Pneumocystis infection: unraveling the colonization-to-disease shift

Abstract

A Commemorative Conference of Pneumocystis Discovery First Centenary was held in Brussels, Belgium, on 5–6 November 2009. A total of 16 keynote speakers from different countries attended the meeting. This conference has allowed the principal European and non-European groups who are working on Pneumocystis infection to gather together, in order to expose the most recent advances accomplished in the basic and translational scientific knowledge of Pneumocystis infection, and to discuss the trends of future research in this area.

The year 2009 marked the 100th anniversary of the first description of the microorganisms that are known today as Pneumocystis[1]. During the past 100 years, the image of Pneumocystis has been dramatically transformed. Basic research on Pneumocystis infection has been hampered by the lack of a reliable in vitro culture system; nevertheless, through the use of molecular techniques and experimental models, progress has been made over the last few years and important research advances have radically changed our conceptions about Pneumocystis development, infection sources, reservoir, taxonomy and clinical manifestations [2].

The Commemorative Conference of Pneumocystis Discovery First Centenary, held in Brussels in November 2009, has allowed the gathering together of the principal European and non-European groups who are working in Pneumocystis infection in order to expose the most recent advances accomplished in the basic and translational scientific knowledge of Pneumocystis infection and has allowed discussion about future research in this area. This meeting report summarizes the highlights of the conference, which was divided into five major sessions.

Biology of Pneumocystis organisms

Pneumocystis organisms are now considered as part of an early diverging lineage of Ascomycetes. Although no robust long-term culture model is available, major advances have been made in our understanding of Pneumocystis biology in the past few years. Key proteins involved in the metabolic pathways, signal transduction cascades, cell wall assembly and mitotic cell cycle have been identified [3]. The lifecycle of Pneumocystis microorganisms is thought to involve a controversial asexual mode of replication via active binary fission of the trophic form and a sexual mode resulting in formation of an ascus (cyst) containing eight ascospores. Although most fungi developing in animals do not complete a sexual cycle in vivo, Pneumocystis species constitute one of a few exceptions. All of the known lifecycle stages of Pneumocystis have been found in the lungs of infected mammals. Trophic forms, sporocytes and mature cysts are usually considered to be the three main morphological forms involved in the Pneumocystis lifecycle. Trophic forms are the most abundant of all Pneumocystis lifecycle stages, representing 90–95% of the total population in the lungs of hosts with pneumocystosis. These forms may be involved in mating, as evidenced by homologs to yeast pheromone receptor genes present in the Pneumocystis carinii genome and the expression of a pheromone receptor protein on the surface of some trophic forms [4].

Sequences for the mitochondrial genome of P. carinii are now available as a result of the Pneumocystis Genome Project [101]. Some data have recently provided strong evidence that the mitochondrial genome of P. carinii is linear. The significance of a linear genome in Pneumocystis is not yet understood, but it has been suggested that it may convey a survival advantage [5].

The ability of P. carinii and Pneumocystis murina to form biofilms has been documented in an in vitro setting. Biofilm formation probably occurs in the mammalian lung and confers several advantages for the survival of Pneumocystis[6].

Epidemiology of Pneumocystis infection

Molecular techniques have revealed a high prevalence of Pneumocystis colonization in wild mammals, probably resulting from active airborne horizontal and vertical transmission mechanisms. Cophylogeny is the evolutionary pattern for Pneumocystis species, which have dwelt in the lungs of mammals for more than 100 million years [7]. The strong host species specificity of Pneumocystis strains suggests that Pneumocystis infection in humans is an anthroponosis, and that humans serve as reservoir for Pneumocystis jirovecii, the sole species found in humans. Airborne transmission of Pneumocystis spp. from host to host has been demonstrated in rodent models and several observations suggest that interindividual transmission occurs in humans in both hospitals, as a nosocomial infection, and in the community [8]. In this sense, a recent study has provided molecular evidence that transmission of P. jirovecii from colonized immunocompetent carrier hosts to susceptible persons may occur [9]. It has been suggested that individuals with chronic pulmonary diseases who are colonized by P. jirovecii have a significant role as major reservoirs and sources of infection [10].

Vertical transmission of Pneumocystis via the transplacental route has been demonstrated in rabbits, but it does not appear to occur in rats or severe combined immunodeficient mice. Only last year, the first molecular evidence of P. jirovecii transplacental transmission in humans was provided by documenting the presence of P. jirovecii DNA in fetal lung and placenta samples, recovered from nonimmunodepressed pregnant women who had a miscarriage [11]. The finding could be of potential clinical importance and could open a new field of research, which should be explored.

Pneumocystis–host interaction

The development of pneumonia, which is only rarely reported in wild mammals, seems to be a rather infrequent event in the natural history of Pneumocystis infection. This view is further supported by the fine adaptation of Pneumocystis organisms to the alveolar microenvironment [7]. Marked immunosuppression constitutes the crucial factor allowing Pneumocystis pneumonia (PcP) to occur, but the severity of the pathogenic effect could be influenced by other factors.

β-D-glucan is one of the major components of the cyst wall of Pneumocystis micoorganims which interacts with alveolar macrophages and alveolar epithelial cells to stimulate the release of inflammatory mediators. An additional interaction is mediated through lactosylceramide in the host cell membrane [3,12]. Accumulating evidence indicates that innate recognition of β-glucan molecules is an important early factor driving the initiation of lung inflammation during PcP. An effective host inflammatory response is necessary to eliminate Pneumocystis infection. However, exuberant inflammation during PcP strongly promotes pulmonary injury. Severe PcP is characterized by lung inflammation involving neutrophils and CD8⁺ T cells, which induce diffuse alveolar damage and impair gas exchange, leading to respiratory failure. In fact, respiratory impairment and death are more closely correlated with the extent of lung inflammation than with the severity of the organism burden present during PcP [3].

Pneumocystis contains a glucosylceramide synthesis gene, which is necessary for organism viability. Glycosphingolipid synthesis inhibitors could be beneficial to decrease both organism burden and host inflammation during infection and offer promise as supplemental therapeutic agents [12].

Clinical spectrum of Pneumocystis infection

Pneumocystis pneumonia is one of the most serious and potentially fatal infections encountered in immonosuppressed patients. Despite advances in the treatment of HIV, mainly the development of highly active antiretroviral therapy, PcP remains the most common opportunistic infection in patients with AIDS. While the incidence of PcP among subjects with HIV infection has decreased in developed countries, the prevalence of AIDS-related PcP in developing countries remains high and poorly controlled. AIDS-related PcP continues to be an overwhelming illness among individuals who are unaware of their HIV infection, those without access to antiretroviral therapy, among subjects who are intolerant or nonadherent to therapy and in cases of failure of prophylaxis, probably related to the emergence of drug-resistant strains [13]. Currently, with the rising number of patients receiving immunosuppressive therapies for malignancies, allogeneic organ transplantations and autoimmune diseases, PcP is being recognized more and more in non-HIV-immunosuppressed individuals in developed countries [14]. The clinical presentation in HIV-infected patients may differ from that in other immunocompromised patients and its diagnosis continues to be challenging.

Cotrimoxazole, an association of trimethoprim and sulfamethoxazole, is the drug of choice for prophylaxis and therapy of Pneumocystis pneumonia. Sulfa drugs (sulfamethoxazole and dapsone) interfere with folate synthesis by competitively inhibiting the enzyme dihydropteroate synthase (DHPS). These drugs exert apparent selective pressure on P. jirovecii, as more DHPS gene mutations are observed in patients previously exposed to sulfa drugs. Although similar DHPS mutations confer resistance to sulfa drugs in other microorganisms, the association between DHPS mutations and clinical resistance in P. jirovecii is unclear because treatment failures have not yet been consistently reported in all studies [15].

Pneumocystis jirovecii colonization has recently been described in subjects with various chronic lung diseases, and accumulating evidence suggests that it may be an important clinical phenomenon [16]. P. jirovecii colonization may induce systemic inflammatory responses in patients with chronic obstructive pulmonary disease. Serum concentrations of several proinflammatory cytokines have been shown to be significantly elevated in colonized patients compared with those in noncolonized controls. Since high levels of airway and systemic inflammatory markers are associated with a faster decline in lung function, the presence of P. jirovecii in these patients could contribute to the pathophysiology of chronic obstructive pulmonary disease by inducing inflammatory changes [17].

Trends in Pneumocystis research

The recent capacity of simultaneous separating/purifying of Pneumocystis fungal cells from host cell debris, and Pneumocystis cystic forms from trophic forms while remaining infectious to the specific host, opened for the first time the possibility of characterizing the biology of each Pneumocystis stage and of substantially increasing our understanding of the Pneumocystis lifecycle [18]. In addition, progress made by the Pneumocystis Genome Project allowed the development of transcriptomic approaches to finely characterize the metabolism and biological properties of each Pneumocystis lifecycle stage [101].

The challenging issue of new potential clinical presentations of Pneumocystis infection (i.e., fetal intrauterine infection, primary infection in young children and infection in patients with chronic pulmonary diseases) will encourage investigators to clarify its role in these conditions [10].

Future clinical research should also include studying the transmission and epidemiology of PcP in populations worldwide, improving the diagnosis of PcP, improving regimens for prophylaxis and treatment in various patient populations, and determining the significance of the DHPS mutations in various populations and in different geographic locations. Furthermore, the threat of emerging resistance to available anti-Pneumocystis drugs highlights the need to continue investigating the biology of this organism in the hope of developing novel treatment strategies.

Acknowledgements

For their contributions to the success of the conference, the authors thank Robert F Miller (Royal Free and University College, UK); Cécile-Marie Aliouat (Institut Pasteur de Lille, France); El Moukhtar Aliouat (Lille Nord de France University, France); Magali Chabé (Institut Pasteur de Lille, France); François Delaporte (Université de Picardie, France); Gillez Nevez (Hôpital Augustin Morvan Centre Hospitalier Universitaire, France); Patricia Roux (Faculté de Médecine Saint-Antoine, France); Anne Totet (Université de Picardie Jules Verne, France); Annie Vitse (University of Lille, France); Olga Matos (Instituto de Higiene e Medicina Tropical, Portugal); Melanie T Cushion (University of Cincinnati College of Medicine, OH, USA), Malcolm A Finkelman (Cape Cod, Inc., MA, USA); Laurence Huang (San Francisco General Hospital, CA, USA); Andrew H Limper (Mayo Clinic, MN, USA); Edna S Kaneshiro (University of Cincinnati, OH, USA); Marta Barrionuevo (Instituto de Salud Carlos III, Spain); Carmen de la Horra (Hospital Universitario Virgen del Rocío, Spain); Sonia Gutiérrez-Rivero (Hospital Universitario Virgen del Rocío, Spain); Francisco J Medrano (Hospital Universitario Virgen del Rocío, Spain); Marco A Montes-Cano (Hospital Universitario Virgen del Rocío, Spain); Juan E Riese (Instituto de Salud Carlos III, Spain) and Philippe Hauser (Hospital Preventive Medicine, Switzerland).

Financial & competing interests disclosure

The meeting was organized in the framework of the project ‘Pneumocystis Pathogenomics: Unravelling the Colonization-to-Disease Shift’ into a Coordination Action supported by the European Commission (ERA-NET PathoGenoMics, ANR-06-PATHO-009-01) and the Spanish Ministry of Science and Innovation (SAF2009-06804-E). The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.

No writing assistance was utilized in the production of this manuscript.