Reduced cytotoxicity by mutation of lysine 590 of Pseudomonas exotoxin can be restored in an optimized, lysine-free immunotoxin.

Immunotoxins, which are fusion proteins of an antibody fragment and a fragment of a bacterial or a plant toxin, induce apoptosis in target cells by inhibition of protein synthesis. ADP-ribosylating toxins often have few lysine residues in their catalytic domain. As they are the target for ubiquitination, the low number of lysines possibly prevents ubiquitin-dependent degradation of the toxin in the cytosol. To reduce this potential degradation, we aimed to generate a lysine-free (noK), Pseudomonas exotoxin (PE)-based immunotoxin. The new generation 24 kDa PE, which lacks all but the furin-cleavage site of domain II, was mutated at lysine 590 (K590) and at K606 in a CD22-targeting immunotoxin and activity was determined against various B cell malignancies in vitro and in vivo. On average, K590 mutated to arginine (R) reduced cytotoxicity by 1.3-fold and K606R enhanced cytotoxicity by 1.3-fold compared to wild type (wt). Mutating K590 to histidine or deleting K590 did not prevent this loss in cytotoxicity. Neither stability nor internalization rate of K590R could explain reduced cytotoxicity. These results highlight the relevance of lysine 590 for PE intoxication. In line with in vitro results, the K606R mutant was more than 1.8-fold more active than the other variants in vivo suggesting that this single mutation may be beneficial when targeting CD22-positive malignancies. Finally, reduced cytotoxicity by K590R was compensated for by K606R and the resulting lysine-free variant achieved wt-like activity in vitro and in vivo. Thus, PE24-noK may represent a promising candidate for down-stream applications that would interfere with lysines.

Zoonoses in the European Union: origin, distribution and dynamics - the EFSA-ECDC summary report 2009.

We present a summary of the main findings of the latest report of the European Food Safety Authority and European Centre for Disease Prevention and Control on zoonoses, zoonotic agents and food-borne outbreaks in the European Union (EU), based on data from 2009. Zoonoses are prevalent and widely distributed across several countries in the EU. The most important highlight of this report was the continuous decrease of human salmonellosis since 2005, probably due to effective control programmes in livestock.

Infection control--a European research perspective for the next decade.

A symposium was held in June 2009 near Freiburg in Germany. Twenty-nine attendees from several European countries participated, most of whom are actively involved in research and hospital infection prevention and control. The following topics were presented and discussed: isolation and screening for control of multidrug-resistant organisms; impact of the environment on healthcare-associated infection (HAI); new technologies to control infection--state of evidence; surveillance of HAI; methodological challenges and research priorities for infection control and control of HAI: learning from each other in a united Europe. This Leader summarises the main issues for debate and the number of consensus points agreed amongst delegates.

Initial surveillance of 2009 influenza A(H1N1) pandemic in the European Union and European Economic Area, April-September 2009.

European Union (EU) and European Economic Area (EEA) countries reported surveillance data on 2009 pandemic influenza A(H1N1) cases to the European Centre for Disease Prevention and Control (ECDC) through the Early Warning and Response System (EWRS) during the early phase of the 2009 pandemic. We describe the main epidemiological findings and their implications in respect to the second wave of the 2009 influenza pandemic. Two reporting systems were in place (aggregate and case-based) from June to September 2009 to monitor the evolution of the pandemic. The notification rate was assessed through aggregate reports. Individual data were analysed retrospectively to describe the population affected. The reporting peak of the first wave of the 2009 pandemic influenza was reached in the first week of August. Transmission was travel-related in the early stage and community transmission within EU/EEA countries was reported from June 2009. Seventy eight per cent of affected individuals were less than 30 years old. The proportions of cases with complications and underlying conditions were 3% and 7%, respectively. The most frequent underlying medical conditions were chronic lung (37%) and cardio-vascular diseases (15%). Complication and hospitalisation were both associated with underlying conditions regardless of age. The information from the first wave of the pandemic produced a basis to determine risk groups and vaccination strategies before the start of the winter wave. Public health recommendations should be guided by early capture of profiles of affected populations through monitoring of infectious diseases.

Experience and lessons from surveillance and studies of the 2009 pandemic in Europe.

Surveillance and studies in a pandemic is a complex topic including four distinct components: (1) early detection and investigation; (2) comprehensive early assessment; (3) monitoring; and (4) rapid investigation of the effectiveness and impact of countermeasures, including monitoring the safety of pharmaceutical countermeasures. In the 2009 pandemic, the prime early detection and investigation took place in the Americas, but Europe needed to undertake the other three components while remaining vigilant to new phenomenon such as the emergence of antiviral resistance and important viral mutation. Laboratory-based surveillance was essential and also integral to epidemiological and clinical surveillance. Early assessment was especially vital because of the many important strategic parameters of the pandemic that could not be anticipated (the 'known unknowns'). Such assessment did not need to be undertaken in every country, and was done by the earliest affected European countries, particularly those with stronger surveillance. This was more successful than requiring countries to forward primary data for central analysis. However, it sometimes proved difficult to get even those analyses from European counties, and information from Southern hemisphere countries and North America proved equally valuable. These analyses informed which public health and clinical measures were most likely to be successful, and were summarized in a European risk assessment that was updated repeatedly. The estimate of the severity of the pandemic by the World Health Organization (WHO), and more detailed description by the European Centre for Disease Prevention and Control in the risk assessment along with revised planning assumptions were essential, as most national European plans envisaged triggering more disruptive interventions in the event of a severe pandemic. Setting up new surveillance systems in the midst of the pandemic and getting information from them was generally less successful. All European countries needed to perform monitoring (Component 3) for the proper management of their own healthcare systems and other services. The information that central authorities might like to have for monitoring was legion, and some countries found it difficult to limit this to what was essential for decisions and key communications. Monitoring should have been tested for feasibility in influenza seasons, but also needed to consider what surveillance systems will change or cease to deliver during a pandemic. International monitoring (reporting upwards to WHO and European authorities) had to be kept simple as many countries found it difficult to provide routine information to international bodies as well as undertaking internal processes. Investigation of the effectiveness of countermeasures (and the safety of pharmaceutical countermeasures) (Component 4) is another process that only needs to be undertaken in some countries. Safety monitoring proved especially important because of concerns over the safety of vaccines and antivirals. It is unlikely that it will become clear whether and which public health measures have been successful during the pandemic itself. Piloting of methods of estimating influenza vaccine effectiveness (part of Component 4) in Europe was underway in 2008. It was concluded that for future pandemics, authorities should plan how they will undertake Components 2-4, resourcing them realistically and devising new ways of sharing analyses.

Incidence and risk factors for community-acquired acute gastroenteritis in north-west Germany in 2004.

In developed countries, acute gastroenteritis (AGE) is a major source of morbidity. However, only a few studies have estimated its incidence and the associated medical burden. This population-based study determined the incidence of community-acquired AGE patients seeking medical care and the relative role of various pathogens. Stool samples from patients with AGE presenting to a general practitioner (GP), pediatrician, or specialist in internal medicine for that reason were screened for various bacterial and viral enteropathogens. A control group was established as well. Incidences were calculated by the number of positive patients divided by the general population. The study was performed in north-west Germany in 2004. The incidence of AGE patients requiring medical consultation was 4,020/100,000 inhabitants. Children (<5 years of age) were at the highest risk (13,810/100,000 inhabitants). Of the patients, 6.6% were tested positive for an enteropathogenic bacteria and 17.7% for a viral agent. The predominant pathogens were norovirus (626/100,000) and rotavirus (270/100,000). Salmonella was the most frequently detected bacteria (162/100,000). The results presented confirm AGE and, specifically, AGE of viral origin as a major public health burden in developed countries.

Zoonotic infections in Europe in 2007: a summary of the EFSA-ECDC annual report.

The European Food Safety Authority and the European Centre for Disease Prevention and Control have just published their Community Zoonoses Report for 2007, analysing the occurrence of infectious diseases transmittable from animals to humans. Campylobacter infections still topped the list of zoonotic diseases in the European Union and the number of Salmonella infections in humans decreased for the fourth year in a row. Cases of listeriosis remained at the same level as in 2006, but due to the severity of the disease, more studies on transmission routes are warranted. The report highlights the importance of continued co-operation between veterinarians and public health specialists, both at the EU level and within Member States.

Sorbitol-fermenting enterohaemorrhagic Escherichia coli O157:H- causes another outbreak of haemolytic uraemic syndrome in children.

An outbreak of haemolytic uraemic syndrome (HUS) among children caused by infection with sorbitol-fermenting enterohaemorrhagic Escherichia coli O157:H- (SF EHEC O157:H-) occurred in Germany in 2002. This pathogen has caused several outbreaks so far, yet its reservoir and routes of transmission remain unknown. SF EHEC O157:H- is easily missed as most laboratory protocols target the more common sorbitol non-fermenting strains. We performed active case-finding, extensive exploratory interviews and a case-control study. Clinical and environmental samples were screened for SF EHEC O157:H- and the isolates were subtyped by pulsed-field gel electrophoresis. We identified 38 case-patients in 11 federal states. Four case-patients died during the acute phase (case-fatality ratio 11%). The case-control study could not identify a single vehicle or source. Further studies are necessary to identify the pathogen's reservoir(s). Stool samples of patients with HUS should be tested with an adequate microbiological set-up to quickly identify SF EHEC O157:H-.

Developing the Community reporting system for foodborne outbreaks.

Investigating and reporting of foodborne outbreaks became mandatory with Directive 2003/99/EC. In 2006 and 2007 the Community reporting system for foodborne outbreaks was further developed in an interdisciplinary approach, which is described in this paper. This involved experts on investigating and reporting foodborne outbreaks as well as experts on communicable diseases in addition to the European Food Safety Authority (EFSA) Task Force for Zoonoses Data Collection, the European Centre for Disease Prevention and Control (ECDC) Advisory Forum and representatives of ECDC, the World Health Organization (WHO), the World Organization for Animal Health (OIE) and the European Commission. European Union Member States participated in a survey regarding their national reporting systems and the needs for information on foodborne outbreaks at the Community level. The acceptability, the functionality and the data quality of the current reporting system were evaluated. The results were used to propose new variables on which data should be reported. Pick-lists were developed to facilitate reporting and better integration of the Community system with Member States' reporting systems. The new system is expected to yield better quality data on foodborne outbreaks relevant for risk assessment and risk management while reducing the work load for Member States.

The surveillance of communicable diseases in the European Union--a long-term strategy (2008-2013).

This article presents the steps and considerations that led to the development of the European Centre for Disease Prevention and Control s (ECDC) long-term strategy for the surveillance of communicable diseases in the European Union (EU) for the years 2008 to 2013. Furthermore, it outlines the key features of the strategy that was approved by the ECDC s Management Board in December 2007.

Longitudinal study of meningococcal carrier rates in teenagers.

To gain actual information concerning the oropharyngeal carriage of Neisseria meningitidis among teenagers aged 15-18 years in Germany especially in a region with increased incidence of meningococal-related diseases prompted the study. Each teenager was swabbed three times with an interval of 2 months between the examinations. The 901 recovered N. meningitidis strains were characterized using serological (serogrouping, serotyping/serosubtyping) and molecular methods (PCR, PFGE) each. The results of the study demonstrate an overall average carrier rate of 18.8% for the three collection periods. There were, however, significant differences between the carrier rates within a given school and of different towns and counties. Of all isolates, 60.6% were not serogroupable. Serogroup B dominated (12.3%), followed by serogroup Y (9.0%) and serogroup C (3.6%). After PCR-based serogrouping of not serogroupable strains the percentages for serogroups enhanced to 18.8% for B, 10.8% for Y and 4.1% for C. Serotyping led to 305 different phenotypes with the most common being 29E:NT:P1.2,5 followed by Y:14:NST. In the 6 study towns the number of different N. meningitidis clones (PFGE types) isolated, varied between 30 and 87. In Wenden, where a prolonged outbreak had taken place, serogroup C (14.8%) was predominant. Only in this town C:2a isolates were found, all belonging to the ST-11/ET-37 complex and 12/13 matched identically to the ET-15 clone. Of the colonized teenagers, 26.7% were carriers over at least 23 weeks, 22.6% with the same strain, 36.0% were carrier for at least 15 weeks. Over all three collection periods 36.7% of the adolescents acquired a new strain. The highest acquisition rate was related to PFGE type 12.

[Surveillance of infectious foodborne diseases after enactment of the German Protection against Infection Act in 2001. Potentials and requirements]. / Surveillance Lebensmittel-übertragener Infektionserkrankungen durch das Infektionsschutzgesetz: Möglichkeiten und Anforderungen.

The epidemiology of infectious foodborne diseases has changed. Outbreaks more frequently occur geographically dispersed or protractedly over longer periods of time, and they often appear as a scatter of seemingly sporadic cases. This hampers and delays the identification of their epidemiological link. The surveillance of infectious foodborne diseases has to be refined accordingly to be able to detect these diffuse outbreaks. The German Protection against Infection Act, enacted in 2001, offers the potential of increased sensitivity due to timely electronic reporting of individual cases and detailed data accompanying each report. In addition to a timely and comprehensive reporting system, subtyping of pathogens has become an invaluable tool in identifying epidemiologically linked cases, i.e. outbreaks. Still, the sensitivity of foodborne disease surveillance still hinges on the willingness of physicians to order stool testing for enteric pathogens (and to report suspected outbreaks to local health departments). Without the active participation of physicians, the chance of detecting outbreaks and successfully investigating them is markedly reduced. Consequently, the general preventive strategy would be jeopardised, namely to understand the (often new) mechanisms by which contamination and disease transmission occur well enough to interrupt them.

Outbreak of hepatitis A in two federal states of Germany: bakery products as vehicle of infection.

In April 2004, increased numbers of hepatitis A were noted in six neighbouring districts in Germany. Exploratory interviews showed that patients had consumed bakery products from company X where two employees had been diagnosed with hepatitis A in February. A case-control study of consumption of products of company X was carried out through telephone interviews. Altogether, 64 cases were identified. Fifty-two cases and 112 controls aged >or=16 years were included in the case-control study. In total, 46/52 cases and 37/112 controls had consumed company X products [odds ratio (OR) 15.5, 95% confidence interval (CI) 6.1-39.7]. Of these, 36/46 cases and 16/37 controls had consumed pastries (OR 4.7, 95% CI 1.8-12.3), 25/46 cases and 12/37 controls had consumed filled doughnuts (OR 2.5, 95% CI 1.0-6.1). Sequence analysis of the VP1-2A junction region indicated 100% strain homology between cases and an infected employee of company X. We recommended reinforcement of hygiene precautions, and consideration of a prolongation of compulsory work absence after post-exposure vaccination.

SurvNet@RKI--a multistate electronic reporting system for communicable diseases.

In 2001 Germany implemented a new electronic reporting system for surveillance of notifiable infectious diseases (SurvNet@RKI). The system is currently being used in all 431 local health departments (LHD), the 16 state health departments (SHD) and the Robert Koch-Institut (RKI), the national agency for infectious disease epidemiology. The SurvNet@RKI software is written in MS Access 97 and Visual Basic and it supports MS Access as well as MS SQL Server database management systems as a back-end. The database is designed as a distributed, dynamic database for 73 reporting categories with more than 600 fields and about 7000 predefined entry values. An integrated version management system documents deletion, undeletion, completion and correction of cases at any time and entry level and allows reproduction of previously conducted queries. Integrated algorithms and help functions support data quality and the application of case definitions. RKI makes the system available to all LHDs and SHDs free of charge. RKI receives an average of 300,000 case reports and 6240 outbreak reports per year through this system. A public web-based query interface, SurvStat@RKI, assures extensive and timely publication of the data. During the 5 years that SurvNet@RKI has been running in all LHDs and SHDs in Germany it has coped well with a complex federal structure which makes this system particularly attractive to multinational surveillance networks. The system is currently being migrated to Microsoft C#/.NET and transport formats in XML. Based on our experiences, we provide recommendations for the design and implementation of national or international electronic surveillance systems.