Standardization of a multiplex assay to identify weak D types in a mixed-race Brazilian population.

RH allele variability is caused by several types of variants, resulting in altered RhD and RhCE phenotypes. Most of the weak D phenotypes in European-derived populations are weak D types 1, 2, or 3, which are not involved in alloimmunization episodes. However, the Brazilian population is racially diverse, and the accuracy of molecular and serologic tests developed in recent years has allowed for the identification of other RH variants, that are common in the Brazilian population, such as weak D type 38 or weak partial 11, the latter involved in alloimmunization cases. Furthermore, patients with these two weak D variants must be transfused with D- red blood cell units, as do patients with weak D type 4 or DAR, which are also common D variants in Brazil. Weak D type 38 and weak partial 11 can be serologically misclassified as weak D types 1, 2, or 3 in patients, based on European experience, or as D- in donors. Additionally, pregnant women may unnecessarily be identified as requiring Rh immune globulin. RhCE phenotypes are reliable indicators of RhD variants. For individuals with the Dce phenotype, the preferred approach is to specifically search for RHD*DAR. However, when encountering DCe or DcE phenotypes, we currently lack a developed method that assists us in rapidly identifying and determining the appropriate course of action for the patient or pregnant woman. Two multiplex assays were proposed: one for the identification of RHD*weak partial 11, RHD*weak D type 38, and RHD*weak D type 3 and another for RHD*weak D type 2 and RHD*weak D type 5. The multiplex assays were considered valid if the obtained results were equivalent to those obtained from sequencing. Expected results were obtained for all tested samples. The proposed multiplex allele-specific polymerase chain reaction assays can be used in the molecular investigation of women of childbearing age, patients, and blood donors presenting a weak D phenotype with DCe or DcE haplotypes in a mixed-race population, such as Brazil.

Transcriptome of iPSC-derived neuronal cells reveals a module of co-expressed genes consistently associated with autism spectrum disorder.

Evaluation of expression profile in autism spectrum disorder (ASD) patients is an important approach to understand possible similar functional consequences that may underlie disease pathophysiology regardless of its genetic heterogeneity. Induced pluripotent stem cell (iPSC)-derived neuronal models have been useful to explore this question, but larger cohorts and different ASD endophenotypes still need to be investigated. Moreover, whether changes seen in this in vitro model reflect previous findings in ASD postmortem brains and how consistent they are across the studies remain underexplored questions. We examined the transcriptome of iPSC-derived neuronal cells from a normocephalic ASD cohort composed mostly of high-functioning individuals and from non-ASD individuals. ASD patients presented expression dysregulation of a module of co-expressed genes involved in protein synthesis in neuronal progenitor cells (NPC), and a module of genes related to synapse/neurotransmission and a module related to translation in neurons. Proteomic analysis in NPC revealed potential molecular links between the modules dysregulated in NPC and in neurons. Remarkably, the comparison of our results to a series of transcriptome studies revealed that the module related to synapse has been consistently found as upregulated in iPSC-derived neurons-which has an expression profile more closely related to fetal brain-while downregulated in postmortem brain tissue, indicating a reliable association of this network to the disease and suggesting that its dysregulation might occur in different directions across development in ASD individuals. Therefore, the expression pattern of this network might be used as biomarker for ASD and should be experimentally explored as a therapeutic target.

Two novel pathogenic variants in MED13L: one familial and one isolated case.

BACKGROUND: Genetic variants involving the MED13L gene can lead to an autosomal dominant syndrome characterised by intellectual disability/developmental delay and facial dysmorphism. METHODS: We investigated two cases (one familial and one isolated) of intellectual disability with speech delay and dysmorphic facial features by whole-exome sequencing analyses. Further, we performed a literature review about clinical and molecular aspects of MED13L gene and syndrome. RESULTS: Two MED13L variants have been identified [MED13L(NM_015335.5):c.4417C>T and MED13L(NM_015335.5):c.2318delC] and were classified as pathogenic according to the ACMG (American College of Medical Genetics and Genomics) guidelines. One of the variants was present in sibs. CONCLUSIONS: The two pathogenic variants identified have not been previously reported. Importantly, this is the first report of a familial case of MED13L nonsense mutation. Although the parents of the affected children were no longer available for analysis, their apparently normal phenotypes were surmised from familial verbal descriptions corresponding to normal mental behaviour and phenotype. In this situation, the familial component of mutation transmission might be caused by gonadal mosaicism of a MED13L mutation in a gonad from either the father or the mother. The case reports and the literature review presented in this manuscript can be useful for genetic counselling.

Leishmania amazonensis induces modulation of costimulatory and surface marker molecules in human macrophages.

Manipulation of costimulatory and surface molecules that shape the extent of immune responses by Leishmania is suggested as one of the mechanisms of evading the host's defences. The experiments reported here were designed to evaluate the expressions of CD11b, CD11c, CD14, CD18, CD54, CD80, CD86, CD206, MHC class II and TLR-2 (Toll-like receptor 2) in human macrophages infected with L. amazonensis. Phenotypic evaluation revealed a negative modulation in CD11b, CD11c, CD14, CD18, CD54 and MHC class II molecules, depending on the level of infection. The results showed that as early as 1 hour after infection no reduction in marker expression occurs, whereas after 24 hours, downregulation of these molecules was observed in macrophages. No significant changes were observed in the expressions of CD80, CD86, CD206 and TLR2. Evidence of the differential modulation of markers expression and that after parasite uptake no reduction in surface marker expression occurs indicates that parasite internalization is not involved in the phenomena of down-modulation.

Responses of Crepis japonica induced by supplemental blue light and UV-A radiation.

Crepis japonica (L.) D.C. (Asteraceae), a weed with antioxidant, antiallergenic, antiviral and antitumor properties displays both medicinal properties and nutritional value. This study aims to assess the effects of a supplementation of blue light and UV-A radiation on the growth, leaf anatomical structure and phenolic profile of the aerial parts of Crepis japonica. Plants were grown under two light treatments: W (control - white light), W + B (white light supplemented with blue light) and W + UV-A (white light supplemented with UV-A radiation). We recorded the length, width, and weight of fresh and dry leaves, the thickness of the epidermis and mesophyll, and stomata density. The phenolic profiles of the aqueous extracts of the aerial parts were analyzed by HPLC-DAD. There was an increase in the leaf size, stomatal density, and phenolic production, and a thickening of the mesophyll and epidermis. UV-A radiation increased the phenolic production more than blue light. Blue light and UV-A radiation both improved the production of caffeic acid by about 6 and 3 times, respectively, in comparison to control. This compound was first reported as a constituent of the extract from the aerial parts together with caftaric acid. UV-A also promoted the production of chlorogenic acid (about 1.5 times in comparison to the control). We observed that the morphological and chemical parameters of C. japonica are modified in response to blue light and UV-A radiation, which can be used as tools in the cultivation of this species in order to improve its medicinal properties and nutritional value.

The surgical challenge of carotid artery and Fallopian canal dehiscence in chronic ear disease: a pitfall for endoscopic approach.

OBJECTIVE: Endoscopic procedures are becoming common in middle ear surgery. Inflammation due to chronic ear disease can cause bony erosion of the carotid artery and Fallopian canals, making them more vulnerable during surgery. The objective of this study was to determine whether or not chronic ear disease increases dehiscence of the carotid artery and Fallopian canals. DESIGN: Comparative human temporal bone study. SETTING: Otopathology laboratory. PARTICIPANTS: We selected 78 temporal bones from 55 deceased donors with chronic otitis media or cholesteatoma and then compared those two groups with a control group of 27 temporal bones from 19 deceased donors with no middle ear disease. MAIN OUTCOME MEASURES: We analysed the middle ear, carotid artery canal and Fallopian canal, looking for signs of dehiscence of its bony coverage, using light microscopy. RESULTS: We found an increased incidence in dehiscence of the carotid artery and Fallopian canals in temporal bones with chronic middle ear disease. The size of the carotid artery canal dehiscence was larger in the middle ear-diseased groups, and its bony coverage, when present, was also thinner compared to the control group. Dehiscence of the carotid artery canal was more frequently located closer to the promontory. The incidence of Fallopian canal dehiscence was significantly higher in temporal bones from donors older than 18 years with chronic middle ear disease. CONCLUSION: The increased incidence of the carotid artery and Fallopian canal dehiscence in temporal bones with chronic middle ear disease elevates the risk of adverse events during middle ear surgery.

Genomic copy number alterations in non-syndromic hearing loss.

Genetic heterogeneity has made the identification of genes related to hearing impairment a challenge. In the absence of a clear phenotypic aetiology, recurrence risk estimates are often based on family segregation and may be imprecise. We profiled by oligonucleotide array-CGH patients presenting non-syndromic hearing loss with presumptive autosomal recessive (n = 50) or autosomal dominant (n = 50) patterns of inheritance. Rare copy number variants (CNVs) were detected in 12 probands; four of the detected CNVs comprised genes previously associated with hearing loss (POU4F3, EYA4, USH2A, and BCAP31) and were considered causative, stressing the contribution of genomic imbalance to non-syndromic deafness. In six cases, segregation of the CNVs in pedigrees excluded them as causative. In one case, segregation could not be investigated, while in another case, a point mutation likely explains the phenotype. These findings show that the presumptive patterns of inheritance were incorrect in at least two cases, thereby impacting genetic counselling. In addition, we report the first duplication reciprocal to the rare ABCD1, BCAP31, and SLC6A8 contiguous deletion syndrome; as with most microduplication syndromes, the associated phenotype is much milder than the respective microdeletion and, in this case, was restricted to hearing impairment.

Dynamics of blaOXA-23 gene transmission in Acinetobacter spp. from contaminated veterinary environmental surfaces: an emerging One Health threat?

BACKGROUND: Carbapenem-resistant Acinetobacter baumannii is a common pathogen associated with healthcare-acquired infections, and robust infection prevention and control protocols exist in human healthcare settings. In contrast, infection prevention and control (IPC) standards are limited in veterinary medicine, necessitating further investigation. AIM: Examine the possible transmission of carbapenem-resistant Acinetobacter spp. in a veterinary practice where a cat was diagnosed with an OXA-23-producing A. baumannii ST2 strain. METHODS: Environmental samples together with nasal and hand swabs from the veterinary personnel were collected. All swabs were screened for the presence of extended-spectrum-ß-lactamase- and carbapenemase-producing Enterobacterales, meticillin-resistant staphylococcus and multi-drug-resistant Acinetobacter spp. Whole-genome sequencing was performed for carbapenemase-producing strains. RESULTS: Of the veterinary staff, 60% carried meticillin-resistant Staphylococcus epidermidis. Environmental evaluation showed that 40% (N=6/15) of the surfaces analysed by contact plates and 40% (N=8/20) by swabs failed the hygiene criteria. Assessment of the surfaces revealed contamination with five OXA-23-producing Acinetobacter spp. strains: an OXA-23-producing Acinetobacter schindleri on the weight scale in the waiting room; and four OXA-23-producing Acinetobacter lwoffii strains, on different surfaces of the treatment room. The blaOXA-23 gene was located on the same plasmid-carrying Tn2008 across the different Acinetobacter spp. strains. These plasmids closely resemble a previously described OXA-23-encoding plasmid from a human Portuguese nosocomial Acinetobacter pittii isolate. Distinctly, the OXA-23-producing A. baumannii ST2 clinical strain had the resistant gene located on Tn2006, possibly inserted on the chromosome. CONCLUSION: The detection of an OXA-23-producing A. baumannii ST2 veterinary clinical strain is of concern for companion animal health and infection, prevention and control. This study established the dynamic of transmission of the plasmid-mediated blaOXA-23 gene on critical surfaces of a small animal veterinary practice. The genetic resemblance to a plasmid found in human nosocomial settings suggests a potential One Health link.

PLP1 duplication at the breakpoint regions of an apparently balanced t(X;22) translocation causes Pelizaeus-Merzbacher disease in a girl.

PLP1 (proteolipid protein1 gene) mutations cause Pelizaeus-Merzbacher disease (PMD), characterized by hypomyelination of the central nervous system, and affecting almost exclusively males. We report on a girl with classical PMD who carries an apparently balanced translocation t(X;22)(q22;q13). By applying array-based comparative genomic hybridization (a-CGH), we detected duplications at 22q13 and Xq22, encompassing 487-546 kb and 543-611 kb, respectively. The additional copies were mapped by fluorescent in situ hybridization to the breakpoint regions, on the derivative X chromosome (22q13 duplicated segment) and on the derivative 22 chromosome (Xq22 duplicated segment). One of the 14 duplicated X-chromosome genes was PLP1.The normal X chromosome was the inactive one in the majority of peripheral blood leukocytes, a pattern of inactivation that makes cells functionally balanced for the translocated segments. However, a copy of the PLP1 gene on the derivative chromosome 22, in addition to those on the X and der(X) chromosomes, resulted in two active copies of the gene, irrespective of the X-inactivation pattern, thus causing PMD. This t(X;22) is the first constitutional human apparently balanced translocation with duplications from both involved chromosomes detected at the breakpoint regions.

First Report of Sexual Reproduction of Fusarium solani f. sp. piperis in Bahia, Brazil.

Black pepper (Piper nigrum L.) is a popular spice native of India, and Brazil is one of its most important producing countries. The main disease of black pepper in Brazil is fusariosis, caused by Fusarium solani f. sp. piperis. Symptoms include leaf chlorosis and defoliation, blight of stems or stem cuttings, and root and foot decay. During surveys conducted in the south of the state of Bahia, municipalities of Taperoá (13°34'S, 39°10'W) and Valencia (13°20'S, 39°14'W), stems of diseased plants covered with red or salmon-colored perithecia were observed, while twigs showed leaf chlorosis, leading to early death of the plants. Ascomata were solitary or in groups, mostly superficial or surrounded by mycelia, globose, subglobose, ovoid, and 122 to 400 µm diameter. Microscopic examination revealed unitunicate, cylindric asci, 60 to 90 × 8.5 to 16 µm, thin-walled, containing eight ascospores arranged obliquely in two rows. Ascospores are hyaline, elliptical to oblong, one-septate, constricted at the central septum, 10 to 16 × 4 to 6.5 µm (means ± S.D.: 13.1 ± 1.4 × 5.1 ± 0.6 µm), length/width (L/W) 1.9 to 3.7. Single-spored cultures were transferred to SNA medium (incubated at 20°C for 7 days with 12-h photoperiod) and on potato dextrose agar (25°C in dark) for characterization. The anamorph is characterized by the presence of chlamydospores, canoe-shaped sporodochial macroconidia with three to four septae, and microconidia formed on long monophialidic conidiophores. Based on morphological markers, isolates were identified as F. solani. The partial fragment of the TEF-1α gene of single-spored isolates (CML 2186, 2187, 2188, 2189, 2190, and 2191) were sequenced. BLAST analysis of the sequence resulted in 94 to 99% identity with a reference strain of F. solani f. sp. piperis (NRRL 22570, CML 1888). For pathogenicity tests, cv. Bragantina was used and two isolates were inoculated as 5-mm diameter mycelial plugs on the stem of four plants each. Four control plants were treated only with sterile culture medium. Plants were maintained in the greenhouse at 25°C and 75 to 85% relative humidity under 70% shade. All inoculated plants showed initial symptoms of stem necrosis in inoculated branches 7 days after inoculation. Symptoms were not observed on stems of control plants. Isolates were successfully reisolated and identified as F. solani f. sp. piperis, fulfilling Koch's postulates. Representative isolates were deposited at the Coleção Micológica de Lavras (CML) at Universidade Federal de Lavras, Brazil. Production of perithecia of the pathogen has been previously reported only in Pará and Espírito Santo States (1,3). It is not yet confirmed if this taxon is homothallic or heterothallic. To our knowledge, this is the first report of the associated teleomorph of F. solani f. sp. piperis infecting and causing black pepper fusariosis in Bahia, Brazil. The results suggest that the spread of ascospores from perithecia is likely to be one of the main inoculum sources of the disease on adjacent vines. There is evidence that this special form of F. solani actually represents a distinct species pathogenic to black pepper (2). References: (1) F. C. Albuquerque and S. Ferraz. Experientiae 22:133, 1976. (2) K. O'Donnell. Mycologia 92:919, 2000. (3) J. A. Ventura et al. Fitopatol. Bras. 11:361, 1986.

Effectiveness of the immunomodulatory extract of Kalanchoe pinnata against murine visceral leishmaniasis.

Previously, we described the protective action of the immunomodulatory extract of Kalanchoe pinnata (Kp) in murine and human cutaneous leishmaniasis. In the present study, we investigated the effectiveness of Kp against visceral leishmaniasis, using the BALB/c mouse model of infection with Leishmania chagasi. Mice receiving oral daily doses of Kp (400 mg/kg) for 30 days displayed significantly reduced hepatic and splenic parasite burden, when compared with untreated animals. Protectiveness was accompanied by a reduction in parasite-specific IgG serum levels, and impaired capacity of spleen cells to produce IL-4, but not IFN-gamma and nitric oxide upon antigen recall in vitro. The reference drug Pentostam (72 mg/kg) given by the intra-peritoneal route on alternate days produced an anti-leishmanial effect similar to oral Kp. Our findings show that the oral efficacy of Kp, seen previously in murine cutaneous leishmaniasis, extends also to visceral leishmaniasis caused by L. chagasi, a difficult to treat and lethal disease of man.

Chromosome imbalances in syndromic hearing loss.

The cause of hearing impairment has not been elucidated in a large proportion of patients. We screened by 1-Mb array-based comparative genomic hybridization (aCGH) 29 individuals with syndromic hearing impairment whose clinical features were not typical of known disorders. Rare chromosomal copy number changes were detected in eight patients, four de novo imbalances and four inherited from a normal parent. The de novo alterations define candidate chromosome segments likely to harbor dosage-sensitive genes related to hearing impairment, namely 1q23.3-q25.2, 2q22q23, 6p25.3 and 11q13.2-q13.4. The rare imbalances also present in normal parents might be casually associated with hearing impairment, but its role as a predisposition gene remains a possibility. Our results show that syndromic deafness is frequently associated with chromosome microimbalances (14-27%), and the use of aCGH for defining disease etiology is recommended.

Patients with nephrolithiasis and blood hypertension have higher calciuria than those with isolated nephrolithiasis or hypertension?

AIM: The aim of this study was to determine urinary excretion of calcium, uric acid and sodium and to evaluate insulin resistance in patients with nephrolithiasis and blood hypertension, isolated and in association, and in healthy controls, in absence of obesity and diabetes. METHODS: The study included 83 non-obese or diabetic patients: 17 with nephrolithiasis and hypertension (group D); 25 with nephrolithiasis (group C); 17 with hypertension (group B) and 24 healthy controls (group A). Urinary analysis was done in 24-hour urine collection and insulin resistance was evaluated through the HOMA-IR index. RESULTS: Calciuria was higher in group D in relation to groups A (P<0.01), B (P<0.01) and C (P=0.01). There was no significant difference between groups A and B (P=0.32), A and C (P=0.10) and B and C (P=0.68). Correlation analysis between urinary calcium detected strong correlation with uric acid in group A, regular in groups B and C and, strong with sodium in groups B and C. No differences were detected in uric acid and sodium excretion or insulin resistance among groups. CONCLUSIONS: Patients with blood hypertension and nephrolithiasis present higher calciuria than healthy people, with hypertension or with lithiasis and do not have the positive correlation observed in these latter groups with renal excretion of uric acid and sodium. These results suggest that impaired renal calcium reabsorption in non-obese or diabetic individuals is involved in the association between hypertension and urolithiasis.

Influence of the magnon-phonon relaxation in the magnon transport under thermal gradient in yttrium iron garnet.

We investigate the influence of the magnon-phonon relaxation processes in the magnon transport under thermal gradient in the ferrimagnetic insulator yttrium iron garnet (YIG). Based on the Boltzmann equation we calculate the magnon contribution in the thermal conductivity in YIG and the longitudinal spin Seebeck effect in YIG/Pt films, including the influence of the magnon relaxation to the lattice through a phenomenological damping parameter. Our results are in good agreement with reported experimental data showing that besides the magnon-magnon relaxation processes, the magnon-phonon relaxation plays an important role in the thermal properties in YIG films.

Immunomodulatory pretreatment with Kalanchoe pinnata extract and its quercitrin flavonoid effectively protects mice against fatal anaphylactic shock.

Previously, we reported the immunosuppressive action of the aqueous extract of Kalanchoe pinnata (Kp) in mice. In the present study, we report on the protective effect of Kp in fatal anaphylactic shock, likewise a Th2-driven immunopathology, and the identification of its active component. Mice daily treated with oral Kp during hypersensitization with ovalbumin were all protected against death when challenged with the allergen, as compared with the 100% mortality in the untreated group. A single intraperitoneal dose 3 h prior to challenge was partially effective. Oral protection was accompanied by a reduced production of OVA-specific IgE antibodies, reduced eosinophilia, and impaired production of the IL-5, IL-10 and TNF-alpha cytokines. In vitro, Kp prevented antigen-induced mast cell degranulation and histamine release. Oral treatment with the quercitrin flavonoid isolated from Kp prevented fatal anaphylaxis in 75% of the animals. These findings indicate that oral treatment with Kp effectively downmodulates pro-anaphylactic inducing immune responses. Protection achieved with quercitrin, although not maximal, suggests that this flavonoid is a critical component of Kp extract against this extreme allergic reaction.

Genomic imbalances pinpoint potential oncogenes and tumor suppressors in Wilms tumors.

BACKGROUND: Wilms tumor (WT) has a not completely elucidated pathogenesis. DNA copy number alterations (CNAs) are common in cancer, and often define key pathogenic events. The aim of this work was to investigate CNAs in order to disclose new candidate genes for Wilms tumorigenesis. RESULTS: Array-CGH of 50 primary WTs without pre-chemotherapy revealed a few recurrent CNAs not previously reported, such as 7q and 20q gains, and 7p loss. Genomic amplifications were exclusively detected in 3 cases of WTs that later relapsed, which also exhibited an increased frequency of gains affecting a 16.2 Mb 1q21.1-q23.2 region, losses at 11p, 11q distal, and 16q, and WT1 deletions. Conversely, aneuploidies of chromosomes 13 and 19 were found only in WTs without further relapse. The 1q21.1-q23.2 gain associated with WT relapse harbours genes such as CHD1L, CRABP2, GJA8, MEX3A and MLLT11 that were found to be over-expressed in WTs. In addition, down-regulation of genes encompassed by focal deletions highlighted new potential tumor suppressors such as CNKSR1, MAN1C1, PAQR7 (1p36), TWIST1, SOSTDC1 (7p14.1-p12.2), BBOX and FIBIN (11p13), and PLCG2 (16q). CONCLUSION: This study confirmed the presence of CNAs previously related to WT and characterized new CNAs found only in few cases. The later were found in higher frequency in relapsed cases, suggesting that they could be associated with WT progression.

In vitro anti-rotavirus activity of some medicinal plants used in Brazil against diarrhea.

Acute diarrhea, especially in children, is a very common disease with worldwide distribution and with a significant public health impact. Rotaviruses have been recognized as the major agents of diarrhea in infants and young children in developed as well as developing countries. In Brazil, diarrhea is one of the principal causes of death, mainly in the infant population. To fight diarrhea, traditional Brazilian medicine uses a great variety of plants. In this work, 12 medicinal plant species were screened for simian (SA-11) and human (HCR3) rotaviruses inhibition in vitro. At non-cytotoxic concentrations, the extracts from Artocarpus integrifolia L. (Moraceae) bark (480 microg/ml) and Spondias lutea L. (Anacardiaceae) leaves (160 microg/ml) had antiviral activity against both viruses. They showed inhibition of 99.2% and 97%, respectively, for human rotavirus, and 96.4% and 96.2% for simian rotavirus. The extracts from Myristica fragrans Houtt (Myristicaceae) seeds (160 microg/ml) and Spongias lutea bark (40 microg/ml) inhibited human rotavirus (90% and 82.2% inhibition, respectively), whereas the extracts from Anacardium occidentale L. (Anacardiaceae) leaves (4 microg/ml) and Psidium guajava L. (Myrtaceae) leaves (8 microg/ml) showed activity only against simian rotavirus (82.2% and 93.8% inhibition, respectively). Our results indicate that the extracts of Artocarpus integrifolia, Myristica fragrans and Spongias lutea can be useful in the treatment of human diarrhea if the etiologic agent is a rotavirus.

Group A streptococcal isolate 64/14 expresses surface plasmin-binding structures in addition to Plr.

A recombinant plasmin receptor (Plr) gene product originally cloned from group A streptococcal isolate 64/14 was analysed for its ability to bind plasmin(ogen) and to account for all the surface plasmin-binding properties of streptococcal isolate 64/14. Functional analysis of recombinant Plr demonstrated that the protein exhibited equal reactivity with human Lys-plasmin and Lys-plasminogen, but significantly lower reactivity with Glu-plasminogen. Plasmin-binding was both inhibitable and elutable by lysine or lysine analogs, and active plasmin bound to recombinant Plr was not neutralized by alpha 2-antiplasmin. Thus, the plasmin-binding properties of recombinant Plr correlated with the plasmin-binding phenotype of the intact streptococcal isolate 64/14. In addition, fluid-phase recombinant Plr could completely inhibit binding of plasmin to either immobilized recombinant Plr or group A streptococcal isolate 64/14 with equal efficiency, indicating that surface-expressed Plr could account for all the plasmin-binding properties of the intact organism. An IgM monoclonal antibody to recombinant Plr that specifically recognized a surface structure on streptococcal isolate 64/14 significantly inhibited the binding of plasmin to the recombinant protein; however, the antibody was not successful at inhibiting plasmin-binding to the intact bacteria, indicating the presence of other plasmin-binding structures on the bacterial surface in addition to Plr.

FRAXA premutation associated with premature ovarian failure.

A family is described in which six females in three generations experienced premature ovarian failure (POF). In three of them a FRAXA premutation was documented and the carrier status of a fourth female could be inferred, because her son had the fragile X syndrome. These findings provide further evidence for a nonrandom association between POF and the FRAXA premutation.

DXS548/FRAXAC1 haplotypes in fragile X chromosomes in the Brazilian population.

In order to investigate the origin of the fragile X mutation in the Brazilian population, we assessed the size of the microsatellite markers DXS548, FRAXAC1 and FRAXAC2 in 72 X chromosomes from unrelated affected males and 64 control chromosomes. We found a significantly different distribution of alleles between fragile X and controls for loci DXS548 and FRAXAC1, but no apparent linkage disequilibrium was detected for the sequence FRAXAC2. The most frequent DXS548/FRAXAC1 haplotypes in affected males were haplotypes 204/158 bp (2-1) and 196/152 bp (6-4). These findings are in accordance with the proposed two main mutational pathways for the generation of FMR-1 alleles that predispose to instability and hyperexpansion.