flu
H,K-ATPase alpha subunit C-terminal membrane topology: epitope tags in the insect cell expression system.

Smolka AJ, Larsen KA, Schweinfest CW, Hammond CE.

Division of Gastroenterology and Hepatology, Department of Medicine, CSB 916, Medical University of South Carolina, 171 Ashley Avenue, Charleston, SC 29425, USA. smolkaaj musc.edu

The H,K-ATPase responsible for gastric acidification is a heterodimeric (alpha and beta subunit) P-type ATPase, an integral protein of parietal cell apical membranes, which promotes the electroneutral exchange of K+ for protons, is stimulated by K+ and is inhibited by 2-methyl-8-(phenylmethoxy)imidazo[1, 2-alpha]pyridine-3-acetonitrile (SCH 28080). Hydropathy analysis of the catalytic alpha subunit has been interpreted in terms of four N-terminal transmembrane domains, a cytoplasmically oriented segment containing ATP binding and phosphorylation sites, and a C-terminal region with four or six putative transmembrane domains. Several lines of evidence implicate the C-terminal region of P-type ATPases in cation-binding and occlusion, conformational changes, and interactions with the beta subunit (HKbeta), making the definition of topology a prerequisite for understanding the structural basis of these functions. Influenza haemagglutinin epitopes (YPYDVPDYA; flu tag) were inserted in predicted hydrophilic segments of the alpha subunit (HKalpha) to establish the membrane orientation of two amino acids with different predicted topologies in the C-terminal four- and six-transmembrane models. Wild-type and mutated HKalpha and HKbeta cDNA species were expressed in insect cells (Sf9) via recombinant baculovirus infection, and expression of H,K-ATPase was verified by immunoblotting with HKalpha- and HKbeta-specific and flu-tag-specific antibodies. Functional assays showed K+-stimulated, SCH 28080-sensitive ATPase activity, confirming neo-native topology in H,K-ATPase heterodimers expressed in Sf9 cells. The topology of flu tags was determined by microsomal protease protection assays in Sf9 cells and immunolabelling of HKalpha and HKbeta in intact and permeabilized Sf9 cells. In addition, MS of native H,K-ATPase tryptic peptides identified cytoplasmically oriented HKalpha residues. The results indicated cytoplasmic exposure of Leu844 and Phe996, and luminal exposure of Pro898, leading to a revised secondary structure model of the C-terminal third of HKalpha.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=10359643&dopt=Abstract flu, flu medicine, tamiflu



flu
[Impact of various strategies on the rates of flu vaccination in the elderly]

[Article in Spanish]

Puig-Barbera J, Ors Zarzoso P, Vilchez Pena C, Lloria Paes F.

Unidad de Programas de Promocion de Salud, Centro de Salud Publica de Castellon, Conselleria de Sanitat, Generalitat Valenciana. juan.puig sanidad.m400.gva.es

OBJECTIVE: To calculate the impact of different interventions on the rates of anti-flu vaccination in people aged 65 and over. DESIGN: Ecological study with analytical components. SETTING: Health centres and clinics in two health areas in the Community of Valencia. PATIENTS AND OTHER PARTICIPANTS: Care units, characterised by their coverage of stable nuclei of 500 or more people, total population of 415,172 inhabitants. INTERVENTIONS: Use of the communication media, personal invitations by post and telephone; follow-up of activity, discussion of results, external evaluations; registration of people to be vaccinated and prior order to nursing staff to vaccinate. MEASUREMENTS AND MAIN RESULTS: The coverage in quartiles of the previous season and the human resources per thousand inhabitants were the structural factors which best explained the vaccination rates. In the multivariate analysis, invitation by letter had relative risk (RR) of 1.062 (95% CI, 1.036-1.088); telephone call, RR 1.075 (95% CI, 1.021-1.132); discussion of external evaluations, RR 1.046 (95% CI, 1.024-1.068); and the order of vaccination to nursing staff, RR 1.056 (95% CI, 1.025-1.088). All these were associated with greater coverage. The absolute difference in coverage achieved between the care units which invited people to be vaccinated by letter and phone, at which nurses vaccinated directly, and which discussed external evaluations, and the units which did not perform these above activities, was 14 points (95% CI, 13.3-14.5), independently of the other factors. CONCLUSIONS: The previous year's coverage, human resources, invitation by letter or phone, the order to nursing to vaccinate and the discussion of external evaluations, were all associated, independently, with higher rates of vaccination.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=10372456&dopt=Abstract flu, flu medicine, tamiflu



flu
Cold/flu knowledge, attitudes and health care practices: results of a two-city telephone survey.

Vingilis ER, Brown U, Sarkella J, Stewart M, Hennen BK.

Population and Community Health Unit, Faculty of Medicine and Dentistry, University of Western Ontario, London. evelyn.vingilis med.uwo.ca

The purpose of this paper is to describe knowledge, attitudes and practices of cold and flu self-care and health care utilization, and to identify the predictors of health care utilization for the cold and flu among residents of London and Windsor. Using a random digit dialing survey method, 417 residents were interviewed between November-December, 1993 and February-March, 1994. This survey revealed good knowledge about colds and flu and understanding of appropriate physician visits. Only seven percent reported a doctor visit for their last cold. Socio-demographic, health status, attitude and knowledge level variables were subjected to a logistic regression analysis to identify which variables predicted self-reported physician visits. Only attitudes and health status showed statistically significant log odds (3.6 and 1.5, respectively). In summary, consistent with other studies, attitude and health status, not knowledge, appear to be significant predictors of physician visits for colds/flu.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=10401174&dopt=Abstract flu, flu medicine, tamiflu



flu
Characterization of immune responses to experimental polyvalent subunit vaccines assembled in iscoms.

Sjolander A, Ahlborg N, Stahl S, Andersson R.

Department of Veterinary Virology, National Veterinary Institute, Biomedicum, Uppsala, Sweden.

Immune responses to experimental polyvalent subunit vaccines assembled in a particulate adjuvant/delivery system, iscoms, are described. The fusion protein ZZ-M5 comprises structures of staphylococcal protein A (ZZ) and the Plasmodium falciparum malaria antigen Pf155/RESA (M5). MHC congenic mice were immunized with ZZ-M5 conjugated to iscoms containing human influenza virus antigen (flu ag, M5-flu-isc) or to iscom matrix (iscom particles without flu ag, M5-isc). Comparison of antibody and T-cell responses to M5-isc and M5-flu-isc demonstrated that the flu ag in M5-flu-isc exhibits carrier-related helper functions and that the assembly of immunogens in M5-flu-isc did not result in any apparent antigenic competition. In addition, assembly of ZZ-M5 and flu ag in iscoms induced an alteration of the IgG subclass profile of the antibody response to M5. The results suggest that assembly of immunogens in iscoms may be a useful approach to the design of subunit vaccines but that both quantitative and qualitative aspects of the immunogenic properties of such constructs should be scrutinized.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=9694516&dopt=Abstract flu, flu medicine, tamiflu



flu
Laboratory containment for influenza A H5N1 virus: level 2, level 3, or level 3+?

Zambon M.

Enteric and Respiratory Virus Laboratory, PHLS Central Public Health Laboratory, London. mzambon phls.co.uk

The recent alert over bird flu (influenza A H5N1) in Hong Kong has ruffled feathers in some countries, including the United Kingdom, as to how the virus should be handled in clinical and research laboratories.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=9718852&dopt=Abstract flu, flu medicine, tamiflu