Studies on the effect of temperature and pH on the inactivation of fish viral and bacterial pathogens.

Disposal of fish by-products in the European Community must comply with Regulation (EC) No 1069/2009 which categorizes animal by-products according to risk, and specifies methods of disposal of by-products according to that risk. There is provision under the regulation for composting or ensiling to be used for by-products from aquatic animals. Biosecurity considerations require knowledge of the parameters of time and temperature, or time and pH, required to inactivate any fish pathogens that may be present. To provide those data, we undertook laboratory studies on the inactivation of a number of fish pathogenic viruses and bacteria at 60 °C, pH 4.0 and pH 12.0 as a preliminary to conducting subsequent trials with the most resistant viruses and bacteria in fish tissues. The most resistant bacterium to 60 °C, pH 4.0 as well as pH 12.0 was Lactococcus garvieae. Its concentration was reduced to the level of sensitivity of the test after 24-48 h exposure to 60 °C, but it survived for at least 7 days at pH 4.0 and 14 days at pH 12.0. The most resistant virus to 60 °C was infectious pancreatic necrosis virus, and to pH 12.0 was infectious salmon anaemia virus. The majority of the viruses tested survived exposure to pH 4.0 for up to 28 days. The results suggest that the process of acid ensiling alone is not an effective method for the inactivation of many viral and bacterial pathogens, and fish by-products would need further treatment by a method approved under the regulation following ensiling, whereas alkaline or heat treatment are likely to provide an increased degree of biosecurity for on-farm processing of mortalities.

Estimation of infectious dose and viral shedding rates for infectious pancreatic necrosis virus in Atlantic salmon, Salmo salar L., post-smolts.

Infectious dose and shedding rates are important parameters to estimate in order to understand the transmission of infectious pancreatic necrosis virus (IPNV). Bath challenge of Atlantic salmon post-smolts was selected as the route of experimental infection as this mimics a major natural route of exposure to IPNV infection. Doses ranging from 10(2) to 10(-4) 50% end-point tissue culture infectious dose (TCID(50)) mL(-1) sea water were used to estimate the minimum infectious dose for a Scottish isolate of IPNV. The minimum dose required to induce infection in Atlantic salmon post-smolts was <10(-1) TCID(50) mL(-1) by bath immersion (4 h at 10 degrees C). The peak shedding rate for IPNV following intraperitoneal challenge using post-smolts was estimated to be 6.8 x 10(3) TCID(50) h(-1) kg(-1) and occurred 11 days post-challenge. This information may be incorporated into mathematical models to increase the understanding of the dispersal of IPNV from marine salmon sites.

Isolation and quantification of infectious pancreatic necrosis virus from ovarian and seminal fluids of Atlantic salmon, Salmo salar L.

Methods for the isolation and quantification of infectious pancreatic necrosis virus (IPNV) from ovarian and seminal fluids of Atlantic salmon are described. Both have utility for the non-lethal detection of IPNV in mature broodstock and for research into vertical transmission. Two experiments are described to check the efficiency of an elution method for the removal of IPNV from milt. The isolation rate for ovarian fluid of females was generally higher than that for seminal fluid of males from the same populations. In IPNV milt mixing experiments up to 99.98% of available IPNV adsorbed to Atlantic salmon spermatozoa and 20-100% of virus eluted using a variety of procedures. Titration of virus from naturally infected milt can be useful in estimating the relative vertical transmission risk from male broodstock.

[Pharmacological treatment of neurogenic detrusor hyperactivity: intrathecal drugs]. / Traitement pharmacologique de l'hyperactivité détrusorienne neurologique: drogues intrathécales.

Intrathecal clonidine has been tested in spinal cord injury patients not supporting or resistant to anticholinergic drugs. Although the acute effect of clonidine on urodynamic parameters was satisfactory, cardiovascular adverse effects limited the long-term efficacy of this treatment. Intrathecal baclofen has a limited effect on overactive bladder in patients with spinal spasticity, but can modify the quality of erections and induce an incapacity to trigger ejaculation by vibratory penile stimulation.

[Radiculopathy following intrathecal baclofen pump implantation]. / Radiculalgie après implantation de pompe à baclofène.

INTRODUCTION: Intrathecally delivered baclofen has been used as a treatment for severe spasticity since 1984. After a successful intrathecal baclofen trial, a programmable drug delivery system was implanted. Few early complications such as infection or hematoma are observed after this surgery. OBJECTIVE: To describe an unusual and unknown complication of intrathecal baclofen therapy. METHOD: We report 2 cases of complications of intrathecal baclofen therapy, radiculalgy, that appeared early after pump implantation. The clinical symptoms and computed tomography (CT) results are described. RESULTS: The first patient described pain, which evoked left S1 radiculopathic features. The second had left L5 radiculopathic involvement. The mean pain level was estimated on a 10-point visual analog scale as 7.5 (range 4-9). Lumbar CT scan showed a conflict between the symptomatic root and the catheter and eliminated other causes of the symptoms. Treatment with analgesic drugs was successful in 1 patient. The other presented with proximal disconnection of the catheter, which led to surgical replacement of the catheter. The pain disappeared after this surgery. CONCLUSION: Intrathecal baclofen therapy with a subcutaneously implanted progammable pump can be complicated by radiculalgy secondary to a conflict between the catheter and symptomatic root. The diagnosis is made by CT lumbar scan. If medical treatment is not sufficient, surgery could be proposed to replace the catheter.

Intrathecal baclofen for treatment of spasticity of multiple sclerosis patients.

We conducted a retrospective study of the case files of 64 multiple sclerosis (MS) patients presenting severe spasticity, who had received intrathecal (IT) baclofen test injections between 1992 and 2004 in a rehabilitation unit. In almost all cases of our series, IT baclofen was proposed to patients who were no longer able to walk. IT baclofen is a safe and effective treatment to reduce spasticity in MS patients. Despite an advanced stage of the disease at the time of pump placement, the complication rate was low and the efficacy of this treatment was maintained over time.

Infectious pancreatic necrosis virus in Atlantic salmon, Salmo salar L., post-smolts in the Shetland Isles, Scotland: virus identification, histopathology, immunohistochemistry and genetic comparison with Scottish mainland isolates.

During mid-June 1999 peak mortalities of 11% of the total stock per week were seen at a sea cage site of Atlantic salmon, Salmo salar L., post-smolts in the Shetland Isles, Scotland. Virus was isolated on chinook salmon embryo (CHSE) cells in a standard diagnostic test and infectious pancreatic necrosis virus (IPNV) identified by enzyme-linked immunosorbent assay. IPNV was confirmed as serogroup A by a cell immunofluorescent antibody test using the cross-reactive monoclonal antibody AS-1. Four weeks after the main outbreak, virus titres in surviving moribund fish were assayed at >10(10) TCID50 g(-1) kidney. Histopathology of moribund fish was characterized by pancreatic acinar cell necrosis and a marked catarrhal enteritis of the intestinal mucosa. In the liver, necrosis, leucocytic infiltration and a generalized cell vacuolation were noted. IPNV-specific immunostaining was demonstrated in pancreas, liver, heart, gill and kidney tissue. The nucleotide sequence of the coding region of segment A was determined from the Shetland isolate. A 1180 bp fragment of the VP2 gene of this isolate was compared with a 1979 reference isolate from mainland Scottish Atlantic salmon, La/79 and another more recent mainland isolate, 432/00. Both A2 isolates were derived from carrier fish without signs of IPN and serotyped by a plaque neutralization test. The Shetland isolate shows a different nucleotide and amino acid sequence compared with the two isolates from carrier fish. These latter isolates showed identical amino acid sequences in the fragment examined, despite the 21 years separating the isolations. Sequence comparisons with other A2 (Sp) isolates on the database confirm all three Scottish isolates are A2 (Sp).

Post-activation depression in various group I spinal pathways in humans.

This investigation was designed to study the effects of post-activation depression in different spinal pathways fed by group I afferents available to investigation in human subjects. It was precipitated by a recent investigation in the cat showing that-contrary to the general assumption-post-activation depression is not a widespread phenomenon in the spinal cord. In 24 healthy subjects comparison was made between the effects of low and high-test stimulus rates on the monosynaptic Ia excitation, known to be subject to post-activation depression, and on oligosynaptic pathways fed by group I afferents. Both the amplitude of monosynaptic H reflexes and the amount of heteronymous monosynaptic Ia facilitation were significantly smaller at high than at low-test stimulus rates (1-2 s compared with 6-8 s between two consecutive stimuli). So was the amount of reciprocal Ia inhibition of tibialis anterior motoneurones. In contrast, the amount of other non-monosynaptic group I effects directed to the same motor nuclei (peroneal-induced excitation of quadriceps motoneurones, disynaptic non-reciprocal group I inhibition of flexor carpi radialis motoneurones, and D1 inhibition of flexor carpi radialis and soleus H reflexes) were enhanced at high stimulus rates. Results in humans confirm that post-activation depression depends on the type of group I afferents, and/or on the target neurones. The functional significance of the discrepancy between post-activation depression in pure Ia pathways and in other group I pathways is discussed with regard to the fusimotor-driven servo-assistance from Ia afferent discharges.

Evidence for cognitive processes involved in the control of steady state of walking in healthy subjects and after cerebral damage.

OBJECTIVE: The involvement of cognitive processes in the control of walking at steady state was studied in 10 healthy subjects and 18 subjects after unilateral vascular brain damage. A dual task paradigm was used to compare the performance level of a probe reaction time (RT) in sitting (simple task) and during standing or walking on a treadmill (dual task conditions). In this latter condition, RTs were classified and analyzed in relation to the different phases of the gait cycle. RESULTS: The results show a marked increase in RT while walking compared to sitting and standing only in stroke subjects. Specific changes in RTs related to the gait cycle phases were observed in both healthy subjects and those after brain damage. It is concluded that walking at steady state is attentionally demanding. The phase-dependent modulations of the RTs suggest that cognitive processes may play a role in the control of the step cycle. The increase of attentional demand during walking in subjects who had suffered a stroke varies, depending on severity of impairments of walking but also on a reduced general attentional capacity. The dual task paradigm provides a sensitive tool in the assessment of walking ability in stroke subjects.

[Botulinum toxin and spinal cord injury]. / Toxine botulique et paraplégie.

OBJECTIVE: To realize a clarification about the interest of the use of botulinum toxin in spinal cord injured patients. METHOD: Interrogation of Medline database (crossing of botulinum toxin and spinal cord injury). We have also analyzed data from R. Poincaré Hospital in 2001. RESULTS: Three articles of the twenty-five selected, treated effectively of botulinum toxin effect in the limbs muscles of spinal cord injured patients. DISCUSSION: There are some indications of botulinum toxin in spinal cord injured patients ASIA C and D when spasticity induce focal functional discomfort. Indications are exceptional for ASIA A and B patients. No study showed improvement of functional abilities after botulinum toxin injection in spinal cord injured patients. Published studies only covered small number of patients.

[Efficacy of botulinum toxin A for the treatment of detrusor hyperreflexia]. / Effet de la toxine botulique A intradétrusorienne pour le traitement de l'hyperactivité vésicale d'origine neurologique.

OBJECTIVE: To determinate the efficacy of botulinum toxin to treat refractory urinary incontinence due to bladder hyperreflexia. METHOD: The international medical literature was reviewed from the Medline and Pubmed database. DISCUSSION: The usual first line treatment of detrusor hyperreflexia is parasympathicolytic drugs and self cathterization. In case of lack of efficacy or severe side effects, a surgical procedure (enterocystoplasty) can be performed. 300 units of Botox injected into the detrusor permit a significant increase of bladder capacity and a significant decrease of maximal detrusor pressure for at last 6 months. But we did not find any double blind controlled studies in this indication and no fundamental studies focused on the mechanism of action of botulinum toxin on the bladder muscle. It is now impossible to conclude on the long term efficacy and toxicity. CONCLUSION: Botulinum toxin injected into the detrusor muscle seems to be an efficient treatment of bladder hyperreflexia for 6 months in patients resistant to parasympathicolytic drugs. Long term efficacy and mechanism of action is actually not known.

[Clinical evaluation of spasticity]. / Evaluation clinique de la spasticité.

Spasticity is one component of the upper motor neurone syndrome. Unlike motor deficit and co-contraction, spasticity is responsive to treatment. Thus it receives more attention than the other features of the upper motor neuron syndrome. In order to treat correctly the patients, we need to know expectations of the patient and family. Detailed clinical evaluation including functional analysis is necessary to determine whether the objectives are reasonable. This evaluation is not always very easy because of complex clinical features. Simply aiming to reduce spasticity is not a final treatment goal. There must be a purpose behind it. It is actually very difficult to evaluate functional effect of spasticity treatments. Few studies show a real functional improvement. Yet patients are satisfied with the results and the goals are often achieved. One explanation is that functional scales usually used do not fit to the spasticity problem.

[Intrathecal therapy for bladder hyperreflexia]. / Traitement de la vessie hyperactive par les drogues intrathécales.

Intrathecal clonidine was tested for the control of bladder hyperreflexia resistant to pararsympathicolytic drugs in spinal cord injured patients. Urodynamic parameters were significantly improved after acute bolus of intrathecal low doses of clonidine. Unfortunately, the chronic intrathecal infusion of clonidine induced cardiovascular side-effects. Intrathecal baclofen did not modified significantly neurogenic bladder dysfunction from spinal lesion. But intrathecal baclofen modified penile erection quality and impaired the ejaculation reflex induced by penile vibratory stimulation in the same population.

Isolation of salmon pancreas disease virus (SPDV) in cell culture and its ability to protect against infection by the 'wild-type' agent.

A Scottish salmon pancreas disease virus (SPDV) has been isolated and its optimum growth conditions determined. Although several fish cell lines have been tested, successful culture was achieved only with CHSE-214 cells. Cytopathic effects were observed after 5 days. The highest virus titres, calculated by microtitration assay, were reached at 15 degrees C. After 7-9 days post-inoculation, CHSE-214 cell supernatants contained between 10(7)-10(5) TCID50 ml(-1) The cultured isolate is chloroform- and pH 3.0-sensitive, and virions are 50-60 nm in diameter. These characteristics are similar to the Irish SPDV isolates. The culture isolate induced typical pancreas disease (PD) lesions in experimentally infected Atlantic salmon and convalescent fish were resistant to experimental infection with PD-infective kidney homogenates obtained by serial in vivo passages from a PD-infected farmed salmon (termed wild-type SPDV). Furthermore, fish immunised with the inactivated cultured virus were protected against a cohabitation challenge with the wild-type virus. Immunised fish sera showed virus-neutralising activity before challenge (7 weeks post-immunisation) and from 3-6 weeks post-challenge, when sera from non-immunised fish did not neutralise the virus. At 6 weeks post-cohabitation challenge, previously immunised fish had neutralising titres of up to 1:65. Following intraperitoneal (i.p.) challenge, immunised fish showed neutralising titres as high as 1:226 at 8 weeks post-challenge. Non-immunised fish injected i.p. with the wild-type virus developed serum-neutralising activity against the cultured isolate when sampled 8 weeks after infection, confirming an antigenic relationship between the wild-type and cultured virus. The results demonstrate that the tissue culture-adapted isolate of SPDV could be successfully used to protect against challenge by the wild-type virus and could therefore have potential use as an inactivated vaccine against PD.

Antigenic comparison of a truncated form of VP2 of infectious pancreatic necrosis (IPN) virus expressed in four different cell types.

A truncated form of the structural protein VP2 (truncVP2) of infectious pancreatic necrosis (IPN) virus encompassing amino acids 147-307 was expressed in bacterial, yeast, piscine and mammalian cells. All four recombinant antigens were recognised by a VP2-specific monoclonal antibody by ELISA and immunoblot analysis. However, the minimum amount of r-truncVP2 needed for detection by these methods varies depending on the cell type used for expression. Furthermore, all four recombinant preparations, when used to immunise Atlantic salmon, were capable of inducing antibodies reactive with whole IPNV in ELISA.

Distribution of viral haemorrhagic septicaemia virus in wild fish species of the North Sea, north east Atlantic Ocean and Irish Sea.

A surveillance programme was initiated on the occurrence and distribution of viral haemorrhagic septicaemia virus (VHSV) in wild marine fish. Six research cruises were undertaken in an 18 mo period during 1997 and 1998, covering the North Sea, the Atlantic waters off the north and west coasts of Scotland and the Irish Sea. A total of 19,293 fish were sampled from 23 different species including cod, haddock, Norway pout, herring and sprat. Individual fish lengths were recorded and the fish were checked for lesions, haemorrhaging and other signs of disease. Pools of organ samples were taken for virus assay. The majority of fish sampled did not display clinical signs indicative of viral haemorrhagic septicaemia. A small number of cod were found with skin lesions and haddock with skin haemorrhaging. Of the 2081 organ and skin sample pools collected, 21 tested positive for VHSV by tissue culture and enzyme-linked immunosorbent assay. Seventeen of the isolates originated from Norway pout Trisopterus esmarkii, one from cod Gadus morhua (skin lesion), one from herring Clupea harengus, one from whiting Merlangius merlangus, and one from a previously unreported host species, poor cod Trisopterus minutus.

Isolation and identification of Viral Haemorrhagic Septicaemia (VHS) viruses from cod Gadus morhua with the ulcus syndrome and from haddock Melanogrammus aeglefinus having skin haemorrhages in the North Sea.

During fish disease surveys for marine rhabdoviruses in 1993 and 1995, the cod ulcus syndrome was seen widely in all ages of cod, especially the 2 to 5+ year classes. Viral Haemorrhagic Septicaemia Virus (VHSV) was isolated from a small proportion of the lesion-positive fish and these isolates were identified by immunofluorescence or ELISA. A serendipitous observation of dermal petechiae on haddock was made. VHSV was isolated from this lesion for the first time indicating a new host species for VHSV.