Pneumocystis jirovecii is a major cause of life-threatening pneumonia in immunosuppressed patients including transplant recipients and those with HIV/AIDS, yet surprisingly little is known about the biology of this fungal pathogen. Here we report near complete genome assemblies for three Pneumocystis species that infect humans, rats and mice. Pneumocystis genomes are highly compact relative to other fungi, with substantial reductions of ribosomal RNA genes, transporters, transcription factors and many metabolic pathways, but contain expansions of surface proteins, especially a unique and complex surface glycoprotein superfamily, as well as proteases and RNA processing proteins. Unexpectedly, the key fungal cell wall components chitin and outer chain N-mannans are absent, based on genome content and experimental validation. Our findings suggest that Pneumocystis has developed unique mechanisms of adaptation to life exclusively in mammalian hosts, including dependence on the lungs for gas and nutrients and highly efficient strategies to escape both host innate and acquired immune defenses.

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Background An outbreak of 29 cases of Pneumocystis jirovecii pneumonia (PCP) occurred among renal and liver transplant recipients (RTR and LTR) in the largest Danish transplantation centre between 2007 and 2010, when routine PCP prophylaxis was not used. Methods P. jirovecii isolates from 22 transplant-cases, 2 colonized RTRs and 19 Pneumocystis-control samples were genotyped by restriction fragment length polymorphism and multi-locus sequence typing analysis. Contact tracing were used to investigate transmission. Potential risk factors were compared between PCP cases and matched non-PCP transplant patients. Results Three unique Pneumocystis genotypes were shared among 19 of the RTRs, LTRs and a colonized RTR in 3 distinct clusters, two of which overlapped temporally. In contrast, Pneumocystis-control samples harbored a wide range of genotypes. Evidence of possible nosocomial transmission was observed. Among several potential risk factors, only CMV viremia was consistently associated with PCP (P = 0.03; P = 0.009). Mycophenolate mofetile was associated with PCP risk only in the RTR population (P = 0.04). Conclusion We identified three large groups infected with unique strains of Pneumocystis and provide evidence of an outbreak profile and nosocomial transmission. LTRs may be infected in PCP outbreaks simultaneously with RTRs and by the same strains, most likely by inter-human transmission. Patients are at risk several years after transplantation, but the risk is highest during the first 6 months post-transplantation. Since patients at risk cannot be identified clinically and outbreaks cannot be predicted, six months of PCP chemoprophylaxis should be considered for all renal and liver transplant recipients.

PacBio RS, a newly emerging third-generation DNA sequencing platform, is based on a real-time, single-molecule, nano-nitch sequencing technology that can generate very long reads (up to 20-kb) in contrast to the shorter reads produced by the first and second generation sequencing technologies. As a new platform, it is important to assess the sequencing error rate, as well as the quality control (QC) parameters associated with the PacBio sequence data. In this study, a mixture of 10 prior known, closely related DNA amplicons were sequenced using the PacBio RS sequencing platform. After aligning Circular Consensus Sequence (CCS) reads derived from the above sequencing experiment to the known reference sequences, we found that the median error rate was 2.5% without read QC, and improved to 1.3% with an SVM based multi-parameter QC method. In addition, a De Novo assembly was used as a downstream application to evaluate the effects of different QC approaches. This benchmark study indicates that even though CCS reads are post error-corrected it is still necessary to perform appropriate QC on CCS reads in order to produce successful downstream bioinformatics analytical results.

Background. There have been numerous reports of clustered outbreaks of Pneumocystis pneumonia (PCP) at renal transplant centers over the past 2 decades. It has been unclear whether these outbreaks were linked epidemiologically to 1 or several unique strains, which could have implications for transmission patterns or strain virulence.Methods. Restriction fragment length polymorphism (RFLP) analysis was used to compare Pneumocystis isolates from 3 outbreaks of PCP in renal transplant patients in Germany, Switzerland, and Japan, as well as nontransplant isolates from both human immunodeficiency virus (HIV)-infected and uninfected patients.Results. Based on RFLP analysis, a single Pneumocystis strain caused pneumonia in transplant patients in Switzerland (7 patients) and Germany (14 patients). This strain was different from the strain that caused an outbreak in transplant patients in Japan, as well as strains causing sporadic cases of PCP in nontransplant patients with or without HIV infection.Conclusions. Two geographically distinct clusters of PCP in Europe were due to a single strain of Pneumocystis. This suggests either enhanced virulence of this strain in transplant patients or a common, but unidentified, source of transmission. Outbreaks of PCP can be better understood by enhanced knowledge of transmission patterns and strain variation.