A novel inhibition ELISA for the detection and monitoring of <Emphasis Type="Italic">Penicillium marneffei</Emphasis> antigen in human serum

The thermally dimorphic fungus Penicillium marneffei is a causative agent of penicilliosis marneffei, a disease considered to be an acquired immune deficiency syndrome (AIDS)-defining illness in Southeast Asia and southern China. We have developed an inhibition enzyme-linked immunosorbent assay (inh-ELISA) incorporating the yeast phase specific mannoprotein-binding monoclonal antibody 4D1 for the detection of P. marneffei infection. In our sample set, the test detected antigenemia in all 45 (100 %) patients with P. marneffei, with a mean antigen concentration of 4.32 μg/ml. No cross-reactivity in this assay was found using serum from 44 additional patients with other fungal infections, such as Aspergillus fumigatus, Cryptococcus neoformans, and Candida albicans, as well as 44 patients with bacterial infections, such as Mycobacterium tuberculosis and Streptococcus suis. Additionally, no reactivity occurred using serum from 31 human immunodeficiency virus (HIV)-infected patients without a history of fungal infections and 113 healthy controls residing in endemic areas. To investigate the potential of the inh-ELISA for disease monitoring, we followed the reduction in antigenemia in six patients who clinically responded to itraconazole and P. marneffei was no longer isolated from their blood or tissues. In contrast, we correlated increased concentrations of antigenemia in patients with relapsed P. marneffei infection with the progression of their clinical symptoms and the isolation of P. marneffei from their clinical specimens. In summary, the P. marneffei inh-ELISA is a promising new assay for the rapid diagnosis of P. marneffei, as well as a tool for evaluating clinical response and clearance of the fungus during treatment.     

Different susceptibilities of yeasts and conidia of Penicillium marneffei to nitric oxide (NO)-mediated fungicidal activity of murine macrophages

Penicillium marneffei is an important opportunistic fungal pathogen. Host defence mechanisms against P. marneffei are not fully understood. We investigated the fungicidal activity of murine peritoneal macrophages against two forms of P. marneffei, conidia and yeast cells, and the involvement of the NO-mediated killing system. Peritoneal macrophages suppressed the intracellular growth of P. marneffei yeast cells and conidia. The number of live yeast cells within macrophages was significantly reduced by activation of macrophages by interferon-gamma (IFN-γ), while a similar response was not observed with conidia. IFN-γ-induced macrophage fungicidal activity against yeast cells was mediated by NO and was almost completely inhibited by N^G-monomethyl- L -arginine ( L -NMMA), a competitive inhibitor of NO synthesis, while N^G-monomethyl- D -arginine ( D -NMMA), an optical isomer of L -NMMA, did not show any influence. NO production by macrophages stimulated with IFN-γ was significantly enhanced when these macrophages were cultured with P. marneffei yeast cells, while conidia did not enhance macrophage NO production. Furthermore, yeast cells were more susceptible to the killing effect of chemically generated NO than conidia. Our results indicate that the yeast form of P. marneffei is more sensitive to the fungicidal activity of IFN-γ-stimulated macrophages than conidia, and suggest that the different effects of two forms of P. marneffei on macrophage NO production and their different susceptibilities to NO may be reasons for the present findings.     

Effects of Talaromyces marneffei infection on mortality of HIV/AIDS patients in southern China: a retrospective cohort study

Objective

Talaromyces marneffei is an opportunistic infection with high morbidity among human immunodeficiency virus (HIV)/AIDS patients in Southeast Asia and southern China. Its effects on mortality in HIV/AIDS patients has not been clearly elucidated.

Diagnosis and Treatment of Diffusible Penicillium marneffei in Human Immunodeficiency Virus-negative Patients: A Challenge for the Physician

Penicillium marneffei infection in human immunodeficiency virus (HIV)-negative patients is addressed far less often. In this article, a small cohort of HIV-negative patients who disseminated P. marneffei infection was included. Sites of infection were found from blood culture, as subcutaneous nodules, or from lymph node biopsy. Fever, rash, swollen lymph nodes, anaemia and weight loss were common characteristics in most infected patients. The signs and symptoms are diverse and create challenges for accurate diagnosis. This paper will assist our understanding of this disease and contribute to an appropriate regime of therapy, thus improving the health of P. marneffei-positive patients.     

Identification and Purification of Specific Penicillium marneffei Antigens and Their Recognition by Human Immune Sera

Disseminated infection with the dimorphic pathogenic fungus Penicillium marneffei is increasingly seen among patients with AIDS in southeast Asian countries. Previous studies have demonstrated the presence of humoral immune responses to this fungus in patient sera; we have confirmed this work using sera from P. marneffei-infected patients (n = 21) to develop Western blots of P. marneffei cytoplasmic yeast antigen (CYA). P. marneffei CYA was then partially purified by liquid isoelectric focusing, and fractions were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting. Immunoenzyme development of the Western blots with pooled sera from patients with P. marneffei infection and with pooled sera from patients with aspergillosis (n = 20), candidiasis (n = 10), cryptococcosis (n = 9), and histoplasmosis (n = 11) revealed three antigens with relative molecular masses of 61, 54, and 50 kDa. These antigens were specifically recognized by the pooled sera from the P. marneffei-infected patients. The 61- and 54-kDa antigens were subsequently purified to homogeneity by preparative gel electrophoresis, and the 50-kDa antigen was partially purified by the same technique. N-terminal amino acid sequencing revealed that the 61-kDa antigen had a strong homology (87% identity) with the antioxidant enzyme catalase. The three antigens were then subjected to SDS-PAGE and Western blotting and to immunoenzyme development with individual patient sera; sera from 86% of P. marneffei-infected patients recognized the 61-kDa antigen, sera from 71% recognized the 54-kDa antigen, and sera from 48% recognized the 50-kDa antigen. These specifically recognized antigens are the first to be purified from P. marneffei and can be used either singly or in combination to detect antibody responses in a large percentage of individuals infected with P. marneffei.     

Sialic Acid-Dependent Recognition of Laminin by Penicillium marneffei Conidia

Immunofluorescence microscopy demonstrated that laminin bound to the surface of Penicillium marneffei conidia. Attachment of P. marneffei conidia in an adherence assay was inhibited by soluble laminin and anti-laminin antibody. N-Acetylneuraminic acid abolished adherence, indicating an interaction mediated by a sialic acid-specific lectin.     

Detection of Specific Antibodies to an Antigenic Mannoprotein for Diagnosis of Penicillium marneffei Penicilliosis

The disseminated and progressive fungal disease Penicillium marneffei penicilliosis is one of the most common infectious diseases in AIDS patients in Southeast Asia. To diagnose systemic penicilliosis, we developed an enzyme-linked immunosorbent assay (ELISA)-based antibody test with Mp1p, a purified recombinant antigenic mannoprotein of P. marneffei. Evaluation of the test with guinea pig sera against P. marneffei and other pathogenic fungi indicated that this assay was specific for P. marneffei. Clinical evaluation revealed that high levels of specific antibody were detected in two immunocompetent penicilliosis patients. Furthermore, approximately 80% (14 of 17) of the documented penicilliosis patients with human immunodeficiency virus tested positive for the specific antibody. No false-positive results were found for serum samples from 90 healthy blood donors, 20 patients with typhoid fever, and 55 patients with tuberculosis, indicating a high specificity of the test. Thus, this ELISA-based test for the detection of anti-Mp1p antibody can be of significant value as a diagnostic for penicilliosis.     

Quality evaluation of mycelial <Emphasis Type="Italic">Antrodia camphorata</Emphasis> using high-performance liquid chromatography (HPLC) coupled with diode array detector and mass spectrometry (DAD-MS)

Background  

Antrodia camphorata (AC) is an important fungus native to Taiwanese forested regions. Scientific studies have demonstrated that extracts of AC possess a variety of pharmacological functions. This study aims to identify the full profile fingerprint of nucleosides and nucleobases in mycelial AC and to assess the quality of two commercial mycelial AC products.     

An update on cutaneous infections

The spectrum of infectious diseases with potential skin involvement, either primary or secondary to systemic disease, is vast. Unusual host inflammatory responses may be elicited by certain pathogens, including reactions with an inherent capacity for misdiagnosis as neoplasia. Current, emerging, re-emerging and/or recently recognized pathogens or infections of major significance include carbapenem-resistant gram-negative bacterial infections, Ebola virus disease, measles, Mycobacterium lepromatosis as a second aetiological agent of leprosy, and Emergomyces africanus infection among those living with HIV/AIDS. The era of molecular microbiology has also seen recent changes in taxonomy, including Talaromyces marneffei (formerly Penicillium marneffei) and Cutibacterium acnes (formerly Propionobacterium acnes). Precise speciation by PCR on formalin-fixed, paraffin embedded tissue, however, remains a challenge despite recent advances in molecular microbiology. Finally, one must remain cognizant of the wide range of potential infective manifestations of the immune reconstitution inflammatory syndrome in HIV-infected individuals receiving antiretroviral therapy.     

The decreased number and function of lymphocytes is associated with Penicillium marneffei infection in HIV-negative patients

Background/purposePenicillium marneffei (P. marneffei) infection, which has been traditionally considered as an indicator of immunosuppression, is one of the most common systemic opportunistic infections in patients with AIDS. Recently, more and more P. marneffei infections have been documented in HIV-negative patients without underlying diseases, which challenges the traditional view that P. marneffei infection is an indicator of immunosuppression. We aimed to evaluate the number and function of lymphocytes in HIV-negative patients with P. marneffei infection.Methods15 HIV-negative P. marneffei-infected patients and 18 healthy controls were recruited and investigated. The number and function of lymphocytes were analyzed by flow cytometry.ResultsMost laboratory tests were within the reference ranges, except for a significant increase in total IgE in P. marneffei-infected patients. Lymphocyte subset analysis showed that the number of CD4⁺ T cells and NK cells was significantly decreased in HIV-negative marneffei-infected patients compared with healthy controls. However, almost half of the marneffei-infected patients still had normal levels of lymphocytes. A further analysis of cell function showed that the activation and proliferation of CD4⁺ T cells, the cytotoxicity of CD8⁺ T cells and NK cells, and the cytokine secretion potential of CD4⁺ T cells and NK cells were all impaired, in comparison with healthy controls.ConclusionsP. marneffei infection has to be regarded as an indicator of immunosuppression. A further investigation of cell function is required in patients with opportunistic infection, as the cell function may be impaired in this condition.     

Kinetics of capture and infection of infective larvae of trichostrongylides and free-living nematodes Panagrellus sp. by Duddingtonia flagrans

Duddingtonia flagrans, a nematode-trapping fungus, has been investigated as an agent for biological control against infective larvae of gastrointestinal nematode parasites of production animals. The initial process of nematode-trapping fungi infection is based on an interaction between the trap structure of the fungus and the surface of the nematode cuticle. This report investigates by light and scanning electron microscopy the kinetics of capture and infection during the interaction of D. flagrans with the infective larvae (L₃) of trichostrongylides and the free-living nematode Panagrellus sp. D. flagrans was cultivated for 7 days in a Petri dish containing agar–water. L₃ and Panagrellus sp. were inoculated in the Petri dishes and the samples consisting of agar–L₃–fungi and agar–Panagrellus sp.–fungi were collected after 10, 20, 30, 40, 50, 60, and 70 min and 3, 4, 5, 10, 15, 20, and 25 h of interaction. All samples were observed by light microscopy. The samples with 1, 5, 15, and 25 h of interaction were also analyzed by scanning electron microscopy. The interaction was monitored up to 25 h. An initial differentiation of predation structures was observed after 30 min of interaction. The presence of traps and of captured L₃ or Panagrellus sp. occurred after 70 min. The live captured nematodes were observed up to 3 h of interaction. However, after 4 h, all Panagrellus sp. were dead. It took 15 h of interaction for the fungus to invade the L₃, and the presence of hyphae inside the nematode near the region of penetration was evident. At this time, the hyphae had filled the whole body of Panagrellus sp. The complete occupation of the body of L₃ occurred at 20 h of interaction and with 25 h the nematode was completely damaged except for the cuticle. Although the double cuticle of L₃ slows the penetration of D. flagrans, it was possible to verify that the process of trap formation and capture occurs quickly when both nematodes were tested, suggesting that the organisms would eventually be killed once in contact with the fungi encouraging the use of the fungus as a biological control agent.     

ACE (I/D) polymorphism and response to treatment in coronary artery disease: a comprehensive database and meta-analysis involving study quality evaluation

Background  

The role of angiotensin-converting enzyme (ACE) gene insertion/deletion (I/D) polymorphism in modifying the response to treatment modalities in coronary artery disease is controversial.     

Additive effects of <Emphasis Type="Italic">LPL</Emphasis>, <Emphasis Type="Italic">APOA5</Emphasis> and <Emphasis Type="Italic">APOE</Emphasis>variant combinations on triglyceride levels and hypertriglyceridemia: results of the ICARIA genetic sub-study

Background  

Hypertriglyceridemia (HTG) is a well-established independent risk factor for cardiovascular disease and the influence of several genetic variants in genes related with triglyceride (TG) metabolism has been described, including LPL, APOA5 and APOE. The combined analysis of these polymorphisms could produce clinically meaningful complementary information.     

N-acetyltransferase 2 (NAT2) gene polymorphisms in Parkinson's disease

Background  

Parkinson's disease (PD) is a movement disorder caused by the degeneration of dopaminergic neurons in the substantia nigra of the midbrain. The molecular basis of this neural death is unknown, but genetic predisposition and environmental factors may cause the disease. Sequence variations in N-acetyltransferase 2 (NAT2) gene leading to slow acetylation process have been associated with PD, but results are contradictory.     

Effective quantitative real-time polymerase chain reaction analysis of the parkin gene (<Emphasis Type="Italic">PARK2</Emphasis>) exon 1–12 dosage

Background  

One of the causes of Parkinson's disease is mutations in the PARK2 gene. Deletions and duplications of single exons or exon groups account for a large proportion of the gene mutations. Direct detection of these mutations can be used for the diagnosis of Parkinson's disease.     

1Novel <Emphasis Type="Italic">MEFV</Emphasis> transcripts in Familial Mediterranean fever patients and controls

Background  

Familial Mediterranean fever is a recessive autoinflammatory disease frequently encountered in Armenians, Jews, Arabs and Turks. The MEFV gene is responsible for the disease. It encodes a protein called pyrin/marenostrin involved in the innate immune system. A large number of clinically diagnosed FMF patients carry only one MEFV mutation. This study aims at studying the MEFV gene splicing pattern in heterozygous FMF patients and healthy individuals, in an attempt to understand the mechanism underlying the disease in these patients.     

No evidence for association between <Emphasis Type="Italic">tau</Emphasis>gene haplotypic variants and susceptibility to Creutzfeldt-Jakob disease

Background  

A polymorphism at codon 129 of the prion protein gene (PRNP) is the only well-known genetic risk factor for Creutzfeldt-Jakob disease (CJD). However, there is increasing evidence that other loci outside the PRNP open reading frame might play a role in CJD aetiology as well.     

High prevalence of germline <Emphasis Type="Italic">STK11</Emphasis>mutations in Hungarian Peutz-Jeghers Syndrome patients

Background  

Peutz-Jeghers syndrome (PJS) is a rare autosomal dominantly inherited disease characterized by gastrointestinal hamartomatous polyposis and mucocutaneous pigmentation. The genetic predisposition for PJS has been shown to be associated with germline mutations in the STK11/LKB1 tumor suppressor gene. The aim of the present study was to characterize Hungarian PJS patients with respect to germline mutation in STK11/LKB1 and their association to disease phenotype.     

Sphingomyelin phosphodiesterase-1 (SMPD1) coding variants do not contribute to low levels of high-density lipoprotein cholesterol

Background  

Niemann-Pick disease type A and B is caused by a deficiency of acid sphingomyelinase due to mutations in the sphingomyelin phosphodiesterase-1 (SMPD1) gene. In Niemann-Pick patients, SMPD1 gene defects are reported to be associated with a severe reduction in plasma high-density lipoprotein (HDL) cholesterol.     

Polymorphism of SERPINE2 gene is associated with pulmonary emphysema in consecutive autopsy cases

Background  

The SERPINA1, SERPINA3, and SERPINE2 genes, which encode antiproteases, have been proposed to be susceptible genes for of chronic obstructive pulmonary disease (COPD) and related phenotypes. Whether they are associated with emphysema is not known.