Identification of genetic alterations by multiple ligation-dependent probe amplification (MLPA) analysis in oligodendrogliomas

Concurrent deletion at 1p/19q is a common signature of oligodendrogliomas, and it may, be identified in low-grade tumours (grade II) suggesting it represents an early event in the development of these brain neoplasms. Additional non-random changes primarily involve CDKN2A, PTEN and EGFR. Identification of all of these genetic changes has become an additional parameter in the evaluation of the clinical patients` prognosis, including good response to conventional chemotherapy. Multiple ligation-dependent probe amplification (MLPA) analysis is a new methodology that allows an easy identification of the oligodendrogliomas` abnormalities in a single step. No need of the respective constitutional DNA from each patient is another advantage of this method. We used MLPA kits P088 and P105 to determine the molecular characteristics of a series of 40 oligodendrogliomas. Deletions at I p and 19q were identified in 45% and 65% of cases, respectively. Alterations of EGFR, CDKN2A, ERBB2, PTEN and TP53 were also identified in variable frequencies among 7% to 35% of tumours. These findings demonstrate that MLPA is a reliable technique to the detection of molecular genetic changes in oligodendrogliomas.

Identification of genes differentially expressed in a strain of the mold Aspergillus nidulans carrying a loss-of-function mutation in the palA gene

To identify genes differentially expressed in a strain of the mold Aspergillus nidulans carrying a loss-of-function mutation in palA, a gene in the pH-responsive signal transduction pathway, suppression subtractive hybridization was performed between RNA isolated from the biA1 and biA1 palA1 strains grown under limiting inorganic phosphate at pH 5.0. We have identified several genes upregulated in the biA1 palA1 mutant strain that play important roles in mitotic fidelity, stress responses, enzyme secretion, signal transduction mechanisms, development, genome stability, phosphate sensing, and transcriptional regulation among others. The upregulation of eight of these transcripts was also validated by Northern blot. Moreover, we show that a loss of function mutation in the palA gene drastically reduced the neutral sugar content of the acid phosphatase PacA secreted by the fungus A. nidulans grown at pH 5.0 compared with a control strain.

Pompe disease in a Brazilian series: clinical and molecular analyses with identification of nine new mutations

Pompe disease (glycogen storage disease type II or acid maltase deficiency) is an inherited autosomal recessive deficiency of acid alpha-glucosidase (GAA), with predominant manifestations of skeletal muscle weakness. A broad range of studies have been published focusing on Pompe patients from different countries, but none from Brazil. We investigated 41 patients with either infantile-onset (21 cases) or late-onset (20 cases) disease by muscle pathology, enzyme activity and GAA gene mutation screening. Molecular analyses identified 71 mutant alleles from the probands, nine of which are novel (five missense mutations c.136T > G, c.650C > T, c.1456G > C, c.1834C > T, and c.1905C > A, a splice-site mutation c.1195-2A > G, two deletions c.18_25del and c.2185delC, and one nonsense mutation c.643G > T). Interestingly, the c.1905C > A variant was detected in four unrelated patients and may represent a common Brazilian Pompe mutation. The c.2560C > T severe mutation was frequent in our population suggesting a high prevalence in Brazil. Also, eight out of the 21 infantile-onset patients have two truncating mutations predicted to abrogate protein expression. Of the ten late-onset patients who do not carry the common late-onset intronic mutation c.-32-13T > G, five (from three separate families) carry the recently described intronic mutation, c.-32-3C > A, and one sibpair carries the novel missense mutation c.1781G > C in combination with known severe mutation c.1941C > G. The association of these variants (c.1781G > C and c.-32-3C > A) with late-onset disease suggests that they allow for some residual activity in these patients. Our findings help to characterize Pompe disease in Brazil and support the need for additional studies to define the wide clinical and pathological spectrum observed in this disease.

Echocardiographic identification of the oblique vein of the left atrium: its relationship to the persistent left superior caval vein

Thus far, little has been written concerning echocarchographic identification of the oblique of the left atrium, or Marshall`s vein. There is much discussion, nonetheless, on the potential significance of the vein, or its ligamentous remnant, as an arrhythmic substrate. We describe here four patients in whom transthoracic echocardiography revealed a venous structure protruding within the cavity of the left atrium. We discuss the possibility that these structures represent Marshall`s vein, albeit probably as part of a persistent left superior caval vein.

Identification and characterization of a new member of snake venom thrombin inhibitors from Bothrops insularis using a proteomic approach

Snake venom C-type lectin-like proteins (CLPs) are ubiquitously found in Viperidae snake venoms and differ from the C-type lectins as they display different biological activities but no carbohydrate-binding activity. Previous analysis of the transcriptome obtained from the Bothrops insularis venom gland showed the presence of two clusters homologous to bothrojaracin (BJC) chains a and P. In an effort to identify a new BJC-like molecule, we used an approach associated with proteomic technologies to identify the presence of the expressed protein and then to purify and characterize a new thrombin inhibitor from B. insularis venom. We also constructed homology models of this protein and BJC, which were compared with other C-type lectin-like family members and revealed several conserved features of this intriguing snake venom toxin family. (C)0 2007 Elsevier Ltd. All rights reserved.

Short communication: Identification of subclinical cow mastitis pathogens in milk by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

Subclinical mastitis is a common and easily disseminated disease in dairy herds. Its routine diagnosis via bacterial culture and biochemical identification is a difficult and time-consuming process. In this work, we show that matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) allows bacterial identification with high confidence and speed (1 d for bacterial growth and analysis). With the use of MALDI-TOF MS, 33 bacterial culture isolates from milk of different dairy cows from several farms were analyzed, and the results were compared with those obtained by classical biochemical methods. This proof-of-concept case demonstrates the reliability of MALDI-TOF MS bacterial identification, and its increased selectivity as illustrated by the additional identification of coagulase-negative Staphylococcus species and mixed bacterial cultures. Matrix-assisted laser desorption-ionization mass spectrometry considerably accelerates the diagnosis of mastitis pathogens, especially in cases of subclinical mastitis. More immediate and efficient animal management strategies for mastitis and milk quality control in the dairy industry can therefore be applied.

A method for simultaneous detection and identification of Brazilian dog- and vampire bat-related rabies virus by reverse transcription loop-mediated isothermal amplification assay

At present, the sporadic occurrence of human rabies in Brazil can be attributed primarily to dog- and vampire bat-related rabies viruses. Reverse transcription loop-mediated isothermal amplification (RT-LAMP) was employed as a simultaneous detection method for both rabies field variants within 60 min. Vampire bat-related rabies viruses could be distinguished from dog variants by digesting amplicons of the RT-LAMP reaction using the restriction enzyme Alwl. Amplification and digestion could both be completed within 120 min after RNA extraction. In addition, the RI-LAMP assay also detected rabies virus in isolates from Brazilian frugivorous bats and Ugandan dog, bovine and goat samples. In contrast, there were false negative results from several Brazilian insectivorous bats and all of Chinese dog, pig, and bovine samples using the RI-LAMP assay. This study showed that the RT-LAMP assay is effective for the rapid detection of rabies virus isolates from the primary reservoir in Brazil. Further improvements are necessary so that the RT-LAMP assay can be employed for the universal detection of genetic variants of rabies virus in the field. (C) 2010 Elsevier B.V. All rights reserved.

Nymphs of the genus Amblyomma (Acari: Ixodidae) of Brazil: descriptions, redescriptions, and identification key

Together with the larval stage, the nymphal stage of ticks of the genus Amblyomma are the most aggressive ticks for humans entering areas inhabited by wildlife and some domestic animals in Brazil. However, due to the absence of morphological descriptions of the nymphal stage of most Brazilian Amblyomma species, plus the lack of an identification key, little or nothing is known about the life history of Amblyomma spp. nymphs in the country. In the present study, morphological description of the nymphal stage, illustrating important external characters through scanning electron microscopy, is provided for nymphs of 15 Amblyomma species that occur in Brazil, for which the nymphal stage had never been described: A. aureolatum, A. auricularium, A. calcaratum, A. coelebs, A. fuscum, A. humerale, A. incisum, A. latepunctatum, A. naponense, A. nodosum, A. ovate, A. pacae, A. pseudoconcolor, A. scalpturatum, A. varium. In addition, the nymphal stage of 12 Amblyomma species, which had been previously described, are redescribed: A. brasiliense, A. cajennense, A. dissimile, A. dubitatum, A. longirostre, A. oblongoguttatum, A. parked, A. parvum, A. romitii, A. rotundatum, A. tigrinum, A. triste. The descriptions and redescriptions totalized 27 species. Only 2 species (A. geayi, A. goeldii) out of the 29 Amblyomma species established in Brazil are not included in the present study. A dichotomous identification key is included to support taxonomic identification of the nymphal stage of 27 Amblyomma species established in Brazil. (C) 2010 Elsevier GmbH. All rights reserved.

The universal newborn hearing screening in Brazil: From identification to intervention

Objective: The objective of the study is to investigate the results of the newborn hearing screening program carried out in a Public Hospital in Brazil, in the first 3 years regarding: (1) the prevalence of hearing impairment; (2) the influence of the universal hearing screening program on the age at which the diagnosis of hearing loss is defined; (3) the cost effectiveness of the program; (4) the outcomes, in terms of the age in which the hearing rehabilitation started. Methods: A descriptive study of the first 3 years after starting the universal newborn hearing screening in a Public Hospital of Bauru, Sao Paulo state, Brazil. The screening method consists of a two-stage screening approach with transient otoacoustic emissions (TOAE), conducted by an audiologist. If the outcome in the second-stage screening is REFER, the infant is submitted to diagnostic follow-up testing and intervention at the Audiology and Speech Pathology Clinic at the University of Sao Paulo, campus of Bauru. The evaluation of the costs of the universal newborn hearing screening program per each screened newborn (around 4000/year) was done based on a proposal by the National Center for Hearing Assessment and Management, of the Utah State University, United States of America. Results: 11,466 newborns were submitted to hearing screening, corresponding to 90.52% of the living newborns. The prevalence of sensorineural hearing loss was 0.96:1000. Of the 11 children with sensorineural hearing loss, eight children received hearing aids and five started the therapeutic process before the age of 1. Currently, four children between the ages of 11 months and 2 years old were submitted to cochlear implant surgery. The cost of hearing screening was US$7.00 and the annual cost of the universal newborn hearing screening program was US$26,940.47. Conclusion: The hospital-based universal newborn hearing screening carried out through the Brazilian National Health System is viable, with promising results. However, in a country such as Brazil, which presents large socio-economic differences, the same type of analyses should be performed in several regions, so as to take into account specific aspects, to implement the newborn hearing screening along with the Public System. (C) 2010 Elsevier Ireland Ltd. All rights reserved.