Genomic profiling reveals distinct routes to complement resistance in Klebsiella pneumoniae

The serum complement (C') system is a first line of defense against bacterial invaders. Resistance to killing by serum enhances the capacity of Klebsiella pneumoniae to cause infection, but is an incompletely understood virulence trait. Identifying and characterising the factors responsible for preventing activation of, and killing by, serum C' could inform new approaches to treatment of K. pneumoniae infections. We have used functional genomic profiling to define the genetic basis of C' resistance in four diverse serum-resistant K. pneumoniae strains (NTUH-K2044, B5055, ATCC43816 and RH201207), and explored their recognition by key complement components. Over 90 genes contributed to resistance in one or more strains, but only three, rfaH, lpp and arnD, were common to all four. Deletion of the anti-terminator rfaH, controlling expression of capsule and O-side chains, resulted in dramatic C' resistance reductions in all strains. The murein lipoprotein gene lpp promoted capsule retention through a mechanism dependent on its C-terminal lysine residue; its deletion led to modest reductions in C' resistance. Binding experiments with the C' components C3b and C5b-9 showed that the underlying mechanism of evasion varied in the four strains: B5055 and NTUH-K2044 appeared to bypass recognition by C' entirely, while ATCC43816 and RH201207 were able to resist killing despite being associated with substantial levels of C5b-9. All rfaH and lpp mutants bound C3b and C5b-9 in large quantities. Our findings show that, even amongst this small selection of isolates, K. pneumoniae adopts differing mechanisms and utilises distinct gene sets to avoid C' attack

Estimativa de populações canina e felina domiciliadas em zona urbana do Estado de São Paulo Estimate of the owned canine and feline populations in urban area in Brazil

OBJETIVO: Dada a importância do conhecimento acerca da população de cães e gatos domiciliados para o adequado planejamento e avaliação das ações de controle da raiva em áreas urbanas, propõe-se a utilização de um valor preciso de estimativa dessas populações com base em indicadores populacionais humanos. MÉTODOS: Foi calculada a razão entre as populações humana e animal (canina e felina) por meio de inspeções domiciliares, no Município de Taboão da Serra, SP. O município foi dividido em duas áreas homogêneas distintas socioeconomicamente, de acordo com o algoritmo da média k, de modo a permitir a comparação das razões homem:animal (cão e gato) das áreas homogêneas. RESULTADOS: A razão entre a população humana e a população canina foi 5,14 e a entre a humana e a felina foi 30,57. Não foi observada diferença significativa ao comparar-se as razões entre as populações humana e animal das áreas homogêneas. CONCLUSÕES: A adoção de uma metodologia de estimativa populacional canina e felina domiciliada, baseada em indicadores populacionais humanos, é a mais indicada e facilmente exeqüível quando comparada ao censo canino.<br>OBJECTIVE: Given the importance of assessing owned dog and cat populations to adequate planning and evaluation of rabies control measures in urban areas, it is proposed the use of an estimate of these populations based on human population parameters. METHODS: The ratio between human population and owned animal (dogs and cats) population was calculated in the municipality of Taboão da Serra, state of São Paulo, Brazil. This municipality was divided into two distinct social and economic homogeneous areas through k-mean algorithm, allowing for comparison between the ratios of the two homogeneous areas. RESULTS: A 5.14 ratio was calculated for human and dog populations and a 30.57ratio for human and feline populations. A significant difference was not observed when comparing the ratios for human and animal populations of the two homogeneous areas. CONCLUSIONS: An estimate of owned dog and cat populations based on the human population is the better choice for assessing the animal population instead of an animal census because it can be easily implemented

Stabilization and Crystallization of a Membrane Protein Involved in Lipid Transport

Lipoteichoic acids (LTA) are ubiquitous cell wall components of Gram-positive bacteria. In Staphylococcus aureus LTA are composed of a polymer with 1,3-linked glycerol phosphate repeating units anchored to the plasma membrane. The anchor molecule is a lipid-linked disaccharide (anchor-LLD) synthesized at the cytoplasmic leaflet of the membrane. The anchor lipid becomes accessible at the outer leaflet of the membrane after the flippase LtaA catalyzes translocation. Recently we have elucidated the structure of LtaA using vapor diffusion X-ray crystallography and in situ annealing. We were able to obtain LtaA crystals after optimization of purification protocols that led to stabilization of LtaA isolated in detergent micelles. Here we report a protocol that describes the purification, stabilization, crystallization, and data collection strategies carried out to determine the structure of LtaA. We highlight key points that can be used to determine crystal structures of other membrane proteins