Biosurfactant production of Piper hispidum endophytic fungi.

AIMS: To evaluate the production of biosurfactants by fungi isolated from the Amazonian species Piper hispidum (Piperaceae), and to determine the physico-chemical properties of the crude biosurfactant obtained from the most promising fungi. METHODS AND RESULTS: A total of 117 endophytic fungi were isolated, and 50 were used to verify the production of biosurfactants. Of these, eight presented positive results in the drop collapse test, and emulsification index ranging from 20 to 78%. The most promising fungi, Ph III 23L and Ph II 22S (identified as Aspergillus niger and Glomerella cingulata, respectively) were recultivated for extraction and analysis of the biosurfactant's physico-chemical characteristics. The cultivation broth that presented the greatest decrease in surface tension (36%) was that of the A. niger, which reduced it from 68·0 to 44·0 mN m-1 . The lowest critical micellar concentration value was found for the same endophyte (14·93 mg ml-1 ). CONCLUSIONS: Endophytes of P. hispidum proved to be interesting producers of biosurfactants and presented promising physico-chemical characteristics for applications in diverse industrial sectors. SIGNIFICANCE AND IMPACT OF THE STUDY: Piper hispidum endophytic fungi can be used as a new source of biosurfactants, as these molecules present a significant market due to their wide industrial applications.

Expressão e caracterização da glicoproteína D do herpesvírus equídeo 1 em Pichia pastoris / Expression and characterization of equid herpesvirus 1 glycoprotein D in Pichia pastoris

O herpesvírus equídeo 1 (EHV-1) apresenta distribuição mundial e causa graves prejuízos à equideocultura. É agente de surtos de doença respiratória, reprodutiva e neurológica, em equídeos jovens e adultos. A glicoproteína D (gD) do envelope viral é essencial para ligação e penetração em células permissivas e direcionamento do sistema imunológico do hospedeiro, induz respostas imunes humorais e celulares, sendo um antígeno apropriado para ser utilizado em vacinas e imunodiagnóstico. O objetivo deste trabalho foi expressar e caracterizar a gD do EHV-1 em Pichia pastoris para posterior utilização como antígeno em técnicas de imunodiagnóstico e formulação de vacinas recombinantes. Uma sequência de DNA que codifica uma forma truncada da gDEHV-1 foi clonada no vetor pPICZαA de expressão em P. pastoris. Obteve-se uma proteína de ~41 kDa, como esperado. A proteína apresentou glicosilação entre 4 kDa e 16 kDa, demonstrada por deglicosilação enzimática. A proteína recombinante foi caracterizada antigenicamente e imunogenicamente por Western blot, utilizando-se anticorpos policlonais equinos anti-EHV-1, e por ELISA indireto em modelo murino, demonstrando que a gD recombinante manteve epítopos similares aos da proteína nativa. Esses resultados sugerem que a gDEHV-1 é um antígeno promissor para uso como imunobiológico no controle do EHV-1.(AU)

Equine herpesvirus 1 (EHV-1) has a worldwide distribution and causes serious damage to horse breeding. It is an agent of respiratory, reproductive and neurological disease outbreaks in young and adult equids. Viral envelope glycoprotein D (gD) is essential for binding and penetration into permissive cells and targeting the host immune system, inducing humoral and cellular immune responses, and is an appropriate antigen for use in vaccines and immunodiagnostics. The objective of this work was to express in Pichia pastoris and to characterize EHV-1 gD for later use as an antigen in immunodiagnostic techniques and formulation of recombinant vaccines. A DNA sequence encoding a truncated form of gDEHV-1 has been cloned into the P. pastoris expression vector pPICZαA. A protein of ~41 kDa was obtained as expected. The protein presented glycosylation between 4 kDa and 16 kDa, demonstrated by enzymatic deglycosylation. The recombinant protein was antigenically and immunogenically characterized by Western blot using equine polyclonal anti-EHV-1 antibodies, and by indirect ELISA in a murine model, demonstrating that the recombinant gD maintained epitopes similar to those of the native protein. These results suggest that gDEHV-1 is a promising antigen for use as an immunobiological in the control of EHV-1.(AU)

Cardosins in postembryonic development of cardoon: towards an elucidation of the biological function of plant aspartic proteinases.

Following on from previous work, the temporal and spatial accumulation of the aspartic proteinases (EC 3.4.23) cardosin A and cardosin B during postembryonic seed development of cardoon (Cynara cardunculus) was studied, mRNA and protein analyses of both cardosins suggested that the proteins accumulate during seed maturation, and that cardosin A is later synthesised de novo at the time of radicle emergence. Immunocytochemistry revealed that the precursor form of cardosin A accumulates in protein bodies and cell walls. This localisation in seeds is different from that previously described for cardoon flowers, suggesting a tissue-dependent targeting of the protein. It is known that procardosins are active and may have a role in proteolysis and processing of storage proteins. However, the presence of procardosin A in seeds could be related to the proposed role of the plant-specific insert in membrane lipid conversion during water uptake and solute leakage in actively growing tissues. This is in accordance with the recently proposed bifunctional role of aspartic proteinase precursor molecules that possess a membrane-destabilising domain in addition to a protease domain. Mature cardosin B, but not its mRNA, was detected in the first hours after seed imbibition and disappeared at the time of radicle emergence. This extracellular aspartic protease has already been implicated in cell wall loosening and remodelling, and its role in seed germination could be related to loosening tissue constraints for radicle protusion. The described pattern of cardosin A and B expression suggests a finely tuned developmental regulation and prompts an analysis of their possible roles in the physiology of postembryonic development.

Risk factors associated with death in children admitted to a paediatric intensive care unit.

The aim of this study was to identify fatal risk factors for children admitted to the paediatric intensive care unit of the Instituto Materno Infantil de Pernambuco, a referral hospital in Recife, a city in the north-east of Brazil. A survey was performed from June 1996 to January 1997. Risk was quantified by the crude and adjusted odds ratio. The 95 per cent confidence interval, likelihood ratio statistics, and the probability (p < 0.05) value were used to test for statistical significance. An association was established between death in children admitted to the paediatric intensive care unit and: (1) age below 2 years old; (2) use of mechanical ventilation and central venous catheter; (3) presence of hospital-acquired infection; (4) length of hospital stay of 2 days or less; and (5) Class 4 clinical severity, according to the Clinical Classification System (CCS). These results allow the identification of the children with a greater risk of death and may contribute to improvements in handling groups of patients with poor prognosis.

Esquistossomose murina esperimental: estudo imunohistoquímicos das células ganglionares mioentéricas / Experimental murine Schistosomiasis: an immunohistochemical study of the ganglionic myoenteric cells

Para estudar a influência do processo granulomatoso esquistossomóticosobre as células ganglionares mioentéricas, foram utilizados 30 camundongos albinos Swiss infectadoscom 50 cercárias da cepa SLM do S. mansoni.O grupo controle foi constituído por dez animais näo infectados. Após sessenta dias de infecçäo, cortes histológicos do intestino delgadocorados por hematoxilina-eosina e P.A.S. demonstraram granulomas periovulares em todas as camadas da parede intestinal. Através do método imunohistoquímico indireto, usando-se a enolase neurônio-específica como marcador, observou-se desorganizaçäo do plexo mioentêrico em áreas contendo granulomas. Além disso, ocorreu rarefaçäo das estaçöes ganglionares, com aparente destruiçäo de células neuronais. A possível contribuiçäo dessas alteraçöes para a sintomatologia da esquistossomose humana é avaliada