Nogo receptor homolog NgR2 expressed in sensory DRG neurons controls epidermal innervation by interaction with Versican.

Primary sensory afferents of the dorsal root ganglion (DRG) that innervate the skin detect a wide range of stimuli, such as touch, temperature, pain, and itch. Different functional classes of nociceptors project their axons to distinct target zones within the developing skin, but the molecular mechanisms that regulate target innervation are less clear. Here we report that the Nogo66 receptor homolog NgR2 is essential for proper cutaneous innervation. NgR2(-/-) mice display increased density of nonpeptidergic nociceptors in the footpad and exhibit enhanced sensitivity to mechanical force and innocuous cold temperatures. These sensory deficits are not associated with any abnormality in morphology or density of DRG neurons. However, deletion of NgR2 renders nociceptive nonpeptidergic sensory neurons insensitive to the outgrowth repulsive activity of skin-derived Versican. Biochemical evidence shows that NgR2 specifically interacts with the G3 domain of Versican. The data suggest that Versican/NgR2 signaling at the dermo-epidermal junction acts in vivo as a local suppressor of axonal plasticity to control proper density of epidermal sensory fiber innervation. Our findings not only reveal the existence of a novel and unsuspected mechanism regulating epidermal target innervation, but also provide the first evidence for a physiological role of NgR2 in the peripheral nervous system.

Determinants of versican-V1 proteoglycan processing by the metalloproteinase ADAMTS5.

Proteolysis of the Glu(441)-Ala(442) bond in the glycosaminoglycan (GAG) ß domain of the versican-V1 variant by a disintegrin-like and metalloproteinase domain with thrombospondin type 1 motif (ADAMTS) proteases is required for proper embryo morphogenesis. However, the processing mechanism and the possibility of additional ADAMTS-cleaved processing sites are unknown. We demonstrate here that if Glu(441) is mutated, ADAMTS5 cleaves inefficiently at a proximate upstream site but normally does not cleave elsewhere within the GAGß domain. Chondroitin sulfate (CS) modification of versican is a prerequisite for cleavage at the Glu(441)-Ala(442) site, as demonstrated by reduced processing of CS-deficient or chondroitinase ABC-treated versican-V1. Site-directed mutagenesis identified the N-terminal CS attachment sites Ser(507) and Ser(525) as essential for processing of the Glu(441)-Ala(442) bond by ADAMTS5. A construct including only these two GAG chains, but not downstream GAG attachment sites, was cleaved efficiently. Therefore, CS chain attachment to Ser(507) and Ser(525) is necessary and sufficient for versican proteolysis by ADAMTS5. Mutagenesis of Glu(441) and an antibody to a peptide spanning Thr(432)-Gly(445) (i.e. containing the scissile bond) reduced versican-V1 processing. ADAMTS5 lacking the C-terminal ancillary domain did not cleave versican, and an ADAMTS5 ancillary domain construct bound versican-V1 via the CS chains. We conclude that docking of ADAMTS5 with two N-terminal GAG chains of versican-V1 via its ancillary domain is required for versican processing at Glu(441)-Ala(442). V1 proteolysis by ADAMTS1 demonstrated a similar requirement for the N-terminal GAG chains and Glu(441). Therefore, versican cleavage can be inhibited substantially by mutation of Glu(441), Ser(507), and Ser(525) or by an antibody to the region of the scissile bond.

Desmoplastic small round cell tumor: a rare cause of a progressive brachial plexopathy.

INTRODUCTION: Desmoplastic small round cell tumor (DSRCT) is an uncommon, embryonic-type neoplasm, typically presenting as an abdominal mass in young men. A single case of DSRCT arising in the peripheral nervous system has been reported previously. METHODS: The clinical course, imaging, electrophysiological, intraoperative, histopathological, molecular findings, and postoperative follow-up are reported. RESULTS: A 43-year-old man presented with slowly progressive right brachial plexopathy. Magnetic resonance imaging revealed an enlarged medial cord with heterogeneous contrast enhancement. Histology showed a "small round cell" neoplasm with a polyphenotypic immunoprofile, including epithelial and mesenchymal markers. A pathognomonic fusion of Ewing sarcoma breakpoint region 1 and Wilms tumor 1 genes (EWSR1/WT1) was present. Treatment involved gross total excision and local radiotherapy. CONCLUSIONS: Our findings confirm the occurrence of DSRCT as a primary peripheral nerve tumor. Despite its usually very aggressive clinical course, prolonged recurrence-free survival may be reached. Histomorphology and immunoprofile of DSRCT may lead to misdiagnosis as small cell carcinoma.

Primary cutaneous follicle center lymphoma with diffuse CD30 expression: a report of 4 cases of a rare variant.

BACKGROUND: CD30 is expressed in aggressive and Epstein-Barr virus-associated forms of B-cell non-Hodgkin lymphomas, but is rarely expressed by the majority of tumor cells in primary cutaneous B-cell lymphomas (CBCLs). The expression of CD30 in CBCLs may be at risk for misinterpretation as an unequivocal indicator of a highly aggressive form of the disease. OBJECTIVE: We report 4 cases of low malignant primary cutaneous follicle center lymphoma (PCFCL) with diffuse and strong expression of CD30 by the majority of neoplastic cells. RESULTS: The patients included 3 men and 1 woman with tumors on the scalp (3 patients) and chest wall (1 patient). The histologic examinations revealed a mixed, diffuse, and follicular growth pattern with CD20(+), bcl-6(+), and bcl-2(-) tumor cells. Seventy percent to 90% of the tumor cells expressed CD30. Clonal rearrangement of immunoglobulin heavy chain genes was found in 1 of 4 cases. None of the 3 cases yielded positivity for Epstein-Barr virus RNA. LIMITATIONS: The study is limited by the small number of patients. CONCLUSIONS: This rare variant of CD30(+) PCFCL needs be distinguished from CD30(+) aggressive B-cell lymphomas. CD30 in this variant of CBCLs may serve as a therapeutic target for anti-CD30 antibody-based strategies.

Primary cutaneous marginal zone lymphoma in children: a report of 3 cases and review of the literature.

: Primary cutaneous marginal zone lymphoma (PCMZL) is one of the most common cutaneous B-cell lymphomas. It affects mostly patients in their fourth decade and manifests with multifocal nodules mostly on the arms and upper trunk in more than half of the patients. PCMZL is, however, rare in children and adolescents, with only 20 cases reported in patients aged 20 and younger. The authors present 3 cases of PCMZL in teenagers. The patients were 2 girls aged 18 and 13 and a 17-year-old boy. Two patients presented with multiple lesions involving various anatomic sites, whereas in 1 patient, 2 small closely opposed papules on the abdomen were seen. Histopathologically, the characteristic appearance of PCMZL was found in 3 of 4 specimens, with nodular infiltrates composed of small lymphocytes in the interfollicular compartment, reactive germinal centers, and plasma cells in small clusters mainly at the periphery of the infiltrates, whereas 1 specimen showed a dense lymphocytic infiltrate with small granulomas. Clonality was demonstrated by monotypic immunoglobulin light chain expression and/or monoclonal rearrangement of the immunoglobulin heavy chain genes. No Borrelia burgdorferi was identified on serology or by polymerase chain reaction in any of the cases. Treatment included excision or administration of antibiotics with complete remission in all the 3 patients indicating that PCMZL in children and young adolescents follows the same indolent course with a tendency for recurrences, but excellent prognosis as in adults. The pertinent literature on PCZL in childhood and adolescence is reviewed.

Prosthetic valve endocarditis and bloodstream infection due to Mycobacterium chimaera.

Prosthetic valve endocarditis (PVE) due to fast-growing nontuberculous mycobacteria (NTM) has been reported anecdotally. Reports of PVE with slowly growing NTM, however, are lacking. We present here one case of PVE and one case of bloodstream infection caused by Mycobacterium chimaera. Randomly amplified polymorphic DNA (RAPD)-PCR indicated a relatedness of the two M. chimaera strains. Both patients had heart surgery 2 years apart from each other. A nosocomial link was not detected.

Spindle-cell variant of primary cutaneous follicle center lymphoma spreading to the hepatobiliary tree, mimicking Klatskin tumor.

Primary cutaneous follicle center lymphoma (pcFCL) is an indolent type of primary cutaneous B-cell lymphoma (pcBCL) rarely disseminating to other organs. PcBCL with spindle-cell morphology has been described as a rare variant of pcFCL but the prognosis data of this variant is sparse. We report a rare case of spindle-cell pcFCL with CD20(+), CD79a(+), CD3(+), Bcl-6(+), Mum-1(-) and CD10(-) tumor cells that infiltrated the hepatic hilum, mimicking a Klatskin tumor. On the basis of the sparse published data on spindle-cell morphology of pcBCL, this growth pattern should elicit awareness of an increased risk of systemic involvement in the otherwise indolent pcFCL.

Versican V0 and V1 direct the growth of peripheral axons in the developing chick hindlimb.

Peanut agglutinin-binding disaccharides and chondroitin sulfate mark transient mesenchymal barriers to advancing motor and sensory axons innervating the hindlimbs during chick development. Here we show that the vast majority of these carbohydrates are at the critical stage and location attached to the versican splice variants V0 and V1. We reveal that the isolated isoforms of this extracellular matrix proteoglycan suppress axon extension at low concentrations and induce growth cone collapse and rapid retraction at higher levels. Moreover, we demonstrate that versican V0 and/or V1, recombinantly expressed in collagen-I gels or ectopically deposited in the hindlimbs of chicken embryos in ovo, cause untimely defasciculation and axon stalling. Consequently, severe disturbances of nerve patterning are observed in the versican-treated embryos. Our experiments emphasize the inhibitory capacity of versicans V0 and V1 in axonal growth and evidence for their function as basic guidance cues during development of the peripheral nervous system.

A natural freshwater origin for two chlamydial species, Candidatus Piscichlamydia salmonis and Candidatus Clavochlamydia salmonicola, causing mixed infections in wild brown trout (Salmo trutta).

Gill disease in salmonids is characterized by a multifactorial aetiology. Epitheliocystis of the gill lamellae caused by obligate intracellular bacteria of the order Chlamydiales is one known factor; however, their diversity has greatly complicated analyses to establish a causal relationship. In addition, tracing infections to a potential environmental source is currently impossible. In this study, we address these questions by investigating a wild brown trout (Salmo trutta) population from seven different sites within a Swiss river system. One age class of fish was followed over 18 months. Epitheliocystis occurred in a site-specific pattern, associated with peak water temperatures during summer months. No evidence of a persistent infection was found within the brown trout population, implying an as yet unknown environmental source. For the first time, we detected 'Candidatus Piscichlamydia salmonis' and 'Candidatus Clavochlamydia salmonicola' infections in the same salmonid population, including dual infections within the same fish. These organisms are strongly implicated in gill disease of caged Atlantic salmon in Norway and Ireland. The absence of aquaculture production within this river system and the distance from the sea, suggests a freshwater origin for both these bacteria and offers new possibilities to explore their ecology free from aquaculture influences.

Coreplication of the major genotype group members of porcine circovirus type 2 as a prerequisite to coevolution may explain the variable disease manifestations.

A member of the family Circoviridae, porcine circovirus type 2 (PCV2), is associated with postweaning multisystemic wasting syndrome (PMWS), a recent emerging disease worldwide. PCV2 is also found in clinically asymptomatic animals. This paradoxical finding makes the syndrome etiology challenging. We developed new assays to study PCV2 with links to syndrome etiology. For analysis, we used PCV2-infected tissues from subclinically infected and diseased piglets. We compared antigen- and PCV2 DNA-derived signals for tissue localization and intensity. Oligonucleotides were designed to the signature motif of the PCV2 capsid open reading frame to discriminate experimentally between PCV2 genotype groups by PCR, in situ hybridization (ISH), and fluorescence in situ hybridization (FISH). Unexpectedly, all PCV2-infected animals carried both PCV2a and PCV2b genotype group members. Using confocal microscopy, genotype single-cell infections and cell superinfections were visible. Additionally, we discriminated replicative DNA from total PCV2 DNA isoforms with FISH. This aided in our inquiry into cellular genotype-specific replication. Importantly, single-genotype-group replication was not observed. In infected cells with replicating virus, both genotype groups were equally present. These findings suggest PCV2 genotype group members relaxed replication regulation requirements and may even point to PCV2 replication cooperativity in vivo. These observations explain the readily seen PCV2 DNA recombinations and the high overall PCV2 genome plasticity. Hence, we suggest a novel mechanism of syndrome etiology that consists of a continuously changing PCV2 genome pool in hosts and pig herds, posing a constant challenge to the individual maturing immune system.

Oligodendrocyte myelin glycoprotein does not influence node of ranvier structure or assembly.

Oligodendrocyte myelin glycoprotein (OMgp) is expressed by both neurons and oligodendrocytes in the CNS. It has been implicated in growth cone collapse and neurite outgrowth inhibition by signaling through the Nogo receptor and paired Ig-like receptor B (PirB). OMgp was also reported to be an extracellular matrix (ECM) protein surrounding CNS nodes of Ranvier and proposed to function as (1) an inhibitor of nodal collateral sprouting and (2) an important contributor to proper nodal and paranodal architecture. However, we show here that the anti-OMgp antiserum used in previous studies to define the functions of OMgp at nodes is not specific. Among all reported nodal ECM components, the antiserum exhibited strong cross-reactivity against versican V2 isoform, a chondroitin sulfate proteoglycan. Furthermore, the OMgp antiserum labeled OMgp-null nodes, but not nodes from versican V2-deficient mice, and preadsorption of the OMgp antiserum with recombinant versican V2 blocked nodal labeling. Analysis of CNS nodes in OMgp-null mice failed to reveal any nodal or paranodal defects, or increased nodal collateral sprouting, indicating that OMgp does not participate in CNS node of Ranvier assembly or maintenance. We successfully identified a highly specific anti-OMgp antibody and observed OMgp staining in white matter only after initiation of myelination. OMgp immunoreactivity decorated the surface of mature myelinated axons, but was excluded from compact myelin and nodes. Together, our results strongly argue against the nodal localization of OMgp and its proposed functions at nodes, and reveal OMgp's authentic localization relative to nodes and myelin.

Age-related differences in human skin proteoglycans.

Previous work has shown that versican, decorin and a catabolic fragment of decorin, termed decorunt, are the most abundant proteoglycans in human skin. Further analysis of versican indicates that four major core protein species are present in human skin at all ages examined from fetal to adult. Two of these are identified as the V0 and V1 isoforms, with the latter predominating. The other two species are catabolic fragments of V0 and V1, which have the amino acid sequence DPEAAE as their carboxyl terminus. Although the core proteins of human skin versican show no major age-related differences, the glycosaminoglycans (GAGs) of adult skin versican are smaller in size and show differences in their sulfation pattern relative to those in fetal skin versican. In contrast to human skin versican, human skin decorin shows minimal age-related differences in its sulfation pattern, although, like versican, the GAGs of adult skin decorin are smaller than those of fetal skin decorin. Analysis of the catabolic fragments of decorin from adult skin reveals the presence of other fragments in addition to decorunt, although the core proteins of these additional decorin catabolic fragments have not been identified. Thus, versican and decorin of human skin show age-related differences, versican primarily in the size and the sulfation pattern of its GAGs and decorin in the size of its GAGs. The catabolic fragments of versican are detected at all ages examined, but appear to be in lower abundance in adult skin compared with fetal skin. In contrast, the catabolic fragments of decorin are present in adult skin, but are virtually absent from fetal skin. Taken together, these data suggest that there are age-related differences in the catabolism of proteoglycans in human skin. These age-related differences in proteoglycan patterns and catabolism may play a role in the age-related changes in the physical properties and injury response of human skin.

Neural crest-derived cells with stem cell features can be traced back to multiple lineages in the adult skin.

Given their accessibility, multipotent skin-derived cells might be useful for future cell replacement therapies. We describe the isolation of multipotent stem cell-like cells from the adult trunk skin of mice and humans that express the neural crest stem cell markers p75 and Sox10 and display extensive self-renewal capacity in sphere cultures. To determine the origin of these cells, we genetically mapped the fate of neural crest cells in face and trunk skin of mouse. In whisker follicles of the face, many mesenchymal structures are neural crest derived and appear to contain cells with sphere-forming potential. In the trunk skin, however, sphere-forming neural crest-derived cells are restricted to the glial and melanocyte lineages. Thus, self-renewing cells in the adult skin can be obtained from several neural crest derivatives, and these are of distinct nature in face and trunk skin. These findings are relevant for the design of therapeutic strategies because the potential of stem and progenitor cells in vivo likely depends on their nature and origin.

A new mouse mutant with cleavage-resistant versican and isoform-specific versican mutants demonstrate that proteolysis at the Glu441-Ala442 peptide bond in the V1 isoform is essential for interdigital web regression.

Two inherent challenges in the mechanistic interpretation of protease-deficient phenotypes are defining the specific substrate cleavages whose reduction generates the phenotypes and determining whether the phenotypes result from loss of substrate function, substrate accumulation, or loss of a function(s) embodied in the substrate fragments. Hence, recapitulation of a protease-deficient phenotype by a cleavage-resistant substrate would stringently validate the importance of a proteolytic event and clarify the underlying mechanisms. Versican is a large proteoglycan required for development of the circulatory system and proper limb development, and is cleaved by ADAMTS proteases at the Glu441-Ala442 peptide bond located in its alternatively spliced GAGß domain. Specific ADAMTS protease mutants have impaired interdigit web regression leading to soft tissue syndactyly that is associated with reduced versican proteolysis. Versikine, the N-terminal proteolytic fragment generated by this cleavage, restores interdigit apoptosis in ADAMTS mutant webs. Here, we report a new mouse transgene, Vcan AA, with validated mutations in the GAGß domain that specifically abolish this proteolytic event. Vcan AA/AA mice have partially penetrant hindlimb soft tissue syndactyly. However, Adamts20 inactivation in Vcan AA/AA mice leads to fully penetrant, more severe syndactyly affecting all limbs, suggesting that ADAMTS20 cleavage of versican at other sites or of other substrates is an additional requirement for web regression. Indeed, immunostaining with a neoepitope antibody against a cleavage site in the versican GAGα domain demonstrated reduced staining in the absence of ADAMTS20. Significantly, mice with deletion of Vcan exon 8, encoding the GAGß domain, consistently developed soft tissue syndactyly, whereas mice unable to include exon 7, encoding the GAGα domain in Vcan transcripts, consistently had fully separated digits. These findings suggest that versican is cleaved within each GAG-bearing domain during web regression, and affirms that proteolysis in the GAGß domain, via generation of versikine, has an essential role in interdigital web regression.

Versican V2 assembles the extracellular matrix surrounding the nodes of ranvier in the CNS.

The CNS-restricted versican splice-variant V2 is a large chondroitin sulfate proteoglycan incorporated in the extracellular matrix surrounding myelinated fibers and particularly accumulating at nodes of Ranvier. In vitro, it is a potent inhibitor of axonal growth and therefore considered to participate in the reduction of structural plasticity connected to myelination. To study the role of versican V2 during postnatal development, we designed a novel isoform-specific gene inactivation approach circumventing early embryonic lethality of the complete knock-out and preventing compensation by the remaining versican splice variants. These mice are viable and fertile; however, they display major molecular alterations at the nodes of Ranvier. While the clustering of nodal sodium channels and paranodal structures appear in versican V2-deficient mice unaffected, the formation of the extracellular matrix surrounding the nodes is largely impaired. The conjoint loss of tenascin-R and phosphacan from the perinodal matrix provide strong evidence that versican V2, possibly controlled by a nodal receptor, organizes the extracellular matrix assembly in vivo.

AID protein expression in chronic lymphocytic leukemia/small lymphocytic lymphoma is associated with poor prognosis and complex genetic alterations.

The biological behavior of chronic lymphocytic leukemia and small lymphocytic lymphoma is unpredictable. Nonetheless, non-mutated IgV(H) gene rearrangement, ATM (11q22-23) and p53 (17p13) deletion are recognized as unfavorable prognosticators in chronic lymphocytic leukemia. The mRNA expression of activation-induced cytidine deaminase (AID), an enzyme indispensable for somatic hypermutation processes, was claimed to be predictive of non-mutated chronic lymphocytic leukemia cells in blood. Here, we evaluated AID protein expression compared with known molecular and immunohistochemical prognostic indicators in 71 chronic lymphocytic leukemia/small lymphocytic lymphoma patients using a tissue microarray approach. We found AID heterogeneously expressed in tumor cells as shown by colocalization analysis for CD5 and CD23. Ki-67 positive paraimmunoblasts of the proliferation centers displayed the highest expression. This observation is reflected by a significant association of AID positivity with a high proliferation rate (P=0.012). ATM deletion was detected in 10% (6/63) of patients and p53 deletion in 19% (13/67) of patients. Moreover, both ATM (P=0.002) and p53 deletion (P=0.004) were significantly associated with AID. IgV(H) gene mutation was seen in 45% (27/60) of patients. Twenty-five percent (17/69) of patients with AID-positive chronic lymphocytic leukemia/small lymphocytic lymphoma displayed a shorter survival than AID-negative chronic lymphocytic leukemia/small lymphocytic lymphoma patients (61 vs 130 months, P=0.001). Although there was a trend, we could not show an association with the IgV(H) gene mutation status. Taken together, our study shows that AID expression is an indicator of an unfavorable prognosis in chronic lymphocytic leukemia/small lymphocytic lymphoma patients, although it is not a surrogate marker for the IgV(H) status. Furthermore, the microenvironment of proliferation centers seems to influence AID regulation and might be an initiating factor in its transformation.

A new emerging genotype subgroup within PCV-2b dominates the PMWS epizooty in Switzerland.

Postweaning multisystemic wasting syndrome (PMWS) is among the most important emerging pig diseases worldwide. Initially, the insidious nature of the disease made it difficult to pinpoint the pathogen. The presence of porcine circovirus type 2 (PCV2) in all PMWS diseased animals led to its acceptance, possibly together with an unknown factor, as the causative agent for PMWS. Also, presence of PCV2 in healthy individuals did not facilitate the understanding of the disease. Phylogenetic classification separates PCV2 viruses into at least two major groups. With the aid of a signature motif, a short amino acid motif encoded within the capsid protein, the viruses are determined as belonging to PCV-2a or PCV-2b. Recently, this classification received more attention, as it seemed to define PCV-2b to be more virulent. This simplification, however, could not be confirmed experimentally. Hence, we investigated whether virus genetic shift was an initiator for the PMWS epizooty in Switzerland. Piglet lymphoid tissues from 1973 to 2005 were investigated by histology, immunohistochemistry (IHC) and PCR. For genotype classification, a sequence amplificate of 137bp was used encompassing the signature motif. The onset of Swiss PMWS epizooty exhibited a marked shift in PMWS diseased and subclinically infected piglets to PCV-2b and specifically to one genotype subgroup. Complementary to these observations, healthy piglets also defined by IHC as negative are positive in the PCR reaction and are void of any PCV-2b virus during epizooty. Consequently, our data support PCV2 genome plasticity as a major contributing factor for PMWS disease manifestation.

Heightened incidence of sporadic Creutzfeldt-Jakob disease is associated with a shift in clinicopathological profiles.

Incidences of human transmissible spongiform encephalopathies are monitored by national registries in the majority of countries in Western Europe. During the past 13 years incidences for Creutzfeldt-Jakob disease (CJD) in Switzerland fluctuated between 0.4 and 2.63 cases/10(6) inhabitants. We have compared clinicpathological patient profiles including geographic and gender distribution, age at disease onset, duration of disease, clinical symptoms, and recognized or hypothetical risk factors for CJD, genetic risk factors, biochemical and histopathological data for two cohorts of Swiss sporadic CJD patients from years of regular sporadic CJD incidence (1996-2000, mean incidence 1.3 cases/10(6) inhabitants, n = 47) to Swiss sporadic CJD patients from years of elevated sporadic CJD incidence (2001-2004, mean incidence 2.3 cases/10(6) inhabitants, n = 73). Sporadic CJD patients from the cohort with elevated sporadic CJD incidence presented with a higher frequency of rare sporadic CJD subtypes. Patients of these subtypes were significantly older and showed a skewed male/female ratio when compared to published patients of identical sporadic CJD-types or to patients from the 1996-2000 cohort and indicates that improved detection of rare sporadic CJD subtypes may have contributed to increased incidence.

Inter-laboratory validation of PCR-based HPV detection in pathology specimens.

The detection and typing of human papilloma virus (HPV) in pathology specimens is gaining increasingly in importance. In the context of the initiative for quality assurance in pathology (QuIP) of the German Society of Pathology and the Professional Association of German Pathologists, four panel laboratories with experience and expertise in polymerase chain reaction (PCR)-based HPV detection were selected to establish an inter-laboratory trial. In a first step, these laboratories performed an internal testing of their own methodologies, which comprised DNA sequencing, multiplex nested PCR and hybridization techniques. Material from 39 samples including paraffin sections and DNA preparations of tissues and plasmids were evaluated by each panel institute according to their own protocols. Despite the different methodologies, a high degree of inter-laboratory reliability was achieved. In this report, we summarise the results. Pretested specimens are available for the external trail and can be ordered from the steering institute via provitro GmbH Berlin ( http://www.provitro.de ). Supplementary data are online available at http://pathologie-ccm.charite.de (rubric "Forschung"), which includes a web-based photo gallery of HPV-associated lesions and their potential association with specific virus types. The initiative is intended to foster the quality assurance of molecular HPV analysis in pathology and its correlation with morphological changes.