rely on clinical impression and functional testing of horizontal eye movements. Diffusion tensor imaging (DTI) provides information about the microintegrity especially of white matter. We show here in a case report how DTI and measures derived from this imaging method can serve as adjunct quantitative markers for disease management in Niemann-Pick Type C. Two approaches are taken--first, we compare the fractional anisotropy (FA) in the white matter globally between a 29-year-old NPC patient and 18 healthy age-matched controls and show the remarkable difference in FA relatively early in the course of the disease. Second, a voxelwise comparison of FA values reveals where white matter integrity is compromised locally and demonstrate an individualized analysis of FA changes before and after 1year of treatment with Miglustat. This method might be useful in future treatment trials for NPC to assess treatment effects.

Clean intermittent self-catheterization after botulinum neurotoxin type A injections: short-term effect on quality of life

OBJECTIVE: To investigate the hypothesis that the need for clean intermittent self-catheterization after botulinum neurotoxin type A injections is outweighed by the efficacy of this treatment, so that clean intermittent self-catheterization is not a burden for patients with refractory idiopathic detrusor overactivity. METHODS: Women undergoing intradetrusor injections of 200 units botulinum neurotoxin type A for refractory idiopathic detrusor overactivity were evaluated prospectively. Clean intermittent self-catheterization was discussed with all patients and its possible need after botulinum neurotoxin type A treatment. As indicator of quality of life, lower urinary tract symptom distress and effect on daily activities were assessed using the validated Urogenital Distress Inventory (UDI-6) and Incontinence Impact Questionnaire (IIQ-7) before and 4 weeks after receiving botulinum neurotoxin type A injections. RESULTS: Mean age of the 65 women was 51 years, and all voided spontaneously before botulinum neurotoxin type A injections. After botulinum neurotoxin type A treatment, 28 (43%) required clean intermittent self-catheterization. Mean UDI-6 and IIQ-7 scores reduced from 61 to 33 (P<.001) and 62 to 30 (P<.001) in women performing clean intermittent self-catheterization and from 60 to 28 (P<.001) and 64 to 25 (P<.001) in those who did not, respectively. Comparison of quality of life in women performing clean intermittent self-catheterization and in those who did not revealed no significant differences before and after botulinum neurotoxin type A treatment. CONCLUSION: Clean intermittent self-catheterization after botulinum neurotoxin type A intradetrusor injections did not impair quality of life in appropriately informed and selected women in the short term. All patients should be informed of the potential need for clean intermittent self-catheterization after botulinum neurotoxin type A injections, and a willingness to do so should be a prerequisite for this still unlicensed off-label treatment.

What a patient with refractory idiopathic detrusor overactivity should know about botulinum neurotoxin type a injection

PURPOSE: We documented the effects of intradetrusor injections of botulinum neurotoxin type A (Botox(R)) for refractory idiopathic detrusor overactivity so that prospective patients maybe properly informed about possible improvement in quality of life, the duration of interinjection intervals and the risk of clean intermittent self-catheterization. MATERIALS AND METHODS: A total of 81 consecutive patients with refractory idiopathic detrusor overactivity treated with intradetrusor injections of 200 U botulinum neurotoxin type A at 20 sites per injection course were evaluated in this prospective, nonrandomized, open label cohort study. The primary outcome was changes in quality of life, as assessed by the short form of the Urogenital Distress Inventory and the Incontinence Impact Questionnaire before and after treatment. Secondary outcomes were the interinjection interval and the need for clean intermittent self-catheterization. RESULTS: After intradetrusor botulinum neurotoxin type A injections there was significant improvement in quality of life, which was sustained after repeat injections. Mean Urogenital Distress Inventory and Incontinence Impact Questionnaire scores decreased from 56 to 26 and 59 to 21 after injection 1 in 81 patients, from 52 to 30 and 51 to 24 after injection 2 in 24, from 40 to 19 and 43 to 17 after injection 3 in 13, from 44 to 17 and 61 to 15 after injection 4 in 6 and from 51 to 17 and 63 to 14 after injection 5 in 4, respectively. The median interinjection interval was 15, 12, 14 and 13 months between injections 1 and 2, 2 and 3, 3 and 4, and 4 and 5, respectively. Considering a post-void residual urine of greater than 100 ml with lower urinary tract symptoms as the indication for clean intermittent self-catheterization, the overall clean intermittent self-catheterization rate after treatment was 43%. CONCLUSIONS: Intradetrusor botulinum neurotoxin type A injections for refractory idiopathic detrusor overactivity significantly improved quality of life. This effect was sustained after repeat injection. More than 2 of 5 patients with refractory idiopathic detrusor overactivity required clean intermittent self-catheterization after botulinum neurotoxin type A injections and all prospective patients should be informed about this.

Immune response of horses to vaccination with the recombinant Hc domain of botulinum neurotoxin types C and D

Botulinum neurotoxins, predominantly serotypes C and D, cause equine botulism through forage poisoning. The C-terminal part of the heavy chain of botulinum neurotoxin types C and D (HcBoNT/C and D) was expressed in Escherichia coli and evaluated as a recombinant mono- and bivalent vaccine in twelve horses in comparison to a commercially available toxoid vaccine. A three-dose subcutaneous immunization of adult horses elicited robust serum antibody response in an ELISA using the immunogen as a capture antigen. Immune sera showed dose-dependent high potency in neutralizing specifically the active BoNT/C and D in the mouse protection assay. The aluminium hydroxide based mono- and bivalent recombinant HcBoNT/C and D vaccines were characterized by good compatibility and the ability to elicit protective antibody titers similar or superior to the commercially available toxoid vaccine.

[Occurrence of Clostridium perfringens type A and type C in piglets of the Swiss swine population]

Necrotizing enteritis (NE) of newborn piglets still represents an economical problem in Swiss pig breeding and production. The aim of our study was to identify risk factors for NE and evaluate the prevalence of C. perfringens with the toxingenes cpb and cpb2 in Swiss pig breeding farms. The prevalence of theses C. perfringens was investigated using fecal swabs followed by bacteriological culturing and genotyping. Close proximity to other breeding farms and large herd sizes were shown to predispose to NE. C. perfringens type C, carrying the genes cpa, cpb and cpb2 were frequently identified in herds with acute outbreaks of NE. Farms not affected by NE or those using prophylactic vaccination against NE were predominantly positive for C. perfringens type A strains with cpb2 and showed much lower prevalence of C. perfringens type C, compared to acutely affected herds. Our results demonstrate that C. perfringens type A strains with cpb2 are not associated with NE. Besides typical necropsy finding, only the identification of cpb can be used for the diagnosis of NE in affected herds.