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Primary cilia are specialised sensory and developmental signalling devices extending from the surface of most eukaryotic cells. Defects in these organelles cause inherited human disorders (ciliopathies) such as retinitis pigmentosa and Bardet-Biedl syndrome (BBS), frequently affecting many physiological and developmental processes across multiple organs. Cilium formation, maintenance and function depend on intracellular transport systems such as intraflagellar transport (IFT), which is driven by kinesin-2 and IFT-dynein motors and regulated by the Bardet-Biedl syndrome (BBS) cargo-adaptor protein complex, or BBSome. To identify new cilium-associated genes, we employed the nematode C. elegans, where ciliogenesis occurs within a short timespan during late embryogenesis when most sensory neurons differentiate. Using whole-organism RNA-Seq libraries, we discovered a signature expression profile highly enriched for transcripts of known ciliary proteins, including FAM-161 (FAM161A orthologue), CCDC-104 (CCDC104), and RPI-1 (RP1/RP1L1), which we confirm are cilium-localised in worms. From a list of 185 candidate ciliary genes, we uncover orthologues of human MAP9, YAP, CCDC149, and RAB28 as conserved cilium-associated components. Further analyses of C. elegans RAB-28, recently associated with autosomal-recessive cone-rod dystrophy, reveal that this small GTPase is exclusively expressed in ciliated neurons where it dynamically associates with IFT trains. Whereas inactive GDP-bound RAB-28 displays no IFT movement and diffuse localisation, GTP-bound (activated) RAB-28 concentrates at the periciliary membrane in a BBSome-dependent manner and undergoes bidirectional IFT. Functional analyses reveal that whilst cilium structure, sensory function and IFT are seemingly normal in a rab-28 null allele, overexpression of predicted GDP or GTP locked variants of RAB-28 perturbs cilium and sensory pore morphogenesis and function. Collectively, our findings present a new approach for identifying ciliary proteins, and unveil RAB28, a GTPase most closely related to the BBS protein RABL4/IFT27, as an IFT-associated cargo with BBSome-dependent cell autonomous and non-autonomous functions at the ciliary base., Competing Interests: The authors have declared that no competing interests exist.

Background: Joubert syndrome (JBTS) and related disorders are defined by cerebellar malformation (molar tooth sign), together with neurological symptoms of variable expressivity. The ciliary basis of Joubert syndrome related disorders frequently extends the phenotype to tissues such as the eye, kidney, skeleton and craniofacial structures., Results: Using autozygome and exome analyses, we identified a null mutation in KIAA0556 in a multiplex consanguineous family with hallmark features of mild Joubert syndrome. Patient-derived fibroblasts displayed reduced ciliogenesis potential and abnormally elongated cilia. Investigation of disease pathophysiology revealed that Kiaa0556 (-/-) null mice possess a Joubert syndrome-associated brain-restricted phenotype. Functional studies in Caenorhabditis elegans nematodes and cultured human cells support a conserved ciliary role for KIAA0556 linked to microtubule regulation. First, nematode KIAA0556 is expressed almost exclusively in ciliated cells, and the worm and human KIAA0556 proteins are enriched at the ciliary base. Second, C. elegans KIAA0056 regulates ciliary A-tubule number and genetically interacts with an ARL13B (JBTS8) orthologue to control cilium integrity. Third, human KIAA0556 binds to microtubules in vitro and appears to stabilise microtubule networks when overexpressed. Finally, human KIAA0556 biochemically interacts with ciliary proteins and p60/p80 katanins. The latter form a microtubule-severing enzyme complex that regulates microtubule dynamics as well as ciliary functions., Conclusions: We have identified KIAA0556 as a novel microtubule-associated ciliary base protein mutated in Joubert syndrome. Consistent with the mild patient phenotype, our nematode, mice and human cell data support the notion that KIAA0556 has a relatively subtle and variable cilia-related function, which we propose is related to microtubule regulation.

Integrity of the Golgi apparatus requires the microtubule (MT) network. A subset of MTs originates at the Golgi itself, which in this case functions as a MT-organizing center (MTOC). Golgi-derived MTs serve important roles in post-Golgi trafficking, maintenance of Golgi integrity, cell polarity and motility, as well as cell type-specific functions, including neurite outgrowth/branching. Here, we discuss possible models describing the formation and dynamics of Golgi-derived MTs. How Golgi-derived MTs are formed is not fully understood. A widely discussed model implicates that the critical step of the process is recruitment of molecular factors, which drive MT nucleation (γ-tubulin ring complex, or γ-TuRC), to the Golgi membrane via specific scaffolding interactions. Based on recent findings, we propose to introduce an additional level of regulation, whereby MT-binding proteins and/or local tubulin dimer concentration at the Golgi helps to overcome kinetic barriers at the initial nucleation step. According to our model, emerging MTs are subsequently stabilized by Golgi-associated MT-stabilizing proteins. We discuss molecular factors potentially involved in all three steps of MT formation. To preserve proper cell functioning, a balance must be maintained between MT subsets at the centrosome and the Golgi. Recent work has shown that certain centrosomal factors are important in maintaining this balance, suggesting a close connection between regulation of centrosomal and Golgi-derived MTs. Finally, we will discuss potential functions of Golgi-derived MTs based on their nucleation site location within a Golgi stack.

The transition zone (TZ) at the ciliary base has emerged as an important regulator of the composition and functions of cilia, which are microtubule-based structures extending from the surfaces of most eukaryotic cells, serving motility, chemo-/mechano-/photosensation and developmental signaling roles. Possessing distinct ultrastructural features such as microtubule-membrane spanning Y-links, the ∼0.2-1.0-μm long TZ is thought to act as a gated cytosolic (size dependent) and membrane diffusion barrier that drives ciliary compartmentalization by preventing unregulated protein exchange between the cilium and the rest of the cell. Multiple proteins associated with ciliary diseases (ciliopathies) such as Meckel-Gruber syndrome (MKS) and nephronophthisis are specifically found in the TZ, and work from a number of model systems, including Chlamydomonas reinharditii, Caenorhabditis elegans and the mouse indicates TZ-gating and associated ciliogenic functions for a number of these proteins. Here we present a suite of assays for probing the structure, function, and molecular composition of the C. elegans TZ, with emphasis on TZ ultrastructure, diffusion barrier kinetics, MKS module assembly hierarchy, and TZ-dependent behaviors., (Copyright © 2015 Elsevier Inc. All rights reserved.)

The primary cilium has emerged as a hotbed of sensory and developmental signaling, serving as a privileged domain to concentrate the functions of a wide number of channels, receptors and downstream signal transducers. This realization has provided important insight into the pathophysiological mechanisms underlying the ciliopathies, an ever expanding spectrum of multi-symptomatic disorders affecting the development and maintenance of multiple tissues and organs. One emerging research focus is the subcompartmentalised nature of the organelle, consisting of discrete structural and functional subdomains such as the periciliary membrane/basal body compartment, the transition zone, the Inv compartment and the distal segment/ciliary tip region. Numerous ciliopathy, transport-related and signaling molecules localize at these compartments, indicating specific roles at these subciliary sites. Here, by focusing predominantly on research from the genetically tractable nematode C. elegans, we review ciliary subcompartments in terms of their structure, function, composition, biogenesis and relationship to human disease.

Cilia are microtubule-based cell appendages, serving motility, chemo-/mechano-/photo- sensation, and developmental signaling functions. Cilia are comprised of distinct structural and functional subregions including the basal body, transition zone (TZ) and inversin (Inv) compartments, and defects in this organelle are associated with an expanding spectrum of inherited disorders including Bardet-Biedl syndrome (BBS), Meckel-Gruber Syndrome (MKS), Joubert Syndrome (JS) and Nephronophthisis (NPHP). Despite major advances in understanding ciliary trafficking pathways such as intraflagellar transport (IFT), how proteins are transported to subciliary membranes remains poorly understood. Using Caenorhabditis elegans and mammalian cells, we investigated the transport mechanisms underlying compartmentalization of JS-associated ARL13B/ARL-13, which we previously found is restricted at proximal ciliary membranes. We now show evolutionary conservation of ARL13B/ARL-13 localisation to an Inv-like subciliary membrane compartment, excluding the TZ, in many C. elegans ciliated neurons and in a subset of mammalian ciliary subtypes. Compartmentalisation of C. elegans ARL-13 requires a C-terminal RVVP motif and membrane anchoring to prevent distal cilium and nuclear targeting, respectively. Quantitative imaging in more than 20 mutants revealed differential contributions for IFT and ciliopathy modules in defining the ARL-13 compartment; IFT-A/B, IFT-dynein and BBS genes prevent ARL-13 accumulation at periciliary membranes, whereas MKS/NPHP modules additionally inhibit ARL-13 association with TZ membranes. Furthermore, in vivo FRAP analyses revealed distinct roles for IFT and MKS/NPHP genes in regulating a TZ barrier to ARL-13 diffusion, and intraciliary ARL-13 diffusion. Finally, C. elegans ARL-13 undergoes IFT-like motility and quantitative protein complex analysis of human ARL13B identified functional associations with IFT-B complexes, mapped to IFT46 and IFT74 interactions. Together, these findings reveal distinct requirements for sequence motifs, IFT and ciliopathy modules in defining an ARL-13 subciliary membrane compartment. We conclude that MKS/NPHP modules comprise a TZ barrier to ARL-13 diffusion, whereas IFT genes predominantly facilitate ARL-13 ciliary entry and/or retention via active transport mechanisms., Competing Interests: The authors have declared that no competing interests exist.

Background: Multiple intracellular transport pathways drive the formation, maintenance, and function of cilia, a compartmentalized organelle associated with motility, chemo-/mechano-/photosensation, and developmental signaling. These pathways include cilium-based intraflagellar transport (IFT) and poorly understood membrane trafficking events. Defects in ciliary transport contribute to the etiology of human ciliary disease such as Bardet-Biedl syndrome (BBS). In this study, we employ the genetically tractable nematode Caenorhabditis elegans to investigate whether endocytosis genes function in cilium formation and/or the transport of ciliary membrane or ciliary proteins., Results: Here we show that localization of the clathrin light chain, AP-2 clathrin adaptor, dynamin, and RAB-5 endocytic proteins overlaps with a morphologically discrete periciliary membrane compartment associated with sensory cilia. In addition, ciliary transmembrane proteins such as G protein-coupled receptors concentrate at periciliary membranes. Disruption of endocytic gene function causes expansion of ciliary and/or periciliary membranes as well as defects in the ciliary targeting and/or transport dynamics of ciliary transmembrane and IFT proteins. Finally, genetic analyses reveal that the ciliary membrane expansions in dynamin and AP-2 mutants require bbs-8 and rab-8 function and that sensory signaling and endocytic genes may function in a common pathway to regulate ciliary membrane volume., Conclusions: These data implicate C. elegans endocytosis proteins localized at the ciliary base in regulating ciliary and periciliary membrane volume and suggest that membrane retrieval from these compartments is counterbalanced by BBS-8 and RAB-8-mediated membrane delivery., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Genomic rearrangements involving genes encoding erythroblast transformation-specific transcription factors are commonly present in prostate cancer. The TMPRSS2-ERG gene fusion that leads to ERG overexpression occurs in ~70% of prostate cancers. Implementation of fusion gene detection in pathological practice, however, has been hampered by the lack of reliable ERG antibodies. The objective of this study was first to compare ERG immunohistochemistry using the recently described antibody EPR3864 with ERG mRNA by quantitative PCR and, second, to investigate ERG immunohistochemistry in diagnostic prostate cancer needle biopsies. We analyzed 41 primary prostate adenocarcinomas obtained by radical prostatectomy and 83 consecutive prostate cancer needle biopsies. In the prostatectomy specimens, immunohistochemical ERG expression was highly concordant with the ERG mRNA overexpression (sensitivity 100% and specificity 85%). ERG overexpression was due to TMPRSS2-ERG gene fusion in all cases. ERG protein expression was identified in 51/83 adenocarcinomas (61%) on needle biopsies. ERG expression was more frequent in tumors infiltrating ≥2 needle biopsies (P<0.001) or occupying ≥50% of a single biopsy (P=0.018). Expression of ERG also occurred in 11/21 (52%) high-grade prostate intraepithelial neoplasia lesions. In 5/87 (6%) needle biopsies containing benign secretory glands, weak ERG staining was focally observed. In all of these cases, respective glands were adjacent to adenocarcinomas. In conclusion, immunohistochemistry for ERG strongly correlated with ERG mRNA overexpression and was specific for prostate cancer on needle biopsies. Therefore, ERG immunohistochemistry is an important adjunctive tool for pathophysiological studies on ERG gene fusions, and might support the pathological diagnosis of adenocarcinoma in a subset of prostate needle biopsies.

Conventional equine influenza vaccination schedules consist of a primary course of two vaccinations given 4-6 weeks apart followed by a third vaccination (booster) given approximately 5 months later. In between the primary course and the third vaccination, horses are generally considered not to be adequately protected against influenza. This study aimed to investigate whether Thoroughbred foals would benefit from a vaccination schedule in which the third vaccination was given earlier than in conventional vaccination schedules. The vaccines used were an inactivated whole virus equine influenza vaccine and an inactivated whole virus combination vaccine containing equine influenza and equine herpesvirus antigens. Four groups of foals were vaccinated with the two vaccines according to a conventional and an accelerated vaccination schedule in which the third vaccination was given 14 weeks after the first administration. In both groups, the fourth vaccination was given at the normally recommended interval of 26 weeks after the third vaccination for the combination vaccine and 52 weeks after the third vaccination with the influenza only vaccine. The horses were 4-11 months of age and seronegative for influenza. Immunological responses after vaccination were monitored for several months using the single radial haemolysis test. The results indicated that 28 weeks after the first vaccination, antibody levels in horses vaccinated according to the accelerated schedule were not significantly higher than in horses vaccinated according to the conventional schedule. In addition, the total level of antibody production (area under the curve) was not significantly different at that point although antibody titres were slightly higher (but not significantly so) between 16-30 weeks in the accelerated schedule. Between the third and fourth doses, horses vaccinated according to the accelerated schedule had antibodies against influenza below the level required for clinical protection for 39 and 18 weeks for the influenza only and the combination vaccine, respectively, whereas those vaccinated according to the conventional schedule had antibody titres below the level for clinical protection for 9-15 weeks in the corresponding period for both vaccines. Horses vaccinated according to the accelerated schedule with the combination vaccine had lower antibody titres after the fourth vaccination than those vaccinated according to the conventional schedule after the third vaccination, although antibody titres prior to vaccination were similar. For the influenza only vaccine, titres after the accelerated fourth administration were not different to those after the conventional third vaccination. There was no benefit from early booster vaccinations with the vaccines used in this study, so for these vaccines the conventional schedule provided better protection than the selected accelerated alternative. This may contrast with some other vaccine formulations, although a direct comparison using similar protocols has not been made.

Objective: To investigate the clinical and electrophysiologic phenotype of Charcot-Marie-Tooth disease (CMT) Type 2 in a large number of affected families., Methods: We excluded CMT Type 1, hereditary neuropathy with liability to pressure palsies, and CMT due to Cx32 gene mutations by DNA analysis. We performed genetic analysis of the presently known CMT Type 2 genes., Results: Sixty-one persons from 18 families were affected. Ninety percent of patients were able to walk with or without the help of aids. Proximal leg muscle weakness was present in 13%. Asymmetrical features were present in 15%. Normal or brisk knee reflexes were present in 36%. Extensor plantar responses without associated spasticity occurred in 10 patients from eight families. Only three causative mutations were identified in the MFN2, BSCL2, and RAB7 genes. No mutations were found in the NEFL, HSPB1, HSPB8, GARS, DNM2, and GDAP1 genes., Conclusions: At group level, the clinical phenotype of Charcot-Marie-Tooth disease (CMT) Type 2 is uniform, with symmetric, distal weakness, atrophy and sensory disturbances, more pronounced in the legs than in the arms, notwithstanding the genetic heterogeneity. Brisk reflexes, extensor plantar responses, and asymmetrical muscle involvement can be considered part of the CMT Type 2 phenotype. The causative gene mutation was found in only 17% of the families we studied.

In 1996, metaidoioplasty was introduced as an alternative to phalloplasty in female-to-male transsexuals. To assess the long-term outcome in 70 consecutive patients (mean follow-up 8 years), we established the characteristics of postoperative events and additional surgical procedures. Metaidoioplasty and primary or secondary scrotoplasty was uneventful in 8 patients. In the other patients, postoperative events included immediate postoperative complications (n = 23), urethral fistulas (n = 26) or strictures (n = 25), or loss (n = 22) or dislocation (n = 34) of testicular prostheses. An average of 2.6 surgical procedures per patient was needed to complete genital confirmation and cope with all events. Additional phalloplasty was performed or scheduled in 17 patients. We conclude that genital reassignment by metaidoioplasty cannot usually be completed in 1 step and that phalloplasty is feasible subsequent to metaidoioplasty. We still consider metaidoioplasty to be a method of choice in selected patients.

In our hands, neourethral stenosis is the main complication following metaidoioplasty in female-to-male transsexuals. We introduce the use of surplus of minor labial skin to correct these stenoses. The surplus was used as a subcutaneously pedicled flap with a 1.5 x 3.5 cm skin paddle to correct the circumferential deficit of neourethral lining at the level of the stenosis. After minimum undermining, the pedicle was retracted laterally to allow for a median external urethrotomy. The skin paddle was turned outside in to fit the resulting longitudinal neourethral defect. Subsequently, the major labial subcutis and skin were approximated in layers to cover the subcutaneous pedicled flap and to close the labioscrotum in the midline. Patients were kept immobilized for 3 days, and a suprapubic catheter was left open for 7 days. This technique was applied successfully in 15 of the 70 female-to-male transsexuals who consequently underwent metaidoioplasty in Amsterdam up to March of 1999. We conclude that the surplus of labial skin ought to be retained during primary surgery because it is an ideal substitute to correct neourethral stenosis.

A late onset axonal Charcot-Marie-Tooth phenotype is described, resulting from a novel mutation in the myelin protein zero (MPZ) gene. Comparative computer modelling of the three dimensional structure of the MPZ protein predicts that this mutation does not cause a significant structural change. The primary axonal disease process in these patients points to a function of MPZ in maintenance of the myelinated axons, apart from securing stability of the myelin layer.

Double-breasted closure of the skin is achieved by deepithelialization of 1 edge of a wound and advancement of the opposite edge over this deepithelialized area. Such closure may prevent contamination or exposure of the implant in cases where immediate breast reconstruction with prosthetic material is obtained after skin sparing mastectomy. The deepithelialization routinely extends as far as the incision to be closed and, consequently, the skin suture at the end of the incision immediately overlies thedeep suture or implant in cases where the implant is not fully covered by muscle. To prevent contamination or exposure of the implant when suture infection or wound dehiscence occurs in these cases, we extend the deepithelialization beyond and around the limit of the skin incision. This allows for complete and secured double-breasted closure of the entire wound and lessens the risk of implant contamination or exposure.

Objective: To characterize and distinguish the types of sensorineural hearing impairment (SNHI) that occur in hereditary motor and sensory neuropathy Type 1a (HMSN-1a) and hereditary neuropathy with liability to pressure palsies (HNPP), which are caused by deletion or frameshift mutation., Study Design: Prospective study., Setting: Ambulatory patients in a university hospital., Patients: Twelve patients with HMSN-1a due to a duplication of the PMP22 gene on chromosome 17p11.2, 16 patients with HNPP due to the common PMP22 deletion (HNPP del), and 11 HNPP patients with a frame shift mutation (heterozygous PMP22 G-insertion) (HNPP mut), all confirmed by molecular genetic analysis., Interventions: Pure-tone audiograms and speech audiograms were obtained., Main Outcome Measures: Results of cross-sectional analysis comprising linear regression of hearing threshold on age., Results: Pure-tone audiograms showed mild to moderate SNHI, predominant at the low and the high frequencies. SNHI showed significant progression by approximately 0.4 dB per year at 0.25 to 4 kHz and up to 1 to 2 dB per year at 4 to 8 kHz. Patients with HMSN-1a had substantial, presumably congenital, SNHI but did not show significant progression beyond presbyacusis. Patients with HNPP showed postnatal onset at age 11 years with progression of SNHI in excess of presbyacusis by 0.4 dB per year. All three types of neuropathy showed normal speech recognition., Conclusions: All three types of neuropathy showed SNHI with normal speech recognition. HMSN-1a showed stable SNHI without progression beyond presbyacusis. HNPP showed progression beyond presbyacusis with postnatal onset. The differences in SNHI may be explained by the differences in PMP22 expression. The progressive SNHI in HNPP might be explained by the liability for exogenous factors associated with this disorder.

We report the findings in five muscle and three sural nerve biopsies, and in one postmortem plexus specimen, from six patients with hereditary neuralgic amyotrophy (HNA). We found that the sensory nerves are definitely involved in HNA despite the mainly motor symptoms, and that lesions in nerves and muscles are more widespread throughout the peripheral nervous system than clinically presumed, but, simultaneously, very focally affect isolated fascicles within individual nerves.

It has been recommended that modern equine influenza vaccines should contain an A/equi-1 strain and A/equi-2 strains of the American and European-like subtype. We describe here the efficacy of a modern updated inactivated equine influenza-herpesvirus combination vaccine against challenge with a recent American-like isolate of equine influenza (A/equine-2/Kentucky/95 (H3N8). The vaccine contains inactivated Influenza strains A-equine-1/Prague'56, A-equine-2/Newmarket-1/'93 (American lineage) and A-equine-2/ Newmarket-2/93 (Eurasian lineage) and inactivated EHV-1 strain RacH and EHV-4 strain V2252. It is adjuvanted with alhydrogel and an immunostim. Horses were vaccinated at the start of the study and 4 weeks later. Four, six and eight weeks after the first vaccination high anti-influenza antibody titres were found in vaccinated horses, whereas at the start of the study all horses were seronegative. After the challenge, carried out at 8 weeks after the first vaccination, nasal swabs were taken, rectal temperatures were measured and clinical signs were monitored for 14 days. In contrast to unvaccinated control horses, vaccinated animals shed hardly any virus after challenge, and the appearance of clinical signs of influenza such as nasal discharge, coughing and fever were reduced in the vaccinated animals. Based on these observations, it was concluded that the vaccine protected against clinical signs of influenza and, more importantly, against virus excretion induced by an American-like challenge virus strain. In a second experiment the duration of the immunity induced by this vaccine was assessed serologically. Horses were vaccinated at the start of the study and 6 and 32 weeks later. Anti-influenza antibody titres were determined in bloodsamples taken at the first vaccination, and 2, 6, 8, 14, 19, 28, 32, 37, 41, 45 and 58 weeks after the first vaccination. Vaccinated horses had high anti-influenza antibody titres, above the level for clinical protection against influenza, against all strains present in the vaccine until 26 weeks after the third vaccination.

Available IBDV vaccines based on classical IBDV strains do not induce full protection in the presence of maternally derived antibodies (MDA) against variant strains. The aim of this study was the generation of an IBDV chimera between both classical and variant strains. Characterization of the generated chimeric IBDV (D78-Chim) was performed in both cell culture and susceptible chickens using the classical vaccine strain D78 for comparison. Growth kinetics on cell culture showed that the D78-Chim replicates comparable to the live vaccine strain D78. Administration of the D78-Chim to SPF chickens resulted in equally high virus neutralizing antibodies titres against both, classical and variant IBDV strains. In vivo experiments in commercial chickens revealed that D78-Chim and D78 induced comparable bursal damage in presence of maternally derived antibodies. Both D78-Chim and D78 viruses were able to breakthrough a similar level of MDA.

The presence of maternal immunity in broilers does not preclude the successful protective immunization with an attenuated live reovirus vaccine at 1-day-old. This was recently demonstrated in a reovirus challenge model based on reovirus isolation from different organs (van Loon et al., 2002). Discrimination between challenge and vaccine virus was made by specific monoclonal antibody. Notably, at the day of challenge the levels of circulating reovirus-specific antibodies proved to be minimal or undetectable. This suggested an antibody-independent vaccination-induced immunity under the chosen conditions. In the present study we assessed the contribution of specific humoral responses in the same experimental model, by evaluating the control of a virulent challenge infection in the complete absence of B cells as a result of cyclophosphamide treatment. These experiments were performed in both SPF white leghorn chickens and commercially available broilers with passively acquired maternal antibodies. Our data demonstrate that the virus is controlled in the absence of actively produced antibodies, and is independent of B lymphocytes. This suggests that cellular immunity is sufficient for protection of broilers with maternal antibodies against reovirus infection following early age vaccination with live reoviral vaccine.

Background: Mutations in the ganglioside-induced differentiation-associated protein 1 gene (GDAP1) were recently shown to be responsible for autosomal recessive (AR) demyelinating Charcot-Marie-Tooth disease (CMT) type 4A (CMT4A) as well as AR axonal CMT with vocal cord paralysis., Methods: The coding region of GDAP1 was screened for the presence of mutations in seven families with AR CMT in which the patients were homozygous for markers of the CMT4A locus at chromosome 8q21.1., Results: A nonsense mutation was detected in exon 5 (c.581C>G, S194X), a 1-bp deletion in exon 6 (c.786delG, G262fsX284), and a missense mutation in exon 6 (c.844C>T, R282C)., Conclusions: Mutations in GDAP1 are a frequent cause of AR CMT. They result in an early-onset, severe clinical phenotype. The range of nerve conduction velocities (NCV) is variable. Some patients have normal or near normal NCV, suggesting an axonal neuropathy, whereas others have severely slowed NCV compatible with demyelination. The peripheral nerve biopsy findings are equally variable and show features of demyelination and axonal degeneration.

Hereditary neuropathy with liability to pressure palsies is associated with a deficiency in the Peripheral Myelin Protein 22 (PMP22). Most hereditary neuropathy with liability to pressure palsies cases are caused by a deletion of a 1.5 Mb region on chromosome 17p11.2-12 encompassing the PMP22 gene. We describe a hereditary neuropathy with liability to pressure palsies family that lacks the common deletion, but carries a small deletion spanning the 3' region of the PMP22 gene, causing only a partial deletion of one copy of the gene.

Purpose of the Review: Secondary axonal atrophy is common in most if not all demyelinating neuropathies and is likely responsible for the majority of clinical symptoms. We review clinical, electrophysiological and morphological evidence for secondary axonal atrophy in demyelinating neuropathies and summarize recent hypotheses on possible pathomechanisms., Recent Findings: Elucidation of genetic defects responsible for hereditary demyelinating neuropathies and insights into axon-Schwann cell interactions have allowed longitudinal studies of genetically defined demyelinating neuropathies and research into the pathomechanism of secondary axonal atrophy., Summary: There is ample clinical electrophysiological and electropathological evidence that secondary axonal atrophy is found in hereditary and demyelinating neuropathies. Recognizing secondary axonal atrophy as a main cause for clinical disability in demyelinating neuropathies is important for the clinician and may reveal a therapeutic target common to all different forms of demyelinating neuropathies.

Charcot-Marie-Tooth disease caused by mutations of the myelin protein zero gene demonstrates considerable phenotypical variability. We describe a 45-year-old female with a peripheral neuropathy with demyelinating and axonal features, pes cavus and pupillary light-near dissociation. She was heterozygous for two mutations in the myelin protein zero gene (His81Tyr and Val113Phe), both present on the same allele. Our patient shows a less severe phenotype than previously described patients with a His81Arg mutation. Multiple mutations in the myelin protein zero gene, as well as Charcot-Marie-Tooth with pupillary abnormalities have previously been described in rare instances. However, concurrent occurrence of both phenomena is a novel finding.

The efficacy of live reovirus vaccines may be determined by challenge via the foot pad route 3 to 4 weeks after vaccination. Swelling and discoloration in the foot pad and shank are scored for a period of 14 days. The major disadvantages of this challenge model are the subjective judgement of gross foot pad and/or shank lesions, that it is very difficult to induce lesions in broilers, and that it causes animal suffering. Other reovirus challenge models are based on reisolation of the virus from different tissues or on scoring microscopic lesions in the tendons. Some disadvantages of these models are that they either cannot be used after vaccination with live reovirus because they cannot discriminate between vaccine and challenge virus or that the microscopic lesions scored need not necessarily be related to the challenge virus but may have been induced by other factors. Therefore, we have attempted to develop a reovirus challenge model that was an improvement on the existing ones, using isolation of reovirus from different organs followed by specific detection of the challenge virus with a monoclonal antibody that can discriminate between challenge and vaccine virus. The reovirus challenge model was examined in specific pathogen free (SPF) White Leghorn chickens and commercial broilers. In vivo studies were conducted to examine the efficacy of an attenuated reovirus vaccine in SPF White Leghorn chickens and commercial broilers with maternal immunity against reovirus. No challenge virus could be detected in any of the organs of the vaccinated chickens 3 and 10 days after challenge. In contrast, challenge virus could be isolated from the unvaccinated control group. At an increased challenge dose all unvaccinated challenge control birds were positive, while the vaccinated chickens were protected. It was shown that 1-day-old vaccination in the presence of maternal immunity was effective. It seemed that protection induced in broilers by the attenuated reovirus vaccine may not have been entirely humoral because in protected birds no antibodies against reovirus were detected by enzyme-linked immunosorbent at the time of challenge. Protection in these birds might therefore have been induced by cellular immunity.