Molecular and genomic characterization of a novel equine molluscum contagiosum-like virus.

Cases of pox-like lesions in horses and donkeys have been associated with poxviruses belonging to different genera of the family Poxviridae. These include the orthopoxviruses vaccinia virus (VACV), horsepoxvirus (HPXV) and cowpoxvirus (CPXV), as well as a potentially novel parapoxvirus and molluscum contagiosum virus (MOCV). However, with the exception of VACV, HPXV and CPXV, the genomic characterization of the causative agents remains largely elusive with only single short genome fragments available. Here we present the first full-length genome sequence of an equine molluscum contagiosum-like virus (EMCLV) directly determined from skin biopsies of a horse with generalized papular dermatitis. Histopathological analysis of the lesions revealed severe epidermal hyperplasia with numerous eosinophilic inclusion bodies within keratinocytes. Virions were detected in the lesions in embedded tissue by transmission electron microscopy. The genome sequence determined by next- and third-generation sequencing comprises 166 843 nt with inverted terminal repeats (ITRs) of 3473 nt. Overall, 20 of the predicted 159 ORFs have no equivalents in other poxviruses. Intriguingly, two of these ORFs were identified to encode homologues of mammalian proteins involved in immune signalling pathways, namely secreted and transmembrane protein 1 (SECTM1) and insulin growth factor-like family receptor 1 (IGFLR1), that were not described in any virus family so far. Phylogenetic analysis with all relevant representatives of the Poxviridae suggests that EMCLV should be nominated as a new species within the genus Molluscipoxvirus.

[CMTM2 is involved in spermiogenesis in mice].

OBJECTIVE: To investigate whether CKLF-like MARVEL transmembrane domain-containing protein 2 (CMTM2) is involved in spermatogenesis in mice. CMTM2 is highly expressed in testis, and could possibly be a potential spermagogenesis specific gene. METHODS: CMTM2-deficient mouse model was generated. Northern, RT-PCR and Western blotting analysis were performed on total RNA derived from wild-type (WT, CMTM2+/+) and CMTM2+/- (heterozygote) and CMTM2-/-(homozygote) mice to examine the CMTM2 level. The number of litters and the number of pups were counted and pregnancy rates calculated. The motility and morphology of the sperm and the histology of testes were analyzed. Serum testosterone and FSH concentrations were also measured. Standard t-tests were used and standard error of means were calculated. RESULTS: CMTM2 was highly expressed in a finely regulated pattern in the mouse testis during spermatogenesis. The body weight of adult mice with CMTM2 deficiency was not significantly different from that of wild type mice. No obvious anatomical or behavioral abnormalities were observed. The testis of CMTM2-/- was smaller than that of CMTM2+/+ mice. The testis diameter in wild mice and CMTM2 null mice were (11.32±1.21) mm vs. (8.29±1.92) mm (P<0.05), and the weights were (101.63±2.33) mg vs. (85.22±2.84) mg (P<0.05), respectively. Female CMTM2 null mice were fertile, indicating that CMTM2 was not required for female gametogenesis. The CMTM2-/- mice produced virtually no sperm, and CMTM2+/- mice sperm count showed a significant decline. In terms of sperm morphorlogy study, more round spermatids could be observed in the heterozygote group, compared with the wild type group; while in the homozygote group, a large amount of round spermatids could be observed because of complete arrest of spermiogenesis. The hormone levels were not significantly different. The CMTM2-/- male mice were sterile due to a late, complete arrest of spermiogenesis. The organized architecture of the seminiferous epithelium of the seminiferous tubules seen in CMTM2+/+ mice was lost in CMTM2-/- mice. CONCLUSION: This study suggests CMTM2 is not required for embryonic development in the mouse but is essential for spermiogenesis, however, further studies are required for more detailed mechanism study.

Anti-inflammatory drugs decrease infection of brain endothelial cells with EHV-1 in vitro.

BACKGROUND: Equine herpesvirus-associated myeloencephalopathy is the result of endothelial cell infection of the spinal cord vasculature with equine herpesvirus-1 (EHV-1) during cell-associated viraemia. Endothelial cell infection requires contact between infected peripheral blood mononuclear and endothelial cells. Inflammation generated during viraemia likely upregulates adhesion molecule expression on both cell types increasing contact and facilitating endothelial cell infection. OBJECTIVES: Evaluating the role of anti-inflammatory drugs in decreasing endothelial cell infection with EHV-1. STUDY DESIGN: In vitro assay, crossover design, multiple drug testing. METHODS: In vitro modified infectious centre assay using immortalised carotid artery endothelial cells or primary brain endothelial cells with plaque counts per well as outcome. Cells were either anti-inflammatory drug treated or left untreated. RESULTS: Significant reduction of plaque count when cells were treated compared with untreated cells. No dose-dependent effect when drug concentrations were increased to 10× dose. Treatment of both peripheral blood mononuclear cells (PBMC) and endothelial cells (EC) is required for significant plaque count reduction. MAIN LIMITATIONS: In vitro study. CONCLUSIONS: Anti-inflammatory drugs decrease infection of endothelial cells likely by reducing contact between EHV-1 infected PBMC and endothelial cells in vitro. The role of adhesion molecules in this process needs further investigation. In vitro results suggest anti-inflammatory drug therapy during EHV-1 infection and viraemia in horses could be clinically relevant.

Detection of canine oral papillomavirus DNA in conjunctival epithelial hyperplastic lesions of three dogs.

Papillomavirus infections are responsible for plaques and papillomas in various locations on the skin and in mucous membranes. The aim of this report was to describe morphologic features of a viral pigmented conjunctival plaque and 2 conjunctival squamous papillomas in 3 dogs, and to investigate these lesions for the presence of papillomavirus DNA by polymerase chain reaction (PCR), DNA sequence analysis, and in situ hydridization (ISH). Histopathology revealed in all neoplasms various degrees of epithelial hyperplasia, acanthosis, and hyperkeratosis with koilocytosis. In all lesions E6, E7, and L1 gene fragments of canine oral papillomavirus (COPV) DNA were detected by PCR and sequencing analysis. ISH revealed COPV DNA in a highly specific pattern within nuclei of the hyperplastic epithelium. The presence of canine papillomavirus in ocular conjunctival plaques and papillomas suggests these benign lesions may have the potential for malignant transformation. This is the first time that the lambdapapillomavirus COPV has been detected in ocular epithelial hyperplastic lesions.

Recurrent uveitis in horses: vitreal examinations with ultrastructural detection of leptospires.

This study documents the examination of 17 horses (both sexes, 3-18 years old) suffering from spontaneous equine recurrent uveitis (ERU). Vitreal samples obtained by pars plana vitrectomy were examined macroscopically and ultrastructurally, and in most cases also by cultural examination, by microscopic agglutination test (MAT) and by polymerase chain reaction. In 24% (4/17) of the animals, ultrastructural examination by electron microscopy revealed intact leptospiral bacteria in the vitreous. The leptospires were detected freely in the vitreous and also incorporated by a phagocyte. They were surrounded by a rim of proteinaceous material which was reduced around a phagocytosed leptospira. Ninety-four per cent (16/17) of the vitreal samples presented significant antibody levels in the MAT, mostly against leptospiral serovar Grippotyphosa. Seventy-five per cent (9/12) of bacterial culture examinations were positive for leptospira. Polymerase chain reaction was positive in all (16/16) examinations performed. Our findings support previous reports suggesting that leptospires play an important role in the pathogenesis of ERU. Interestingly, this study found leptospires after secondary and later acute episodes. A persistent leptospiral infection is therefore suggested as the cause of ERU.

Two independent cases of spontaneously occurring branchioblastomas in koi carp (Cyprinus carpio).

This report documents 2 cases of branchioblastomas in koi carp (Cyprinus carpio). Macroscopically, both cases were characterized by well-demarcated, pale red nodular masses located at the left first branchial arch and the right pseudobranch, respectively. Histologically, the neoplasias were composed of blast-like cells that differentiated into cartilage and branchial lamellae embedded in abundant fibrous connective tissue. Based on these findings, a branchioblastoma was diagnosed.

Comparative morphologic and immunohistochemical investigation of spontaneously occurring thymomas in a colony of European hamsters.

This study documents the characteristics of a large series of spontaneously occurring thymomas in a laboratory colony of European hamsters (Cricetus cricetus). Thymomas are rare organotypic neoplasms originating from the thymic epithelial compartment. Because the hamster thymomas largely resembled their human counterparts, the recent World Health Organization (WHO) classification of human thymic epithelial tumors was used. Forty hamsters of both sexes aged 3-29 months were examined macroscopically and histologically. In 22 (55%) of the 40 animals, necropsy revealed enormous whitish masses in the anterior mediastinum, with a diameter ranging from 0.5 to 4.5 cm and a lobulated structure. The anatomy of the thymus region was normal in the remaining 18 hamsters. Histologically, the tumors presented as thymuslike organoid structures with areas of medullary and cortical differentiation and a predominance of lymphoid cells. A network of epithelial cells in the cortical areas, demonstrated immunohistochemically with a cross-reactive antibody against pancytokeratin, supported the diagnosis of thymoma. Cortical lymphocytes showed positive staining with cross-reacting antibodies against CD3 and terminal deoxynucleotidyl transferase, characteristic of immature T cells. On the basis of these findings, the tumors were classified as B1 thymomas, in some cases with AB or B2 components, according to the new WHO classification for human thymic epithelial tumors.

Immunoglobulin adsorption in patients with idiopathic dilated cardiomyopathy.

BACKGROUND: Idiopathic dilated cardiomyopathy (IDC) frequently is a progressive disease without causative therapy options. Following the hypothesis that in certain patients autoantibodies against cardiac structures may induce, maintain, or promote the progression of the disease, we investigated whether the elimination of these autoantibodies through immunoadsorption would improve cardiac function. METHODS AND RESULTS: This prospective case-control study included 34 patients with IDC. Each patient presented with moderate to severe heart failure and evidence of autoantibodies directed against beta(1)-adrenoceptors (beta(1)-AABs). Seventeen patients received standard medical therapy (control group), whereas 17 were also treated with immunoadsorption (treatment group) to eliminate beta(1)-AABs. A 1-year follow-up included echocardiographic assessment of left ventricular ejection fraction and internal diameters, beta(1)-AAB levels, and clinical status every 3 months. Within 1 year, the mean+/-SD left ventricular ejection fraction rose from 22.3+/-3.3% to 37.9+/-7.9% (P=0.0001) in the treatment group, with a relative increase of 69.9%. However, in the control group, no overall increase was seen (from 23.8+/-3.0% to 25.2+/-5.9%, P=0. 3154). Left ventricular diameter in diastole decreased by 14.5% from 74.5+/-7.1 to 63.7+/-6.0 mm in the treatment group (P=0.0001) and by 3.8% (P=0.2342) in the control group. In the treatment group, the NYHA functional rating improved after immunoadsorption (P=0.0001). beta(1)-AABs did not increase anew. CONCLUSIONS: In IDC, the use of immunoadsorption is superior to the use of standard medical therapy. It significantly improves cardiac performance and clinical status.

Weaning from mechanical cardiac support in patients with idiopathic dilated cardiomyopathy.

BACKGROUND: Implantation of mechanical cardiac support systems (MCSS) in patients with idiopathic dilated cardiomyopathy (IDC) may improve cardiac function and allow explantation of the device. We report of long-term effects of ventricular unloading on cardiac function, humoral anti-beta1-adrenoceptor autoantibodies (A-beta1-AABs), and myocardial fibrosis. METHODS AND RESULTS: Seventeen patients in New York Heart Association functional class IV with nonischemic IDC received MCSS. All had a cardiac index of < 1.6 L x min(-1) x m(-2) of body surface area, a left ventricular ejection fraction (LVEF) of <16%, and a left ventricular internal diameter in diastole (LVIDd) of >68 mm and tested positive for A-beta1-AABs. Echocardiographic evaluation, serum tests for A-beta1-AABs, and histological assessment of myocardial fibrosis were performed before and after MCSS implantation. The mean support duration was 230+/-201 days. Six patients died, four were transplanted, and two are still on MCSS. Five patients with significant cardiac recovery (mean LVIDd, 54+/-2.3 mm; LVEF, 47+/-3.7%) were weaned after 160 to 794 days and are now device free for 51 to 592 days. A-beta1-AABs disappeared gradually during MCSS without increase after weaning; cardiac function and volume density of fibrosis remained normal. Nine patients' cardiac function hardly improved during ventricular unloading. CONCLUSIONS: Cardiac function can be normalized in selected patients with end-stage IDC by MCSS. The degree of preoperative myocardial fibrosis may be an indicator for outcome; A-beta1-AABs can be used to monitor myocyte recovery. Weaning from MCSS offers an alternative to cardiac transplantation in certain patients.

[Temporary mechanical left heart support. Recovery of heart function in patients with end-stage idiopathic dilated cardiomyopathy]. / Temporäre mechanische Linksherzentlastung. Erholung der Herzfunktion bei Patienten mit idiopathischer dilatativer Kardiomyopathie im Endstadium.

BACKGROUND: Implantation of a mechanical cardiac support system (MCSS) in patients with idiopathic dilated cardiomyopathy (IDC) may improve cardiac function and allow explantation of the device. Our experience now includes 13 patients who have been "weaned" from MCSS and we report about the overall results of this treatment as well as the effects of ventricular unloading on cardiac function, anti-beta 1-adrenoceptor-autoantibody (A-beta 1-AAB) level and the degree of myocardial fibrosis. METHODS: 13 patients with non-ischemic IDC who had been admitted here in cardiogenic shock (CI < 1.61.min-1.m2, left ventricular ejection fraction [LVEF] < 16% and left ventricular internal diameter in diastole [LVIDd] > 68 mm) and who all tested positive for A-beta 1-AABs were implanted with an uni-(12 patients) or a biventricular (1 patient) mechanical assist device. Echocardiographic evaluation and A-beta 1-AAB-level-monitoring was routinely performed after implantation and explantation of the MCSS and the degree of myocardial fibrosis was assessed at the time of implantation and after explantation. RESULTS: During a mean duration of mechanical support of 236 +/- 201 days (range: 30 to 794 days), LV-EF improved to a mean of 46% and LVIDd decreased to a mean value of 56 mm in these 13 patients. A-beta 1-AABs decreased and disappeared 11.7 weeks after implantation of the device and did not reincrease thereafter. The highly pathologic degree of fibrosis at the time of implantation diminished to normal values about 1 year after explantation. One patient died of anesthesiologic complications and another patient shortly presented with a new episode of cardiac insufficiency 6 months after explantation. He was implanted again with an univentricular assist device was successfully transplanted 3 weeks later. Mean observation period of the remaining 11 patients now amounts to 12.6 +/- 9.77 (range: 3 to 26) months after explantation of the device--as of May, 31, 1997--with a cumulative observation period of 139 patient months. CONCLUSION: Temporary implantation of a MCSS may normalize cardiac function in selected patients with IDC. The striking degree of myocardial fibrosis can reduce to normal values after explantation of the device. A-beta 1-AABs disappear during ventricular unloading and do not increase thereafter. "Weaning" from mechanical device may constitute an alternative treatment to cardiac transplantation in selected patients.