Prevailing effectiveness of the 2009 influenza A(H1N1)pdm09 vaccine during the 2010/11 season in Sweden.

Sixty per cent of the Swedish population received the monovalent AS03-adjuvanted pandemic influenza vaccine in the autumn of 2009. We assessed the age-specific effectiveness of this pandemic vaccine against hospitalisation with laboratory-confirmed influenza A(H1N1)pdm09 during the season 2010/11, in the age group from six months to 64 years in Sweden. The screening method was applied to available surveillance data. Our results suggest a prevailing effectiveness of 72% (95% confidence interval (CI): 63­80%) with the highest effectiveness among children, six months to nine years-old (92%, 95%CI: 80­97%). However, there were limitations in data quality and study design due to the lack of systematic recording of administered vaccinations, which underline the importance of preparing for an evaluation when planning for large public health actions. Despite these limitations, we believe the results reflect true, high prevailing vaccine effectiveness. Indeed, there were fewer deaths caused by influenza and the impact of influenza on intensive care units was less severe during the 2010/11 season in Sweden than in countries with lower pandemic vaccination coverage. The association between the pandemic vaccine and narcolepsy has increased the importance of assessing the risks and benefits of the vaccination; studies on the effectiveness and the duration of protection are needed for this.

Isolation rooms for highly infectious diseases: an inventory of capabilities in European countries.

Isolation of patients with highly infectious diseases (HIDs) in hospital rooms with adequate technical facilities is essential to reduce the risk of spreading disease. The European Network for Infectious Diseases (EUNID), a project co-funded by European Commission and involving 16 European Union member states, performed an inventory of high level isolation rooms (HIRs, hospital rooms with negative pressure and anteroom). In participating countries, HIRs are available in at least 211 hospitals, with at least 1789 hospital beds. The adequacy of this number is not known and will depend on prevailing circumstances. Sporadic HID cases can be managed in the available HIRs. HIRs could also have a role in the initial phases of an influenza pandemic. However, large outbreaks due to natural or to bioterrorist events will need management strategies involving healthcare facilities other than HIRs.

EuroNHID checklists for the assessment of high-level isolation units and referral centres for highly infectious diseases: results from the pilot phase of a European survey.

Healthcare settings have been identified as preferential for the transmission of many agents causing highly infectious diseases (HIDs). Infection control procedures strongly reduce the risk of transmission of HIDs in hospital settings, when adequately applied. The main objective of the European Network for Highly Infectious Diseases (EuroNHID), a network co-funded by the European Commission, is to assess the current capabilities for dealing with HIDs in Europe, specifically in the context of infection control and healthcare worker (HCW) safety, through conducting an on-the-field survey of high-level isolation units (HLIUs)/referral centres for the management of HIDs in participating countries. During the first year of the project's activities, specifically designed, evidence-based checklists were developed. This review introduces the EuroNHID checklists as a standard tool for the assessment of hospital capabilities concerning infection control and HCW safety in the management of patients with HIDs, and presents preliminary results from five HLIUs.

European concepts for the domestic transport of highly infectious patients.

Highly infectious diseases involve clinical syndromes ranging from single to multiorgan infections and pose a constant threat to the public. In the absence of a definite treatment for most causative agents, patients benefit from maximum supportive care as clinical conditions may deteriorate in the short term. Hence, following initial case identification and isolation, rapid transportation to a specialized treatment unit must be considered in order to minimize the risk of secondary infections, but this is limited by available infrastructure, accessible care en route and the patient's clinical condition. Despite the development of consensus curricula for the clinical management of highly infectious patients, medical transportation lacks a common European approach. This article describes, as examples, three current European concepts for the domestic relocation of highly infectious patients by ground vehicles and aircraft with respect to national legislation and geography.

A variety of respiratory viruses found in symptomatic travellers returning from countries with ongoing spread of the new influenza A(H1N1)v virus strain.

Clinical specimens from 79 symptomatic individuals with a recent history of travel to countries with verified transmission of influenza A(H1N1)v (North America) were tested with a multiple real-time PCR targeting a broad range of agents that may cause acute respiratory infection. This analysis revealed that besides four cases of influenza A(H1N1)v, other respiratory viruses were diagnosed in almost 60% of the samples. These observations are a reminder that many different viral transmissions occur simultaneously in countries with ongoing spread of influenza A(H1N1)v. The findings demonstrate that the definition of suspected cases by clinical and epidemiological criteria has only a poor capacity for discriminating influenza A(H1N1)v from other viral infections.

Effective control measures limited measles outbreak after extensive nosocomial exposures in January-February 2008 in Gothenburg, Sweden.

In January-February 2008, one imported case of measles initiated a series of exposures with around 380 nosocomial secondary contacts. Susceptible individuals were traced early and control measures were initiated that managed to limit the consequences considerably. Only four secondary cases were identified by the end of March. This minor outbreak illustrates the importance and efficiency of early control measures as well as the fact that the risk of measles outbreaks still exists in a country that has high measles, mumps, rubella vaccination coverage among children.

A curriculum for training healthcare workers in the management of highly infectious diseases.

The SARS epidemic, the threat of bioterrorism, and recent examples of imported highly infectious diseases (HID) in Europe have all highlighted the importance of competent clinical and public health management of infectious disease emergencies. Although the European Union of Medical Specialists in Europe and the Infectious Diseases Society of America have developed curricula for training in infectious disease medicine, neither of those mentions training in the management of HIDs. The European Network for Infectious Diseases (EUNID, http://www.eunid.com) is a European Commission co-funded network of experts in HID management, created to help improve the preparedness for HID emergencies within Europe. One of EUNID's agreed tasks is the development of a curriculum for such a training. Between April 2005 and September 2006, EUNID developed a curriculum and accompanying training course on the basis of a questionnaire that was sent to all country representatives and discussion, followed by amendment of drafts shared through the project website, and a final consensus meeting. The resulting curriculum consists of a two-module course covering the core knowledge and skills that healthcare workers need to safely treat a patient who has, or who may have, an HID. The first module introduces theoretical aspects of HID management, including disease-specific knowledge, infection control, and the public health response, through didactic teaching and class-based discussion. The second module involves a "skill station" and a clinical scenario, and equips trainees with relevant practical skills, including the use of specialised equipment and teamwork practice in patient management. Together, the curriculum and course contribute to the creation of a common framework for training healthcare professionals in Europe, and although they are designed primarily for clinicians that are directly involved in patient care, they are relevant also to public health professionals and others who may be involved in HID management and emergency response.

Human cathelicidin, hCAP-18, is processed to the antimicrobial peptide LL-37 by extracellular cleavage with proteinase 3.

Cathelicidins are a family of antimicrobial proteins found in the peroxidase-negative granules of neutrophils. The known biologic functions reside in the C-terminus, which must be cleaved from the holoprotein to become active. Bovine and porcine cathelicidins are cleaved by elastase from the azurophil granules to yield the active antimicrobial peptides. The aim of this study was to identify the physiological setting for cleavage of the only human cathelicidin, hCAP-18, to liberate the antibacterial and cytotoxic peptide LL-37 and to identify the protease responsible for this cleavage. Immunoelectron microscopy demonstrated that both hCAP-18 and azurophil granule proteins were present in the phagolysosome. Immunoblotting revealed no detectable cleavage of hCAP-18 in cells after phagocytosis. In contrast, hCAP-18 was cleaved to generate LL-37 in exocytosed material. Of the 3 known serine proteases from azurophil granules, proteinase 3 was solely responsible for cleavage of hCAP-18 after exocytosis. This is the first detailed study describing the generation of a human antimicrobial peptide from a promicrobicidal protein, and it demonstrates that the generation of active antimicrobial peptides from common proproteins occurs differently in related species. (Blood. 2001;97:3951-3959)

Skin chamber technique for study of in vivo exudated human neutrophils.

The development of new techniques for isolation of neutrophils extravasated in vivo have been essential for studying the dynamics of the inflammatory response in humans. Methods for generating inflammatory skin reactions were first presented in the mid 1950s, and later a skin blistering technique based on suction was introduced. With this procedure, small areas of denuded dermis, called "skin windows", are created and covered with special chambers containing a medium that attracts exudated neutrophils. By comparing the neutrophils collected in such chambers with those isolated from peripheral blood, it is possible to investigate the functional modifications that neutrophils undergo when attracted to an inflammatory process. The skin-blister chamber technique represents an aseptic, non-traumatic and reproducible model of inflammation that can be used to study in vivo activated human neutrophils. The background, methodological aspects and options of this technique are described, together with the functional characteristics of exudated neutrophils.

Vertebral hydatid cyst infection (Echinococcus granulosus): a case report.

Spinal echinococcosis is a rare but serious condition. Within bone tissue hydatid cysts enlarge by daughter cyst formation. The value of drug treatment in bone echinococcosis is questionable. The aim of surgery is therefore removal of all the cysts. The best way to achieve this is at the first operation early in the progress of the disease. An anterior or circumferential approach is generally required to give the necessary accessibility. Owing to diffuse spread of the infection within the bone and the canal, recurrence is common. If neurological deterioration occurs, reintervention is necessary.

Galectin-3 activates the NADPH-oxidase in exudated but not peripheral blood neutrophils.

Galectin-3, a lactose-binding mammalian lectin that is secreted from activated macrophages, basophils, and mast cells, was investigated with respect to its ability to activate the human neutrophil NADPH-oxidase. The galectin-3-induced activity was determined with in vivo exudated cells (obtained from a skin chamber) and compared with that of peripheral blood neutrophils. Galectin-3 was found to be a potent activator of the NADPH-oxidase only in exudated neutrophils and the binding of galectin-3 to the surface of these cells was increased compared with peripheral blood cells. Different in vitro priming protocols resulting in degranulation were used to mimic the exudation process in terms of increasing the receptor exposure on the cell surface. Galectin-3 could induce an oxidative response similar to that in exudated cells only after a significant amount of the intracellular organelles had been mobilized. This increase in oxidative response was paralleled by an increased binding of galectin-3 to the surface of the cells. The major conclusion of the study is that galectin-3 is a potent stimulus of the neutrophil respiratory burst, provided that the cells have first experienced an extravasation process. The results also imply that the neutrophil response to galectin-3 could be mediated through receptors mobilized from intracellular granules, and we report the presence of galectin-3-binding proteins in such organelles.

Desensitization of formyl peptide receptors is abolished in calcium ionophore-primed neutrophils: an association of the ligand-receptor complex to the cytoskeleton is not required for a rapid termination of the NADPH-oxidase response.

Binding of ligands to N-formyl peptide chemoattractant receptors exposed on human neutrophils generates signals in the cells that induce an activation of the superoxide anion producing NADPH-oxidase. Ligand binding is followed by a rapid association of the ligand-receptor complex with the cytoskeleton, a process leading to desensitization of the cells with respect to NADPH-oxidase activation. We show that neutrophils that have experienced an intracellular calcium rise obtained through interaction with the calcium-specific ionophore ionomycin are "primed" with respect to the FMLP-induced production of superoxide anions. Mobilization of FMLP receptors from intracellular pools is one well-known mechanism behind the primed response. Based on our finding that ionomycin-treated neutrophils could not be desensitized, we suggest that the lack of association between the ligand-receptor complex and the cytoskeleton is an additional priming mechanism. Since in vivo-exudated neutrophils, which also had mobilized intracellular organelles, could be desensitized, we suggest that the abolished desensitization in ionomycin-treated neutrophils is not due to an inability of newly recruited receptors to couple to the cytoskeleton. We show that a rapid termination of FMLP-induced superoxide anion production is obtained in both desensitizable and nondesensitizable neutrophils, suggesting that the desensitization phenomenon is of limited importance in the oxidase termination process.

Invasive Haemophilus influenzae type b (Hib) infection in an adult patient with a selective deficiency of antibody to the Hib capsular polysaccharide.

A 31-year-old woman without any underlying disease contracted severe invasive Haemophilus influenzae type b (Hib) infection but developed no antibodies to the Hib capsular polysaccharide. Serum immunoglobulin levels were normal, but she had an isolated deficiency of antibody to Hib. Subsequently, immunization with a tetanus toxoid-conjugated Hib vaccine induced only a minimal response. However, she had a protective level of antibody (> 1 microgram/mL) after the fifth vaccination.

Phorbol myristate acetate-induced NADPH oxidase activity in human neutrophils: only half the story has been told.

The paradigm for activation of the neutrophil NADPH oxidase with the protein kinase C activator phorbol myristate acetate (PMA) states that the oxidase assembles the plasma membrane and that the metabolites generated are released extracellularly. This paradigm is challenged by the results presented. Most of the PMA-induced oxidase activity measured as chemiluminescence and dichlorofluorescein fluorescence was insensitive to scavengers of superoxide anion and hydrogen peroxide. This indicates that oxidase activity also takes place in a cellular compartment that the scavengers cannot reach. From the results obtained with granule-deficient HL-60 cells and cord blood neutrophils, we suggest that the scavenger-insensitive part of the NADPH oxidase activity in normal neutrophils resides in an intracellular compartment identical to or originating from granules. Our results also indicate that specific and azurophil granules have to be in very close contact to allow the generated oxygen metabolites to reach and react with myeloperoxidase.

Mobilization of granules and secretory vesicles during in vivo exudation of human neutrophils.

The extent of mobilization of four different intracellular compartments was measured during in vivo exudation of neutrophils into skin chambers and compared with resting neutrophils obtained from blood. Exudation of neutrophils induced increased surface expression of alkaline phosphatase, complement receptor 1, and Mac-1, and a complete loss of L-selectin. The increase in the content of surface molecules in the plasma membrane is in accordance with complete mobilization of secretory vesicles. Granule matrix proteins were secreted into the chamber fluid by the exudated neutrophils and the exocytosed proteins were recovered in the skin chamber fluid. Release of gelatinase from gelatinase granules was 38.1%, lactoferrin release from specific granules was 21.9%, and myeloperoxidase release from azurophil granules was 7.0%, clearly illustrating a hierarchy in mobilization among granules. When exudate neutrophils were stimulated with FMLP, additional mobilization of granules was observed and the rank order regarding release was preserved. This is the first report to evaluate the mobilization of secretory vesicles during in vivo exudation of human neutrophils. It is shown that secretory vesicles are regulated exocytotic vesicles that are fully mobilized during in vivo exudation. Once exocytosed, secretory vesicles are not re-formed within a period of 6 h.

Neutrophil alkaline phosphatase activity increase in bacterial infections is not associated with a general increase in secretory vesicle membrane components.

The content of alkaline phosphatase (ALP) was determined in neutrophils isolated from patients with acute bacterial infections by a standard enzyme assay. Compared with control cells, patient cells exhibited about a fivefold increase in ALP activity. There was no difference between the ALP Km values of control and patient cells, which indicates that the elevated activity in patient cells was due to the presence of increased amounts of the enzyme. The ALP isozyme in both cell types was determined to be the tissue-unspecific ALP. The fact that much of the ALP activity was measurable only in the presence of detergent suggested that the enzyme was localized in the secretory vesicles, a putative reservoir of plasma membrane components. The amount and subcellular distribution of two other secretory vesicle membrane proteins, i.e., cytochrome b and complement receptor 3, were not altered; hence, we conclude that there was no general increase in amounts of secretory vesicle membrane constituents in the patient cells.

Quantitative slot-blot chemiluminescence assay for determination of myeloperoxidase from human granulocytes.

Using slot blot, we show that myeloperoxidase (MPO), a constituent of azurophil granules of neutrophil polymorphonuclear leukocytes, can be measured quantitatively using a commercially available chemiluminescence kit, originally developed for detection of specific proteins on Western blots. MPO is determined through its ability to catalyze the oxidation of luminol, resulting in the emission of light which is recorded on a photographic film. The sensitivity of the method is high and allows MPO from less than 100 cells to be detected. This method was used to determine MPO in exudate fluid and in neutrophil fractions following disintegration and subcellular fractionation of the postnuclear supernatant on two-layer Percoll gradients.

Successful interferon-gamma therapy in a chronic granulomatous disease (CGD) patient suffering from Staphylococcus aureus hepatic abscess and invasive Candida albicans infection.

A 16-year old boy with an early history of recurrent lower respiratory tract infections exhibited symptoms of prolonged septic fever and liver abscess. Cultures from liver puncture were positive for Staphylococcus aureus, and the patient initially responded to antibacterial therapy. After a period of 4 months, however, the infection relapsed, and further treatment with broad antibacterial, antifungal and tuberculostatic drugs was ineffective. Neither soluble nor particulate stimuli were found to elicit the respiratory burst response in granulocytes from the patient. Spectral analysis of granulocyte cytochrome-b confirmed the diagnosis of chronic granulomatous disease. Since the patient's physical condition deteriorated severely during the prolonged (10-week) septic course, immunosupportive interferon-gamma was added to the anti-microbial therapy. With this regime, the fever subsided and the general condition of the patient improved dramatically. He could be discharged from hospital 9 weeks after the introduction of interferon-gamma and was, at an elective follow-up control 1 month later, convalescing and showed no signs of active infection.

Phagocytosis following translocation of the neutrophil b-cytochrome from the specific granule to the plasma membrane is associated with an increased leakage of reactive oxygen species.

The effect of neutrophil b-cytochrome translocation on the respiratory burst activation generated during phagocytosis of yeast particles was investigated. Secretion of neutrophil specific granules was induced by the calcium ionophore ionomycin prior to phagocytosis. The secretory process is associated with a translocation from the specific granules to the plasma membrane of the respiratory burst b-cytochrome. Respiratory burst activity was measured as release of hydrogen peroxide in the absence of azide (extracellular leakage) and in the presence of azide (total production). The subcellular localization of the b-cytochrome was found to affect the extracellular release of hydrogen peroxide in that a plasma membrane localization was associated with a significantly increased release during phagocytosis. It should be pointed out, however, that most of the hydrogen peroxide, both in control and in ionomycin-treated cells, is produced intracellularly, probably in the phagosomes.

N-acetylcysteine enhances receptor-mediated phagocytosis by human neutrophils.

The effect of the sulphur compound N-acetylcysteine (NAC) on certain receptor-mediated cellular functions [chemiluminescence (CL), phagocytosis and degranulation] in human neutrophils was studied, to evaluate how a scavenger of certain toxic oxygen product can protect the phagocyte and the bystander tissue cells from oxidative damage. When using IgG-opsonized yeast particles as stimulating agent, preincubating the neutrophils with NAC (0.25 mg/ml = 1.5 mM) increased both the CL response and phagocytosis. Higher concentrations of NAC (0.50-1.00 mg/ml = 3-6 mM) decreased the CL response, whereas the phagocytic capacity was still enhanced. This effect was more pronounced with adherent neutrophils than with neutrophils in suspension. No increased CL or phagocytic activity was, however, induced by NAC when C3bi-opsonized particles were used as a prey. From the fact that NAC (i) inhibited extracellularly localized myeloperoxidase dependent activities, and (ii) had no effect on neutrophils from patients with chronic granulomatous disease (CGD), we conclude that the scavenger effect of NAC not only reduces the accumulation of oxidative metabolites per se, but also enhances receptor-mediated phagocytosis by protecting Fc(IgG)-receptors from oxidative damage mediated by myeloperoxidase (MPO) and hydrogen peroxide (H2O2). Since NAC can increase phagocytosis and reduce the extracellularly produced oxidative metabolites, we furthermore conclude that NAC possesses some ideal properties as an anti-inflammatory agent.