Isolation and characterization of BliAI, an isoschizomer of ClaI from Bacillus licheniformis

The restriction endonuclease BliAI, an isoschizomer of ClaI, which recognizes the sequence 5'- AT¼CGAT - 3', was purified from a natural isolate identified as Bacillus licheniformis. The restriction endonuclease was isolated from cell extracts using single-step purification by phosphocellulose column chromatography. The restriction endonuclease is active at 37°C and over a wide range of pH and salt concentration. The molecular weight of the purified restriction enzyme is consistent with a value of 39 kDa.

A endonuclease de restrição BliAI, um isoesquisômero de ClaI, que reconhece a seqüência 5'- AT¼CGAT - 3', foi purificada de um isolado natural identificado como Bacillus licheniformis. A endonuclease de restrição em questão foi isolada a partir de um extrato celular em um único passo cromatográfico utilizando uma coluna contendo a resina fosfocelulose. A endonuclease de restrição é ativa à 37°C e em uma ampla escala de pH e concentração de sais. O peso molecular da enzima purificada corresponde a um valor de kDa 39.

BpuAmI: a novel SacI neoschizomer from Bacillus pumilus discovered in an isolate from Amazon Basin, recognizing 5'-GAG FONT FACE=Symbol>¯ /FONT>CTC-3'

A strain of Bacillus pumilus was isolated and identified from water samples collected from a small affluent of the Amazon River. Type II restriction endonuclease activity was detected in these bacteria. The enzyme was purified and the molecular weight of the native protein estimated by gel filtration and SDS-PAGE. The optimum pH, temperature and salt requirements were determined. Quality control assays showed the complete absence of "nonspecific nucleases." Restriction cleavage analysis and DNA sequencing of restriction fragments allowed the unequivocal demonstration of 5´GAG FONT FACE=Symbol>¯ /FONT>CTC3´ as the recognition sequence. This enzyme was named BpuAmI and is apparently a neoschizomer of the prototype restriction endonuclease SacI. This is the first report of an isoschizomer and/or neoschizomer of the prototype SacI identified in the genus Bacillus.

Uma linhagem de Bacillus pumilus foi isolada e identificada de amostras de águas coletadas em um pequeno Igarapé do Rio Amazonas. Foi detectada atividade de restrição do tipo II nesta bactéria. A enzima foi purificada e o peso molecular da proteína nativa foi estimado por gel filtração e por eletroforese em gel de poliacrilamida. Foram determinados, o pH e temperatura ótimos e as necessidades de sais. Os ensaios do controle de qualidade mostraram uma ausência completa de "nucleases não específicas". As analises das clivagens e o seqüenciamento do DNA dos fragmentos de restrição permitiram uma demonstração inequívoca de que 5´GAG FONT FACE=Symbol>¯ /FONT>CTC 3´ é a seqüência de reconhecimento da enzima. Esta enzima foi denominada de BpuAmI e aparentemente é um neoesquisômero da enzima protótipo SacI. Este é o primeiro relato de um isoesquisômero e/ou neoesquisômero da enzima protótipo SacI identificada no gênero Bacillus.

BpuAmI: a novel SacI neoschizomer from Bacillus pumilus discovered in an isolate from Amazon Basin, recognizing 5'-GAGü®CTC-3'

Uma linhagem de Bacillus pumilus foi isolada e identificada de amostras de águas coletadas em um pequeno Igarapé do Rio Amazonas. Foi detectada atividade de restrição do tipo II nesta bactéria. A enzima foi purificada e o peso molecular da proteína nativa foi estimado por gel filtração e por eletroforese em gel de poliacrilamida. Foram determinados, o pH e temperatura ótimos e as necessidades de sais. Os ensaios do controle de qualidade mostraram uma ausência completa de "nucleases não específicas". As analises das clivagens e o seqüenciamento do DNA dos fragmentos de restrição permitiram uma demonstração inequívoca de que 5üLGAG¼CTC 3üL é a seqüência de reconhecimento da enzima. Esta enzima foi denominada de BpuAmI e aparentemente é um neoesquisômero da enzima protótipo SacI. Este é o primeiro relato de um isoesquisômero e/ou neoesquisômero da enzima protótipo SacI identificada no gênero Bacillus.

Resultados preliminares de um projeto sobre a ecologia dos flebotomíneos vetores de leishmaniose tegumenar no estado da Bahia / Preliminary results of a project on the ecology of the cutaneous leishmaniasis vectors phlebotominae in the State of Bahia

Preliminary data of a project about the ecology of the Phlebotominae (sandflies) in a tegumentary leishmaniasis area of coconut plantation in the south of the State of Bahia, Brazil are presented. There are 60 dwellings far of one another, where existed 31 dogs and 229 inhabitants. Among them 41.5 per cent were Montenegro positive; 37.5 per cent from these had scars of healed ulcers and 8.8 per cent had active ulcers. 53 per cent of the house had dogs from which 22 per cent were seropositive; from the 7 dogs with ulcers only 3 were seropositive. 14 per cent of the inspected house harboured sandflies inside them. During two years of observations 72 sentinels hamsters were maintened at houses of patients with leishmaniasis ulcers, but they did not get the infection. During two years of observation, monthly collectings of sandflies were made inside house, chicken pen, curral, tree trunks and open field around a house with a patient with leishmaniasis. The following results were already obtained about the vectors: 5,614 specimens were identified as belonging to fourteen different species of sandflies. Among them Lutzomyia whitmani (92 per cent) and Lutzomyia intermedia (4.8 per cent) were the most abundant species. They are very anthropophilic and can be found inside houses and at peridomestic sites. Probably they are the principal vectors of the disease at domestic places. The other twelve species were less frequent and more found at sylvatic places in inspite they also bite the man. Generally the biting activity of most of the species of the area begins at 5 p.m. in the dusk and reaches its peak at 0 hour a.m., when begins declining until disapear at 7 a.m. L. whitmani was similarly collected with the same density in all lunar phases while L. intermedia was more abundant during the new moon phase. Most of the hundreds sandflies collected during the second year of observations, remains preserved in liquid nitrogen, watching for the adjustment of PCR molecular techniques to be processed for determination of the vector natural infection rates with leishmanias. Final results on all the project will be published as soon as the examination of such material has been processed.