On Temnocephala axenos Monticelli, 1898 (Platyhelminthes, Temnocephalida): taxonomic status and designation of a neotype.

Temnocephala axenos Monticelli, 1898 was described based on specimens from an unidentified host collected in Blumenau, Santa Catarina, Brazil. Information about type locality was imprecise and the host was later identified as Aegla laevis (Latreille, 1818). However, it is known that A. laevis is not present on the eastern side of the Andes. Also, only histological preparations from one specimen studied by Monticelli are currently available in the Museum für Naturkunde Berlin, but it showed none of the taxonomic characters needed for the characterization of the species. Although the updated description of the species based on Uruguayan specimens, neither the author nor the several previous studies about the species showed a search for the type material, a resolution for the misidentification of the type host or the imprecise type locality due to the subsequent geographical division of the municipality cited in the description. The Uruguayan specimens were not even geographically close to the type locality and a neotype was not designed to validate the species' taxonomic status again. Specimens from Santa Catarina and Paraná States, Brazil, were studied, as well as restudied Argentinean specimens. The new data were compared with the update description of the species. The historical background and the discussion about geographical origins and hosts of the species, as well as a designation of a neotype, allow comparative material of the type locality and type host to exist, eliminating doubts about the identification of T. axenos.

Semen quality is affected by HLA class I alleles together with sexually transmitted diseases.

BACKGROUND: The human leukocyte antigen (HLA) locus includes several genes with key roles in antigen presentation and immune response, some of them inclusively found to be associated with non-obstructive azoospermia. Still, HLA connections to other infertility phenotypes such as semen hyperviscosity (SHV), asthenozoospermia (AST), and oligozoospermia (OLI) have been often neglected. OBJECTIVES: In this work, we aimed to evaluate the association of HLA class I and II genes with SHV, AST, and OLI phenotypes while exploring a possible role in an adaptive immune response to sexually transmitted diseases (STD). MATERIALS AND METHODS: Whole-exome sequencing was performed in a Portuguese cohort of 71 infertility cases and 68 controls, followed by HLA typing using a specific software-HLA*PRG:LA tool. Molecular screenings of seven STD were carried out in a subset of 72 samples (30 cases and 42 controls). RESULTS: Statistical tests uncovered three protective alleles: HLA-A*11:01, associated with all forms of male infertility (p = 0.0006); HLA-DQB1*03:02 with SHV and OLI (PSHV  = 0.0303, POLI  = 0.0153); and HLA-A*29:02 with OLI (p = 0.0355), which was found to interfere in sperm number together with HPV (p = 0.0313). Five risk alleles were also identified: two linked with SHV (HLA-B*50:01, p = 0.0278; and HLA-C*06:02, p = 0.0461), another one with both SHV and OLI (HLA-DQA1*05:01, PSHV  = 0.0444 and POLI =0.0265), and two with OLI (HLA-C*03:03, p = 0.0480; and HLA-DQB1*03:01, p = 0.0499). Here, HLA-C*03:03 carriers tend to be HPV infected. CONCLUSIONS: The application of HLA*PRG:LA tool to the study of male infertility provided novel insights for an HLA correlation with semen quality, namely among SHV and OLI phenotypes. The discovery of an HLA-A*29:02/HPV crosstalk, together with former reports of HLA alleles conferring resistance-susceptibility to diverse human pathogens, raises the hypothesis of a mechanistic link between male infertility, HLA polymorphism, and host response to STD.

Portuguese consensus document for the management of alpha-1-antitrypsin deficiency.

Alpha-1-antitrypsin deficiency (AATD) is a genetic autosomal codominant disorder caused by mutations in SERPINA1 gene. It is one of the most prevalent genetic disorders, although it remains underdiagnosed. Whereas at international level there are several areas of consensus on this disorder, in Portugal, inter-hospital heterogeneity in clinical practice and resources available have been adding difficulties in reaching a diagnosis and in making therapeutic decisions in this group of patients. This raised a need to draft a document expressing a national consensus for AATD. To this end, a group of experts in this field was created within the Portuguese Pulmonology Society - Study group on AATD, in order to elaborate the current manuscript. The authors reviewed the existing literature and provide here general guidance and extensive recommendations for the diagnosis and management of AATD that can be adopted by Portuguese clinicians from different areas of Medicine. This article is part of a supplement entitled "Portuguese consensus document for the management of alpha-1-antitrypsin deficiency" which is sponsored by Sociedade Portuguesa de Pneumologia.

Alpha-1 antitrypsin deficiency caused by a novel mutation (p.Leu263Pro): Pi*ZQ0gaia - Q0gaia allele.

Severe alpha-1 antitrypsin deficiency (AATD) is generally associated with PI*ZZ genotype and less often with combinations of PI*Z, PI*S, and other rarer deficiency or null (Q0) alleles. Severe AATD predisposes patients to various diseases, including pulmonary emphysema. Presented here is a case report of a young man with COPD and AATD. The investigation of the AATD showed a novel mutation p.Leu263Pro (c.860T>C), which was named Q0gaia (Pi*ZQ0gaia). Q0gaia is associated with very low or no detectable serum concentrations of AAT.

Rare double sex and mab-3-related transcription factor 1 regulatory variants in severe spermatogenic failure.

The double sex and mab-3-related transcription factor 1 (DMRT1) gene has long been linked to sex-determining pathways across vertebrates and is known to play an essential role in gonadal development and maintenance of spermatogenesis in mice. In humans, the genomic region harboring the DMRT gene cluster has been implicated in disorders of sex development and recently DMRT1 deletions were shown to be associated with non-obstructive azoospermia (NOA). In this work, we have employed different methods to screen a cohort of Portuguese NOA patients for DMRT1 exonic insertions and deletions [by multiplex ligation probe assay (MLPA); n = 68] and point mutations (by Sanger sequencing; n = 155). We have found three novel patient-specific non-coding variants in heterozygosity that were absent from 357 geographically matched controls. One of these is a complex variant with a putative regulatory role (c.-223_-219CGAAA>T), located in the promoter region within a conserved sequence involved in Dmrt1 repression. Moreover, while DMRT1 domains are highly conserved across vertebrates and show reduced levels of diversity in human populations, two rare synonymous substitutions (rs376518776 and rs34946058) and two rare non-coding variants that potentially affect DMRT1 expression and splicing (rs144122237 and rs200423545) were overrepresented in patients when compared with 376 Portuguese controls (301 fertile and 75 normozoospermic). Overall our previous and present results suggest a role of changes in DMRT1 dosage in NOA potentially also through a process of gene misregulation, even though DMRT1 deleterious variants seem to be rare.

Severe α-1 antitrypsin deficiency caused by Q0(Ourém) allele: clinical features, haplotype characterization and history.

α-1 Antitrypsin deficiency (AATD) caused by null alleles is associated with the total lack of protein and generally it translates into more severe clinical features of pulmonary disease. This is the case of Q0(Ourém) , a rare variant found in several families of Central Portugal caused by the L353fsX376 mutation. A total of 41 patients carrying at least one copy of Q0(Ourém) were evaluated for SERPINA1 levels, respiratory function values and lung parenchyma status (chest X-ray and computerized tomography scan). Q0(Ourém) haplotype background was characterized using seven microsatellites flanking SERPINA1 and Q0(Ourém) age was estimated by a statistical method relying on the decay of haplotype sharing at linked markers (DHSMAP). Homozygous patients showed a compromised lung function and extensive emphysema. SQ0(Ourém) , although having serum levels below the 11 µM threshold, did not necessarily result in signs of disease. MQ0(Ourém) were found to be a heterogeneous group, mainly composed of normal individuals. Eight Q0(Ourém) haplotypes were identified and the allele was estimated to have arisen 650 years ago. Q0(Ourém) was associated with mild to severe AATD and has a single origin, probably linked to the major Ourém settlements where the occurrence of severe AATD may not be explained by recent consanguinity.

alpha1-Antitrypsin null alleles: evidence for the recurrence of the L353fsX376 mutation and a novel G-->A transition in position +1 of intron IC affecting normal mRNA splicing.

alpha1-Antitrypsin (PI) deficiency is a common autosomal recessive disorder associated with emphysema and liver disease, which may result from a wide spectrum of mutations causing a reduction of serum levels (deficient alleles) or a total lack of circulating protein (null alleles). We report two different alleles associated with the absence of isoelectric focusing banding patterns in Portuguese patients with emphysema. The first allele, Q0(ourém), results from the recurrence of the defining mutation of the Q0(mattawa) variant (L353fsX376) on a M3 normal background. The second allele, Q0(porto), has a novel G-->A mutation at position +1 of the intron IC (IVS1C+1G-->A), which restricts mononuclear phagocyte transcripts to mRNA species resulting from the direct splice of exon IA to exon II. The absence of this normal splice alternative in the liver, where PI is primarily synthesized, provides a basis for the pathogenic effects of this mutation.

Patterns of haplotype diversity within the serpin gene cluster at 14q32.1: insights into the natural history of the alpha1-antitrypsin polymorphism.

The levels of haplotype diversity associated with different alpha1-antitrypsin (PI) alleles were assessed by the analysis of three microsatellites located within or close to corticosteroid-binding globulin (CBG), alpha1-antitrypsin [PI-(TG)n] and protein C inhibitor [PCI-(TG)n] loci in three populations with different historic backgrounds: Portugal, the Basque Country and São Tomé Príncipe (Gulf of Guinea). Unlike the more distant PCI-(TG)n repeat, allelic variation at PI-(TG)n reflected distinct phases of mutational recovery of microsatellite diversity around different founder alleles and showed a considerable differentiation between alpha1-antitrypsin protein variants. In accordance with population history, the Basque sample presented overall reduced levels of microsatellite variation. The African sample, although presenting the highest PCI-(TG)n diversity, showed a lineage-specific reduction in PI-(TG)n heterozygosity within the oldest M1Ala213 variant that could have been caused by (1) selection at a closely linked locus or (2) biases in the microsatellite mutation process leading to a stable equilibrium distribution. Age estimates of alpha1-antitrypsin variants based on microsatellite variation suggest that the Z deficiency allele appeared 107-135 generations ago and could have been spread in Neolithic times. The S mutation has an older 279- to 470-generation age, indicating that its high frequencies in Iberia did not result from a recent bottleneck and that PI*S could have originated in this region. M2 and M3 types had lower age estimates than would be expected from their wide geographical distributions, suggesting that their dispersion in Europe might have been preceded by important bottlenecks.

Haplotype analysis of the apolipoprotein E and apolipoprotein C1 loci in Portugal and São Tomé e Príncipe (Gulf of Guinea): linkage disequilibrium evidence that APOE*4 is the ancestral APOE allele.

The joint distributions of phenotypes from the apolipoprotein E gene (APOE) and from a closely linked restriction site polymorphism at the apolipoprotein C1 locus (APOC1) were studied in population samples from Portugal and São Tomé e Príncipe (Gulf of Guinea), a former Portuguese colony that was originally populated by slaves imported from the African mainland. The frequencies of the APOE alleles (*2, *3, and *4) in Portugal and São Tomé fitted the ranges of variation generally observed in European and African populations, respectively. Haplotype analysis showed that in both populations the strength of linkage disequilibrium was highest for the APOE*2 allele and lowest for the APOE*4 allele, suggesting that the origin of the APOE alleles followed a 4-->3-->2 pathway and thus providing independent confirmation of the results from sequence homology studies with nonhuman primates. In accordance with global trends in the distribution of human genetic variation, the European sample from Portugal presented more intense linkage disequilibrium between APOE and APOC1 than the African sample from São Tomé where, despite the short 4-kb distance that separates the 2 loci, the level of association between the APOC1 alleles and APOE*4 was nonsignificant.

[The importance of mdx mouse in the physiopathology of Duchenne's muscular dystrophy]. / Importância do camundongo mdx na fisiopatologia da distrofia muscular de Duchenne.

The mdx mouse develop an X-linked recessive muscular dystrophy (locus Xp21.1) and lack dystrophin expression. Despite showing less intense myofibrosis and scarce deposition of fatty tissue, mdx mice are considered an adequate animal model for studies on the pathogenesis of Duchenne-type muscular dystrophy. Marked histological alterations in the muscular tissues associated to myonecrosis and inflammatory mononuclear cell infiltrate (lymphocytes, monocytes/macrophages) suggest a participation of the immune system in this myopathy. Modulation of the extracellular matrix (ECM) components in the muscular tissue during all phases (onset, myonecrosis and regeneration) of disease, indicate an important role for the ECM driving inflammatory cells to the foci of lesion. Therefore mdx mice should be regarded as an important tool for studies on pathogenetic mechanisms of Duchenne-type muscular dystrophy. Such experimental model would allow development of new therapeutic approaches for increasing survival and clinical amelioration.

Altered deposition of extracellular matrix components in the skeletal muscle and lymph node of the MDX dystrophic mouse.

1. MDX mice derived from a colony of C57BL/10ScSn mice develop an X-linked recessive muscular dystrophy, thus providing an adequate model to study the pathogenesis of muscular dystrophy. 2. Skeletal myofibers of MDX mutant mice were heterogeneous, with disorganization of myofilaments and the absence of immunolabelling for dystrophin with monoclonal antibody DY4/6D3. 3. Marked deposition of reticulin, collagenic fiber (types, I, IV) and laminin (LN) were consistently present mostly around lesioned and necrotic myofibers associated with an intense inflammatory reaction, whereas strong immunolabelling for TIII-C, TIV-C and FN was often associated with regenerated fibers. 4. During the onset (3 weeks of postnatal life) of disease and height of myonecrosis (5-6 weeks of postnatal life), popliteal lymph nodes showed dense argyrophilic meshwork, intense immunolabelling for collagens types I and IV, FN, LN and enlargement of the hili which were packed with mononuclear cells. Such alterations, albeit less intense, were still observed in MDX mice with 20 weeks of postnatal life. 5. The results support the view that ECM components might be influencing the migration of inflammatory cells and the process of myonecrosis in the skeletal muscle of MDX dystrophic mice.

Altered deposition of extracellular matrix components in the skeletal muscle and lymph node of the MDX dystrophic mouse

1. MDX mice derived from a colony of C57BL/10ScSn mice develop an X-linked recessive muscular dystrophy, thus providing an adequate to study the pathogenesis of muscular dystrophy. 2. Skeletal myofibers of MDX mutant mice were heterogenous, with disorganization of myofilaments and the absence of immunolabelling for dystrophin with monoclonal antibody DY4/6D3. 3. Marked deposition of reticulin, collagenic fiber (types I, IV) and laminin (LN) were consistently present mostly around lesioned and necrotic myofibers associated eith an intense inflammatory reaction, whereas strong immunolabelling for TIII-C, TIV-C and FN was often associated with regenerated fibers. 4. During the onset (3 weeks of postnatal life) of disease and height of myonecrosis (5-6 weeks of postnatal life), popliteal lymph nodes showed dense argyrophilic meshwork, intense immunolabeling for collagens types I and IV, FN, LN and enlargement of the hili which were packed with mononuclear cells. Such alterations, albeit less intense, were still observed in MDX mice with 20 weeks of postnatal life. 5 The results support the view that ECM components might be influencing the migration of inflammatory cells and the process of myonecrosis in the skeletal muscle of MDX dystrophic mice