Mycobacterium tuberculosis lineage: a naming of the parts

There have been many reports of groups of related Mycobacterium tuberculosis strains described variously as lineages, families or clades. There is no objective definition of these groupings making it impossible to define relationships between those groups with biological advantages. Here we describe two groups of related strains obtained from an epidemiological study in Tanzania which we define as the Kilimanjaro and Meru lineages on the basis of IS6110 restriction fragment length polymorphism (RFLP), polymorphic GC rich sequence (PGRS) RFLP and mycobacterial interspersed repeat unit (MIRU) typing. We investigated the concordance between each of the typing techniques and the dispersal of the typing profiles from a core pattern. The Meru lineage is more dispersed than the Kilimanjaro lineage and we speculate that the Meru lineage is older. We suggest that this approach provides an objective definition that proves robust in this epidemiological study. Such a framework will permit associations between a lineage and clinical or bacterial phenomenon to be tested objectively. This definition will also enable new putative lineages to be objectively tested

Primary screening of blood donors by nat testing for HCV-RNA: development of an "in-house" method and results Triagem primária de doadores de sangue por teste de ácidos nucléicos: desenvolvimento de um método não-comercial e resultados

An "in-house" RT-PCR method was developed that allows the simultaneous detection of the RNA of the Hepatitis C Virus (HCV) and an artificial RNA employed as an external control. Samples were analyzed in pools of 6-12 donations, each donation included in two pools, one horizontal and one vertical, permitting the immediate identification of a reactive donation, obviating the need for pool dismembering. The whole process took 6-8 hours per day and results were issued in parallel to serology. The method was shown to detect all six HCV genotypes and a sensitivity of 500 IU/mL was achieved (95% hit rate). Until July 2005, 139,678 donations were tested and 315 (0.23%) were found reactive for HCV-RNA. Except for five false-positives, all 310 presented the corresponding antibody as well, so the yield of NAT-only donations was zero, presenting a specificity of 99.83%. Detection of a window period donation, in the population studied, will probably demand testing of a larger number of donations. International experience is showing a rate of 1:200,000 - 1:500,000 of isolated HCV-RNA reactive donations.<br>Desenvolveu-se uma metodologia própria ("in-house") baseada em RT-PCR, que permite detectar simultaneamente o RNA do vírus HCV e de um RNA artificial empregado como controle externo. As amostras são analisadas em pools de 6-12 doações, cada doação sendo incluída em dois pools diferentes, um horizontal e um vertical, permitindo a identificação imediata de uma doação reativa, sem a necessidade de desmembrar-se um pool reativo. O processo todo consumiu de 6-8 horas diárias e os resultados foram emitidos em paralelo à sorologia. O método detectou os seis genótipos de HCV, com um limite de sensibilidade de 500 UI/mL (95% hit rate). Até julho de 2005 haviam sido testadas 139.678 doações com a detecção de 315 (0,23%) doações reativas para HCV-RNA. Exceto cinco falso-positivas, todas estas doações também apresentavam o respectivo anticorpo, portanto não se detectou nenhuma doação em janela imunológica. A especificidade foi de 99,83%. A detecção de amostra em janela imunológica, nesta população de doadores, provavelmente demandará a análise de um número maior de doações, espelhando-se na experiência internacional que tem mostrado a detecção de amostras HCV-RNA isoladas em 1:200.000 - 1:500.000 doações

Kar9p-independent Microtubule Capture at Bud6p Cortical Sites Primes Spindle Polarity before Bud Emergence in Saccharomyces cerevisiae

Spindle orientation is critical for accurate chromosomal segregation in eukaryotic cells. In the yeast Saccharomyces cerevisiae, orientation of the mitotic spindle is achieved by a program of microtubule–cortex interactions coupled to spindle morphogenesis. We previously implicated Bud6p in directing microtubule capture throughout this program. Herein, we have analyzed cells coexpressing GFP:Bud6 and GFP:Tub1 fusions, providing a kinetic view of Bud6p–microtubule interactions in live cells. Surprisingly, even during the G1 phase, microtubule capture at the recent division site and the incipient bud is dictated by Bud6p. These contacts are eliminated in bud6Δ cells but are proficient in kar9Δ cells. Thus, Bud6p cues microtubule capture, as soon as a new cell polarity axis is established independent of Kar9p. Bud6p increases the duration of interactions and promotes distinct modes of cortical association within the bud and neck regions. In particular, microtubule shrinkage and growth at the cortex rarely occur away from Bud6p sites. These are the interactions selectively impaired at the bud cortex in bud6Δ cells. Finally, interactions away from Bud6p sites within the bud differ from those occurring at the mother cell cortex, pointing to the existence of an independent factor controlling cortical contacts in mother cells after bud emergence