Need and relevance.
Co-operation between academic, health organizations and industry is promoted by the European Confederation of Medical Mycology, covering the mycological societies of 19 European countries. One of the key activities of this platform is the implementation of Surveillance Working Groups. These are extremely useful to acquire an overview of the clinical significance of major mycoses and of changing epidemiological trends and promote awareness of the emergence of resistance against antifungal drugs.
An ECMM Working Group was founded at the ECMM congress in Budapest, August 2002 as a continuation of an existing informal international network, to focus European attention on the much overlooked but important, potentially life-threatening systemic and disseminated infections by Pseudallescheria and Scedosporium. Due to the therapy-refractory character of these fungi, morbidity and mortality by such infections is high. There is a high degree of genetic diversity within both species, which diminishes the predictive value of standard antifungal susceptibility data. Consequently the infectious diseases united under the umbrella term Pseudallescheriasis provide a potent model for the development of new strategies for control of therapy-refractory emerging opportunists. The consortium will obtain insight into the occurrence and genetic variability of these fungi, and provide data on possible sources of contamination and infection routes. A multidisciplinary approach in health care of the immunocompromised patient population is necessary to understand the emergence of new fungal diseases. Improved diagnostics, at the generic level and down to the (sub)specific level, can be developed and disseminated to the European clinician, which is expected to greatly stimulate awareness of Pseudallescheria, Scedosporium and other fungal infections. This synergistic approach of the pan-European network will lead to an expertise centre with public data bank containing information on strains and their genetic make-up, clinical cases, and antifungal susceptibilities, and will contribute to diagnostic protocols and guidelines for therapy. A genomic approach will be implemented to select key-genes involved in the resistance to antimycotics, providing a basis for the development of novel and dedicated antifungals by pharmaceutical companies, and to efficient treatment protocols.
Infections by Pseudallescheria / Scedosporium have a relatively high degree of virulence. This intrinsic virulence can be deduced from the extremely infrequent occurrence of the species in air but the high prevalence in the lungs of susceptible patients: P. boydii is with a frequency of 8.6% the second most common filamentous fungus in the lungs of CF patients. In Europe around 18,000 patients will be diagnosed with acute leukemia alone and around 13,000 die of this disease each year. 99,000 patients will be treated for a hematologic malignancy and around 18,800 will be bone-marrow or organ transplanted. About 5-24% of the patients undergoing lung or allogenic bone-marrow transplantation and of the patients with acute leukemia will develop invasive mycoses. Roughly estimated, 5-6000 cases concern filamentous fungi, which will in the vast majority be the cause of death. Many of these are unnecessary deaths because the underlying disease can be cured or temporarily stabilized. Important is the present lack of sufficient early diagnostics, including the identification of the infectious agent. Due to significant differences in the susceptibility of fungi to antimycotic agents, an universal recommendation for an antifungal therapy without knowledge about the individual fungal isolate is not possible. For Pseudallescheria / Scedosporium there are major problems to be solved: (1) the agents of disease are often not or inappropriately recognized, and (2) if recognized, current therapeutic regimens are inadequate, and (3) knowledge on the biology, clinical potential, virulence, and routes of transmission and infection is lacking. Investigation of the scattered data on this species complex is hardly possible without (4) an extended data bank.
All members of the Network are kindly requested to put the following footnote on their publications:
*Communication of the ECMM Working Group on Pseudallescheria and Scedosporium (www.Scedosporium-ECMM.com).
OVERVIEW OF ACTIVITIES
(1) Clinical surveillance, epidemiology and isolation
A preliminary inquiry of a number of strictly comparable university hospitals revealed that the frequency of Pseudallescheria and Scedosporium infections was very different. This is possibly explained by two major problem areas: isolation, and diagnosis.
Presence of the fungus in non-sterile body sites is often concealed by rapidly growing colonizers such as Aspergillus and Candida. Selective protocols and guidelines of incubations times and conditions are therefore necessary. It is noted in CF lungs A. fumigatus may appear first, and subsequently the therapy-refractory Pseudallescheria appears after administration of antifungals against Aspergillus. This suggests that Pseudallescheria was present but overlooked.
The histopathology of Scedosporium is identical to that of aspergillosis and therefore the species are frequently misidentified as such. With serology, cross-reactivity is noted with phylogenetically remote fungi like Cryptococcus neoformans. In culture the species may be highly polymorphic. This has led to the frequent redescription of the fungus as a new species and even new genus, the last of which was Polycytella, in 1987. In members of our consortium with increased awareness their frequency currently is estimated to be fatal in about 1-3:106 patients per year, and increasing. It is expected that upon re-evaluation of archival autopsy specimens, numerous cases ascribed to Aspergillus will turn out to have been caused by a Pseudallescheria or Scedosporium species.
Aims: Obtain a balanced picture of the clinical presentation of Pseudallescheria / Scedosporium, including colonization and invasion, and routes of dissemination within the patient.
Expected practical outcome: Isolation / detection protocols; collection of strains for research; epidemiology and frequency.
(1) Define the real occurrence of the different forms of pseudallescheriasis (with a particular emphasis in CF patients),
(2) Define guidelines for mycological examination of clinical samples (with an emphasis on sputum samples from CF patients),
(3) Improve isolation protocols from any clinical sample,
(4) Improve the serological diagnosis of scedosporiosis and analyis of the antigenic equipment of the fungus,
(4) Identify specific antigens allowing the development of a western-blot procedure or of an ELISA test using a monoclonal antibody directed towards one of these specific antigens,
(5) Improve the histopathological examination of biopsies and necropsies with a MAb-based kit or using a molecular biological approach,
(6) Describe routes of dissemination within the patient,
(7) Establish numbers of genotypes within the patient,
(8) Preserve strains for later research.
Steering committee: Jean-Philippe Bouchara, Gerhard Haase, Sevtap Arikan.
(2) Virulence, animal models, antifungals and therapy
Their variable but always remarkably high tolerance to antimycotics makes the species a severe threat for the hospitalised population. In these patients, immediate and adequate antimycotic therapy is mandatory. However, this is hampered by the bewildering genetic diversity of the Pseudallescheria. Published MIC values have limited predictive value for effective treatment of the patient at hand. Thus there is a severe gap in our knowledge on these organisms, with the unnecessary loss of patients as a probable consequence. The proposed concerted action will provide a major leap forward towards the development of reliable treatment protocols. The increasing frequency, the high case fatality rate in immunocompromised patients, and the recent availability of genomic techniques to solve many of the problems, warrant concerted action to diminish the mortality caused by this disease.
Aims: Obtain insight into the virulence factors of Pseudallescheria / Scedosporium, including colonization and invasion, and routes of dissemination in the patient; development of adequate antifungal therapy.
Expected practical outcome: Insight in virulence factors; animal models; therapeutic guidelines.
(1) Prepare a genomic bank for S. apiospermum which will be use for the identification and characterization of some putative virulence factors of the fungus (e.g. adhesins, anti-oxidant systems, proteases, etc.),
(2) Elaborate and develop vertebrate and non-vertebrate animal models for invasive disease, and perhaps also for ingestion,
(3) Establish correlation of in vitro MIC and in vivo outcome in animal model,
(4) Develop effective antifungal (combination) therapy for P. boydii and S. prolificans.
(5) Develop phagocytose assays for P. boydii.
Steering committee: Paul Verweij, Josep Guarro, Aristea Velegraki, Bernard Cimon.
(3) Biology, diagnostics and transmission
Why particular fungal species are emerging? Pseudallescheria is normally associated with heavily polluted environments, such as sewage, mud of geese-inhabited village ponds, manure and agricultural soil. Its frequency in the environment is strongly enhanced by the behaviour of mankind in rural and urban settings, where nitrogenic compounds are ubiquitous pollutants. In the unpolluted environment the species is very rare; there are only occasional reports of the fungus recovered from the intestinal tract of amphibians. Despite their occurrence in soil and water in temperate climates, the species are consistently thermotolerant and have the ability to survive at very low oxygen pressures. This may indicate a natural ecological niche involving the use of a warm-blooded animal as a vector of dissemination.
The jump of the fungus from the unknown natural niche to the man-dominated environment may be accompanied by selection of virulent genotypes and thus by an evolution towards higher average degrees of virulence. The possibility of such events is suggested by the presence of genetic entities within the species with different ecology, such as predominantly clinical vs. environmental. The dung-polluted rural environment has created an enormous window of opportunity for the species. Therefore the fungus species may develop towards higher virulence without jeopardizing its own existence. Apparently it is best for the species to propagate with large numbers, regardless if this interferes with mobilization of eventual hosts. More rapidly growing genotypes thus have an increased probability of transmission and thus have a higher evolutionary fitness. If this hypothesis is correct, it is likely that genotypes with higher virulence and micro-aerophily are selected and that the fungus indeed becomes more virulent in the course of its evolution. From our own data it is apparent that several genotypes with marked differences in growth rate occur next to each other in the same ecosystem, which suggests that the slower genotypes have not yet been outcompeted by the virulent ones. It is probable that new genotypes are continuously brought in. The human creation of new environments which enhance the eutrophic opportunistic pathogen Pseudallescheria leads to the prediction that more virulent and more resistant genotypes will be selected in the course of short-term evolution. This would mean that the current problems with infection of the immunocompromised patient by Pseudallescheria and Scedosporium will aggravate. The problem of resistance to antifungal therapy facing a growing population of susceptible patients increases concomitantly. Using Pseudallescheria as a model, our research will provide insight into the question why only some of opportunists, out of the 100,000 species known, are emerging. This is a fundamental question in any research on virulence of opportunistic fungi such as Candida or Aspergillus.
Aims: Obtain insight into the environmental ecology of Pseudallescheria / Scedosporium, including routes of transmission, and evolutionary events during adaptation to the man-dominated environment.
Expected practical outcome: Hygienic guidelines.
(1) Description of phylogenetic structure of P. boydii and related species,
(2) Description of intraspecific variability using independent markers,
(3) Revealing processes of selection and of speciation by competition experiments,
(4) Description of genetic processes of breeding, compatibility and clonality,
(5) Refined diagnostic protocols using culture- and nonculture-methods,
(6) Revealing natural ecological niche of the species,
(7) Experimental set-up of routes of airborne transmission including hijack with Aspergillus conidia,
(8) Exploring possibilities of development of P. boydii as a biomarker for environmental pollution.
Steering committee: Sybren de Hoog, Johannes Rainer, Juan Rodriguez-Tudela.
In order to obtain materials for study covering variability of the species, a pan-European surveillance system is mandatory. A web-based data base will be built up and made accessible for all participants for streamlining formatted data management and safeguarding of strains through central collections. The collection will be largely virtual: all data on strains will be stored centrally, but the strains do not need to be stored centrally. Central storage is recommended for strains that are at risk to be lost, such as those isolated during patient surveillance in hospitals, where the infrastructure for long-term preservation may be absent. Francoise Symoens (IHEM, Brussels) is prepared to store all threatened strains and smaller collections, and to make working collections available to all participants of the network. All participants will share their strains with any other participant freely.
With any research in the network, we will take sets of reference strains into account, as strains may have a different status. The nomenclatural type strains (i.e. first strains on which species descriptions were based) are available at CBS. We will select sets of methodical reference strains, in order to enhance comparability of research outcome. It is recommended to include these strains in any study. The set will be selected on (a) genetic diversity, and (b) strains that were used in laborious experiments, such as animal experiments or phagocytosis assays.
Aims: Building a central, web-accessible information centre aupporting workers in themes (1-3) above.
Expected practical outcome: General purpose data bank.
(1) Establishment of a webdatabase accessible for all the participants.
(2) Distribution of reference strains for research purposes and long term preservation of the clinical isolates no preserved in hospital centres,
(3) Preparation of a form enhancing deposition of strains in data base and one of the culture reference centres.
Steering committee: Vincent Robert, Francoise Symoens, Regine Horré.
Sybren de Hoog