Immunization with Pneumocystis carinii A121-85 antigen activates immune function against P. carinii.

BACKGROUND: Pneumocystis pneumonia (PcP), which is caused by Pneumocystis carinii, is a life-threatening infection that affects immunocompromised individuals. Unfortunately, chemoprophylaxis and dapsone are only effective for half of the patients with PcP, indicating that additional preventive methods are needed. We predicated the pneumocystis surface protein A12 sequence 1-85 by DNAStar software and BepiPred, and identified it as a potential vaccine candidate by bioresearch. METHODS: We used recombinant A121-85 as antigen to immunized mice and detected serum titer of IgG, expression of inflammatory factors by EILSA, qRT-PCR and flow cytometry. RESULTS: Our results showed that immunization with recombinant A121-85 increased the serum titer of IgG, promoted the secretion of T lymphocytes, increased the expression of inflammatory factors, and elevated lung inflammatory injury in mice. CONCLUSIONS: Our findings suggest that A121-85 is a potential vaccine target for preventing Pneumocystis carinii. The evaluation of A121-85-elicited antibodies in the prevention of PcP in humans deserves further investigation.

Characteristics and risk factors for Pneumocystis jirovecii pneumonia in patients with idiopathic membranous nephropathy.

Pneumocystis jirovecii pneumonia (PCP) is a life-threatening opportunistic infection in idiopathic membranous nephropathy (IMN) patients, who are treated with immunosuppressive drugs. However, the risk factors of infection and their prognosis are rarely investigated. We aimed to characterize the clinical manifestations of PCP in patients with IMN, and to understand their risk factors, so that we can provide early warnings to patients with high risk and potential poor prognosis. We conducted a retrospective observational study of IMN patients in a referral center in China, from Jan 2012 to Dec 2018. Clinical and laboratory data were collected separately at the time of IMN and PCP diagnosis. Patients with PCP were matched to those without by gender and age at a ratio of 1:4. The risk factors and prognostic factors were determined by univariate and multivariate logistic regression analysis. A total of 879 patients with IMN were included, with a median follow-up of 267 (interquartile range (IQR) 64,842) days. In total, 26 (2.96%) of them were diagnosed with PCP. The infection rate increased to 3.87% among patients who received corticosteroids, and it further increased to 5.49% in those received over 0.5mg/kg prednisone. Univariate analysis indicated that initial usage of corticosteroids, use of cyclophosphamide, reduced estimated glomerular filtration rate (eGFR), and higher 24-h proteinuria were related to the PCP susceptibility. Multivariate analysis revealed that corticosteroid treatment and reduced eGFR increased the risk of the Pneumocystis jirovecii infection. The case fatality rate of the PCP patients was 23.08%, and increased to 75% among patients requiring invasive ventilation. Univariate analysis indicated that pulmonary insufficiency, invasive ventilation, decreased eGFR, and increased lactate dehydrogenase at presentation were linked to poor prognosis. PCP is not rare in patients with IMN, especially those on corticosteroids, and presented with decreased eGFR. Considering the high case fatality rate, further studies are in need for prevention and management of these patients.

Diagnosing Pneumocystis jirovecii pneumonia: A review of current methods and novel approaches.

Pneumocystis jirovecii can cause life-threatening pneumonia in immunocompromised patients. Traditional diagnostic testing has relied on staining and direct visualization of the life-forms in bronchoalveolar lavage fluid. This method has proven insensitive, and invasive procedures may be needed to obtain adequate samples. Molecular methods of detection such as polymerase chain reaction (PCR), loop-mediated isothermal amplification (LAMP), and antibody-antigen assays have been developed in an effort to solve these problems. These techniques are very sensitive and have the potential to detect Pneumocystis life-forms in noninvasive samples such as sputum, oral washes, nasopharyngeal aspirates, and serum. This review evaluates 100 studies that compare use of various diagnostic tests for Pneumocystis jirovecii pneumonia (PCP) in patient samples. Novel diagnostic methods have been widely used in the research setting but have faced barriers to clinical implementation including: interpretation of low fungal burdens, standardization of techniques, integration into resource-poor settings, poor understanding of the impact of host factors, geographic variations in the organism, heterogeneity of studies, and limited clinician recognition of PCP. Addressing these barriers will require identification of phenotypes that progress to PCP and diagnostic cut-offs for colonization, generation of life-form specific markers, comparison of commercial PCR assays, investigation of cost-effective point of care options, evaluation of host factors such as HIV status that may impact diagnosis, and identification of markers of genetic diversity that may be useful in diagnostic panels. Performing high-quality studies and educating physicians will be crucial to improve the rates of diagnosis of PCP and ultimately to improve patient outcomes.

Veno-Venous Extracorporeal Membrane Oxygenation for Severe Pneumocystis jirovecii Pneumonia in an Immunocompromised Patient without HIV Infection.

Pneumocystis jirovecii pneumonia (PJP) occurs in immunocompromised hosts and is classified as PJP with human immunodeficiency virus (HIV) infection (HIV-PJP) and PJP without HIV infection (non-HIV PJP). Non-HIV PJP rapidly progresses to respiratory failure compared with HIV-PJP possibly due to the difference in immune conditions; namely, the prognosis of non-HIV PJP is worse than that of HIV PJP. However, the diagnosis of non-HIV PJP at the early stage is difficult. Herein, we report a case of severe non-HIV PJP successfully managed with veno-venous extracorporeal membrane oxygenation (V-V ECMO). A 54-year-old woman with neuromyelitis optica was treated with oral corticosteroid, azathioprine, and methotrexate. She admitted to our hospital for fever, dry cough, and dyspnea which developed a week ago. On admission, she required endotracheal intubation and invasive ventilation for hypoxia. A chest computed tomography (CT) scan revealed ground-glass opacity and consolidation in the both lungs. Grocott staining and PCR analysis of bronchoalveolar lavage fluid indicated the presence of fungi and Pneumocystis jirovecii, respectively, whereas serum HIV-antibody was negative. The patient was thus diagnosed with non-HIV PJP and was treated with intravenous pentamidine and corticosteroid pulse therapy for PJP. However, hypoxia was worsened; consequently, V-V ECMO assistance was initiated on day 7. The abnormal chest CT findings and hypoxia were gradually improved. The V-V ECMO support was successfully discontinued on day 14 and mechanical ventilation was discontinued on day 15. V-V ECMO could be a useful choice for respiratory assistance in severe cases of PJP among patients without HIV infection.

Outbreak-Causing Fungi: Pneumocystis jirovecii.

Pneumocystis jirovecii pneumonia (PCP) is an important cause of morbidity in immunocompromised patients, with a higher mortality in non-HIV than in HIV patients. P. jirovecii is one of the rare transmissible pathogenic fungi and the only one that depends fully on the host to survive and proliferate. Transmissibility among humans is one of the main specificities of P. jirovecii. Hence, the description of multiple outbreaks raises questions regarding preventive care management of the disease, especially in the non-HIV population. Indeed, chemoprophylaxis is well codified in HIV patients but there is a trend for modifications of the recommendations in the non-HIV population. In this review, we aim to discuss the mode of transmission of P. jirovecii, identify published outbreaks of PCP and describe molecular tools available to study these outbreaks. Finally, we discuss public health and infection control implications of PCP outbreaks in hospital setting for in- and outpatients.

Clinical, genetic and immunological characteristics of 40 Chinese patients with CD40 ligand deficiency.

CD40 ligand (CD40L) deficiency is a rare but life-threatening primary immunodeficiency caused by mutations in the CD40L gene. Here, we investigated a cohort of 40 genetically diagnosed CD40L-deficient patients from the Chinese mainland, analysed their clinical and genetic data, and examined CD40L expression, the proportion of T cell subsets, B cell subsets and T follicular helper (Tfh) cells. The aim was to provide a complete picture of CD40L deficiency. Initial presentations of the patient cohort mainly involved recurrent fever (47.5%) and sinopulmonary infection (42.5%). Life-threatening infections (42.5%), caused by various pathogens, were the most serious threats faced by CD40L-deficient patients, while neutropenia (57.5%) remained the most common complication. Opportunistic infections, including Pneumocystis carinii pneumonia and invasive fungal disease associated with Talaromyces marneffei, were also common in the cohort. In addition, seven patients (17.5%) suffered BCGitis/BCGosis, which is a major problem facing a planned immunization programme in China. It was intriguing that reduced IgM levels were observed in 12.5% of patients, while normal or elevated IgA levels were shown in 47.5% of patients. Thirty-seven unique mutations were identified in 40 patients; of these, 10 were novel. Furthermore, we observed a lower percentage of NK cells, Tfh cells, and central memory CD4+ T cells, and an extremely small class-switched memory B cell population, in CD40L-deficient patients. Patients who underwent hematopoietic stem cell transplantation experienced better disease remission. Taken together, our data establish the largest database about CD40L deficiency in China and provide genetic, immunologic and clinical information about Chinese CD40L-deficient patients.

Serum Soluble Interleukin-2 Receptor Is a Biomarker for Pneumocystis jirovecii Pneumonia among Patients with Rheumatoid Arthritis under Methotrexate Therapy.

Rheumatoid arthritis (RA) is a chronic autoimmune disease characterized by systemic joint inflammation and may manifest as interstitial pneumonia (IP). Methotrexate (MTX) is one of the main therapeutic drugs used for RA, but MTX could cause severe side effects, including Pneumocystis jirovecii pneumonia (PCP) and IP. Owing to similar symptoms, it is sometimes difficult to discriminate MTX therapy-associated PCP (MTX-PCP) and MTX therapy-associated IP (MTX-IP). Soluble interleukin-2 receptor (sIL-2R) is considered a marker of T-cell activation, and serum sIL-2R levels are elevated in RA and PCP. This led us to hypothesize that serum sIL-2R is a potential biomarker for discriminating MTX-PCP and MTX-IP. Accordingly, we carried out a retrospective analysis of 20 MITX-PCP cases, 30 MTX-IP cases, and as controls, 16 patients with RA-associated IP (RA-IP) and 13 patients with PCP without MTX treatment (PCP group). C-reactive protein and alveolar-arterial oxygen differences were higher in the MTX-PCP group than those in the RA-IP and MTX-IP groups. Importantly, serum levels of sIL-2R in MTX-PCP were significantly higher than those in other three groups. Based on the receiver operating characteristic curve, the cut-off level of sIL-2R resulting in the highest diagnostic accuracy for MTX-PCP was 1,311.5 U/mL, discriminating between MTX-PCP and other groups with 91.7% sensitivity and 78.6% specificity. Thus, patients with MTX-PCP show a higher degree of systemic inflammation, severe hypoxemia, and increased sIL-2R levels compared with those in MTX-IP cases. In conclusion, serum sIL-2R could be a biomarker for PCP diagnosis among patients with RA under MTX therapy.

Diffusion of Pneumocystis jirovecii in the surrounding air of patients with Pneumocystis colonization: frequency and putative risk factors.

In a prospective bicentric study, Pneumocystis jirovecii excretion and diffusion was explored in air samples collected in the rooms occupied by 17 Pneumocystis-colonized patients. P. jirovecii DNA was detected by real-time PCR in the air collected from 3 patients' rooms (17.6%), with identical genotypes in corresponding clinical and air samples. Pneumocystis DNA was detected for 2/3 patients with autoimmune disease treated with corticosteroids versus 1/6 patients with hematologic disease and 0/5 kidney transplant recipients. These data confirm the possible excretion of the fungus by Pneumocystis-colonized patients and thus bring additional arguments for the prevention of airborne transmission in hospital wards.

Pulmonary co-infections by Pneumocystis jirovecii and Aspergillus fumigatus in non-HIV patients: A report of two cases and literature review.

Pneumocystis jirovecii is the causative agent of Pneumocystis pneumonia (PcP), a common and often life-threatening opportunistic infection in HIV-infected patients. However, non-HIV, immunocompromised patients are at risk of PcP as well, whereas the mortality appears to be higher among these patients. Pneumocystis co-infections with other microorganisms are less frequent and only sparse reports of combined PcP and invasive pulmonary fungal infections exist in the literature, especially in the non-HIV patients. Two cases of pulmonary co-infections by P. jirovecii and Aspergillus fumigatus are presented. Both patients were non-HIV infected, the first one was suffering from crescentic IgA nephropathy under immunosuppressive treatment and the second from resistant non-Hodgkin lymphoma under chemotherapy. Both patients were treated with intravenous trimethoprim/sulphamethoxazole (TMP/SMX) combined with voriconazole. The first patient showed gradual clinical improvement while the outcome for the second patient was unfavourable. In addition, a literature review of the previous published cases of co-infection by P. jirovecii and other fungi in non-HIV patients was performed. Our target was to provide comprehensive information on this kind of infections, highlighting the importance of clinical suspicion.

Evidence for a Pneumocystis carinii Flo8-like transcription factor: insights into organism adhesion.

Pneumocystis carinii (Pc) adhesion to alveolar epithelial cells is well established and is thought to be a prerequisite for the initiation of Pneumocystis pneumonia. Pc binding events occur in part through the major Pc surface glycoprotein Msg, as well as an integrin-like molecule termed PcInt1. Recent data from the Pc sequencing project also demonstrate DNA sequences homologous to other genes important in Candida spp. binding to mammalian host cells, as well as organism binding to polystyrene surfaces and in biofilm formation. One of these genes, flo8, a transcription factor needed for downstream cAMP/PKA-pathway-mediated activation of the major adhesion/flocculin Flo11 in yeast, was cloned from a Pc cDNA library utilizing a partial sequence available in the Pc genome database. A CHEF blot of Pc genomic DNA yielded a single band providing evidence this gene is present in the organism. BLASTP analysis of the predicted protein demonstrated 41 % homology to the Saccharomyces cerevisiae Flo8. Northern blotting demonstrated greatest expression at pH 6.0-8.0, pH comparable to reported fungal biofilm milieu. Western blot and immunoprecipitation assays of PcFlo8 protein in isolated cyst and tropic life forms confirmed the presence of the cognate protein in these Pc life forms. Heterologous expression of Pcflo8 cDNA in flo8Δ-deficient yeast strains demonstrated that the Pcflo8 was able to restore yeast binding to polystyrene and invasive growth of yeast flo8Δ cells. Furthermore, Pcflo8 promoted yeast binding to HEK293 human epithelial cells, strengthening its functional classification as a Flo8 transcription factor. Taken together, these data suggest that PcFlo8 is expressed by Pc and may exert activity in organism adhesion and biofilm formation.

Relationship Between Pneumocystis carinii Burden and the Degree of Host Immunosuppression in an Airborne Transmission Experimental Model.

To quantitatively assess the risk of contamination by Pneumocystis depending on the degree of immunosuppression (ID) of the exposed rat hosts, we developed an animal model, where rats went through different doses of dexamethasone. Then, natural and aerial transmission of Pneumocystis carinii occurred during cohousing of the rats undergoing gradual ID levels (receivers) with nude rats developing pneumocystosis (seeders). Following contact between receiver and seeder rats, the P. carinii burden of receiver rats was determined by toluidine blue ortho staining and by qPCR targeting the dhfr monocopy gene of this fungus. In this rat model, the level of circulating CD4(+) and CD8(+) T lymphocytes remained significantly stable and different for each dose of dexamethasone tested, thus reaching the goal of a new stable and gradual ID rat model. In addition, an inverse relationship between the P. carinii burden and the level of circulating CD4(+) or CD8(+) T lymphocytes was evidenced. This rat model may be used to study other opportunistic pathogens or even co-infections in a context of gradual ID.

Pneumocystis jirovecii--from a commensal to pathogen: clinical and diagnostic review.

Pneumocystis pneumonia is an opportunistic disease caused by invasion of unicellular fungus Pneumocystis jirovecii. Initially, it was responsible for majority of morbidity and mortality cases among HIV-infected patients, which later have been reduced due to the introduction of anti-retroviral therapy, as well as anti-Pneumocystis prophylaxis among these patients. Pneumocystis pneumonia, however, is still a significant cause of mortality among HIV-negative patients being under immunosuppression caused by different factors, such as transplant recipients as well as oncologically treated ones. The issue of pneumocystosis among these people is particularly emphasized in the article, since rapid onset and fast progression of severe symptoms result in high mortality rate among these patients, who thereby represent the group of highest risk of developing Pneumocystis pneumonia. In contrast, fungal invasion in immunocompetent people usually leads to asymptomatic colonization, which frequent incidence among healthy infants has even suggested the possibility of its association with sudden unexpected infant death syndrome. In the face of emerging strains with different epidemiological profiles resulting from genetic diversity, including drug-resistant genotypes, the colonization phenomenon desires particular attention, discussed in this article. We also summarize specific and sensitive methods, required for detection of Pneumocystis invasion and for distinguish colonization from the disease.

A Case of Pneumonia Caused by Pneumocystis jirovecii Resistant to Trimethoprim-Sulfamethoxazole.

A 50-year-old male visited the outpatient clinic and complained of fever, poor oral intake, and weight loss. A chest X-ray demonstrated streaky and fibrotic lesions in both lungs, and chest CT revealed multifocal peribronchial patchy ground-glass opacities with septated cystic lesions in both lungs. Cell counts in the bronchoalveolar lavage fluid revealed lymphocyte-dominant leukocytosis, and further analysis of lymphocyte subsets showed a predominance of cytotoxic T cells and few T helper cells. Video-assisted wedge resection of the left upper lobe was performed, and the histologic examination was indicative of a Pneumocystis jirovecii infection. Trimethoprim-sulfamethoxazole (TMP-SMX) was orally administered for 3 weeks; however, the patient complained of cough, and the pneumonia was aggravated in the follow-up chest X-ray and chest CT. Molecular studies demonstrated mutations at codons 55 and 57 of the dihydropteroate synthase (DHPS) gene, which is associated with the resistance to TMP-SMX. Clindamycin-primaquine was subsequently administered for 3 weeks replacing the TMP-SMX. A follow-up chest X-ray showed that the pneumonia was resolving, and the cough was also alleviated. A positive result of HIV immunoassay and elevated titer of HCV RNA indicated HIV infection as an underlying condition. This case highlights the importance of careful monitoring of patients with P. jirovecii pneumonia (PCP) during the course of treatment, and the molecular study of DHPS mutations. Additionally, altering the anti-PCP drug utilized as treatment must be considered when infection with drug-resistant P. jirovecii is suspected. To the best of our knowledge, this is the first case of TMP-SMX-resistant PCP described in Korea.

Scalable preparation and differential pharmacologic and toxicologic profiles of primaquine enantiomers.

Hematotoxicity in individuals genetically deficient in glucose-6-phosphate dehydrogenase (G6PD) activity is the major limitation of primaquine (PQ), the only antimalarial drug in clinical use for treatment of relapsing Plasmodium vivax malaria. PQ is currently clinically used in its racemic form. A scalable procedure was developed to resolve racemic PQ, thus providing pure enantiomers for the first time for detailed preclinical evaluation and potentially for clinical use. These enantiomers were compared for antiparasitic activity using several mouse models and also for general and hematological toxicities in mice and dogs. (+)-(S)-PQ showed better suppressive and causal prophylactic activity than (-)-(R)-PQ in mice infected with Plasmodium berghei. Similarly, (+)-(S)-PQ was a more potent suppressive agent than (-)-(R)-PQ in a mouse model of Pneumocystis carinii pneumonia. However, at higher doses, (+)-(S)-PQ also showed more systemic toxicity for mice. In beagle dogs, (+)-(S)-PQ caused more methemoglobinemia and was toxic at 5 mg/kg of body weight/day given orally for 3 days, while (-)-(R)-PQ was well tolerated. In a novel mouse model of hemolytic anemia associated with human G6PD deficiency, it was also demonstrated that (-)-(R)-PQ was less hemolytic than (+)-(S)-PQ for the G6PD-deficient human red cells engrafted in the NOD-SCID mice. All these data suggest that while (+)-(S)-PQ shows greater potency in terms of antiparasitic efficacy in rodents, it is also more hematotoxic than (-)-(R)-PQ in mice and dogs. Activity and toxicity differences of PQ enantiomers in different species can be attributed to their different pharmacokinetic and metabolic profiles. Taken together, these studies suggest that (-)-(R)-PQ may have a better safety margin than the racemate in human.

Therapeutic efficacies of chitosan against Pneumocystis pneumonia of immunosuppressed rat.

This study was designed to investigate the therapeutic efficacy of chitosan on Pneumocystis pneumonia (PCP) in immunosuppressed rats. The PCP rat model was established using intramuscular injections of dexamethasone sodium phosphate. To estimate treatment effects of chitosan on rat PCP, weight gain, lung weight, lung weight/body weight (LW/BW) ratio and per cent survival were measured and the HSP70 mRNA expression of Pneumocystis carinii was detected using real-time PCR analysis. Rat lung tissues were stained with HE, and their pathological changes, inflammatory cells and alveolar macrophages were observed by light microscopy. Rat lymphocyte numbers and the concentrations of IL-10, IFN-γ and TNF-α were measured by flow cytometry and ELISA analysis. Additionally, the ultrastructure of P. carinii was examined by electron microscopy to evaluate the effects of chitosan on the protist. Our results demonstrated that chitosan has some apparent treatment effects on rat PCP by reducing HSP70 mRNA expression and lung inflammation, increasing the concentrations of IL-10 and IFN-γ as well as CD4(+) T-lymphocyte numbers, reducing the CD8(+) T-lymphocyte numbers and the concentration of TNF-α and inducing significant ultrastructural damage to P. carinii. Although its precise therapeutic mechanism has yet to be determined, these results lay a theoretical foundation for PCP chitosan therapy.

Primary prophylaxis of bacterial infections and Pneumocystis jirovecii pneumonia in patients with hematological malignancies and solid tumors : guidelines of the Infectious Diseases Working Party (AGIHO) of the German Society of Hematology and Oncology (DGHO).

Bacterial infections are the most common cause for treatment-related mortality in patients with neutropenia after chemotherapy. Here, we discuss the use of antibacterial prophylaxis against bacteria and Pneumocystis pneumonia (PCP) in neutropenic cancer patients and offer guidance towards the choice of drug. A literature search was performed to screen all articles published between September 2000 and January 2012 on antibiotic prophylaxis in neutropenic cancer patients. The authors assembled original reports and meta-analysis from the literature and drew conclusions, which were discussed and approved in a consensus conference of the Infectious Disease Working Party of the German Society of Hematology and Oncology (AGIHO). Antibacterial prophylaxis has led to a reduction of febrile events and infections. A significant reduction of overall mortality could only be shown in a meta-analysis. Fluoroquinolones are preferred for antibacterial and trimethoprim-sulfamethoxazole for PCP prophylaxis. Due to serious concerns about an increase of resistant pathogens, only patients at high risk of severe infections should be considered for antibiotic prophylaxis. Risk factors of individual patients and local resistance patterns must be taken into account. Risk factors, choice of drug for antibacterial and PCP prophylaxis and concerns regarding the use of prophylactic antibiotics are discussed in the review.

Pneumocystis jirovecii colonization in chronic pulmonary disease.

Pneumocystis jirovecii causes pneumonia in immunosuppressed individuals. However, it has been reported the detection of low levels of Pneumocystis DNA in patients without signs and symptoms of pneumonia, which likely represents colonization. Several studies performed in animals models and in humans have demonstrated that Pneumocystis induces a local and a systemic response in the host. Since P jirovecii colonization has been found in patients with chronic pulmonary diseases it has been suggested that P jirovecii may play a role in the physiopathology and progression of those diseases. In this report we revise P. jirovecii colonization in different chronic pulmonary diseases such us, chronic obstructive pulmonary disease, interstitial lung diseases, cystic fibrosis and lung cancer.

[Pneumocystis jiroveci pneumonia: first documented case in Libreville, Gabon]. / Pneumonie à pneumocystis jiroveci: premier cas documenté à Libreville (Gabon).

The purpose of this report is to describle the first documented case of pneumocystosis in Gabon. Diagnosis was confirmed based on microbiological methods in a 59-year-old immunodepressed HIV-positive man. Diagnosis was further documented by observation of Pneumocystis jiroveci cysts in the broncho-alveolar lavage (BAL) after Gomori-Grocott color reaction. The patient responded well to treatment with Cotrimoxazole, corticoids and oxygen.