Gene/Protein Disease Symptom Drug Enzyme Compound
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Chytridiomycosis is a recently identified fungal disease associated with global population declines of frogs. Although the fungus, Batrachochytrium dendrobatidis, is considered an emerging pathogen, little is known about its population genetics, including the origin of the current epidemic and how this relates to the dispersal ability of the fungus. In this study, we use multilocus sequence typing to examine genetic diversity and relationships among 35 fungal strains from North America, Africa and Australia. Only five variable nucleotide positions were detected among 10 loci (5918 bp). This low level of genetic variation is consistent with the description of B. dendrobatidis as a recently emerged disease agent. Fixed (i.e. 100%) or nearly fixed frequencies of heterozygous genotypes at two loci suggested that B. dendrobatidis is diploid and primarily reproduces clonally. In contrast to the lack of nucleotide polymorphism, electrophoretic karyotyping of multiple strains demonstrated a number of chromosome length polymorphisms.
Mol Ecol 2003 Feb
PMID:Multilocus sequence typing suggests the chytrid pathogen of amphibians is a recently emerged clone. 1253 90

Since the express-diagnostics of mycoses in immune-deficit patients still remains an acute problem, we developed an effective test system (Kan-Am) to detect DNA Candida albicans, which is a leader in the list of causative agents of candidosis. A comparison study of three PCR-systems used to detect a broad spectrum of fungoid pathogens was carried out, and a universal system (FungAm), which ensures the detection of DNAs of above 78 strains of 25 types of pathogenic fungi, was selected. The results of clinical testing of the species-specific and universal PCR-systems are well confirmed by the culture method, and they are indicative of the efficacy of applying them for the diagnostics of mycoses in neonatology. The use of the mentioned systems is a promising factor for the express-diagnostics of mycoses in immunodeficiency patients. The high sensitivity of the method makes it possible to detect 10 to 100 cells of a causative agent in 100 mcl of the examined biological material, which is compatible with the culture method. A kit of dry reagents (IonoMix) designed for an accelerated sample preparation and isolation, from them, of DNAs on the basis of Chelex-100 and of proteinase K was worked out; the kit is portable and meant for a long-term storing.
Mol Gen Mikrobiol Virusol 2003
PMID:[Development of a PCR-based method for diagnostics of mycoses]. 1280 Jul 74

Invasive mycoses are associated with a high mortality rate, and their incidence is increased in immunologically deficient patients. From a diagnostic and therapeutic perspective, these infections represent a significant challenge to medicine. In addition to new antifungal agents, drug combinations are an important therapeutic resource, which might be exploited clinically, owing to the multiplicity of fungal targets against which currently available agents are active. In this review, we examine the experimental data regarding the combination of conventional antifungal agents with cytokines, antibacterial agents, calcineurin inhibitors and drugs under development characterized by novel mechanisms of action.
Trends Mol Med 2003 Jun
PMID:Molecular targeted treatments for fungal infections: the role of drug combinations. 1282 16

As the cowpea rust fungus penetrates the wall of a cowpea epidermal cell, resistant and susceptible plants exhibit different ultrastructural and cytochemical changes within the epidermal protoplast. To examine plant gene expression at this stage of infection, cytoplasm was extracted from individual inoculated or uninoculated epidermal cells before the fungal penetration peg reached the cell lumen. Initial differential colony hybridization screening of an expressed sequence tag library constructed from globally amplified cDNAs generated from the inoculated resistant cells resulted in 80 clones (out of 835) with a differential hybridization pattern. Further slot-blot screening and screening of the amplified cDNAs generated from inoculated or uninoculated, resistant or susceptible cells revealed 28 separate genes, mostly with no matching sequences in the databases, that were up-regulated in response to the growth of the fungus through the wall of resistant or susceptible cells. Five genes, including those coding for beta- and alpha-tubulin, were found to be down-regulated specifically in inoculated, susceptible cells, and five were specifically up-regulated in inoculated, resistant cells, including a PR-10 homolog and a phenylalanine ammonia-lyase gene. Probing the amplified cDNAs from each cell type for the expression of cell death-related genes revealed that an LLS1 homolog (vuLLS1), cloned from cowpea, was up-regulated by infection in both resistant and susceptible cells and that a homolog of HSR203J was differentially up-regulated in resistant cells. These data show that changes in gene expression predicting the subsequent expression of susceptibility or hypersensitive resistance to fungal infection occur prior to the fungus entering the cell lumen.
Mol Plant Microbe Interact 2003 Sep
PMID:cDNAs generated from individual epidermal cells reveal that differential gene expression predicting subsequent resistance or susceptibility to rust fungal infection occurs prior to the fungus entering the cell lumen. 1297 7

Pathogenesis-related (PR) proteins are plant proteins that are induced in response to pathogen attack. PR proteins are grouped into independent families based on their sequences and properties. The PR-4 family comprises class I and class II chitinases. We have isolated a full-length cDNA encoding a chitinase from maize which shares a high degree of nucleotide and amino acid sequence homology with the class II chitinases of the PR-4 family of PR proteins. Our results indicate that fungal infection, and treatment either with fungal elicitors or with moniliformin, a mycotoxin produced by the fungus Fusarium moniliforme, increase the level of ZmPR4 mRNA. In situ mRNA hybridization analysis in sections obtained from fungus-infected germinating embryos revealed that ZmPR4 mRNA accumulation occurs in those cell types that first establish contact with the pathogen. ZmPR4 mRNA accumulation is also stimulated by treatment with silver nitrate whereas the application of the hormones gibberellic acid or acetylsalicylic acid has no effect. Wounding, or treatment with abscisic acid or methyl jasmonate, results in accumulation of ZmPR4 mRNA in maize leaves. Furthermore, the ZmPR4 protein was expressed in Escherichia coli, purified and used to obtain polyclonal antibodies that specifically recognized ZmPR4 in protein extracts from fungus-infected embryos. Accumulation of ZmPR4 mRNA in fungus-infected maize tissues was accompanied by a significant accumulation of the corresponding protein. The possible implications of these findings as part of the general defence response of maize plants against pathogens are discussed.
Plant Mol Biol 2003 Jul
PMID:Fungus- and wound-induced accumulation of mRNA containing a class II chitinase of the pathogenesis-related protein 4 (PR-4) family of maize. 1367 64

Improvements in two-dimensional gel electrophoresis, mass spectrometry, and bioinformatics provide new tools to characterize proteins involved in a physiological process, such as the immune response of the insect model Drosophila melanogaster. Profiling of the proteins present in the hemolymph (insect blood) of noninfected flies versus flies infected with bacteria or fungi was performed by two-dimensional gel electrophoresis, silver or Coomassie staining, and image analysis. Through this differential analysis, more than 70 out of 160 spots were up- or down-regulated by at least 5-fold after microbial infection. Coomassie staining, in-gel digestion, and database searches yielded the identity of a series of proteins that are directly involved in the Drosophila immune system. This included proteases, protease inhibitors, and recognition molecules such as prophenoloxydase-activating enzymes, serpins, and Gram-negative binding protein-like. Proteins with a potential function in the immune response were also identified, such as an odorant binding protein, peptidylglycine alpha-hydroxylating monooxygenase, and transferrin, affording new candidates for further investigation of innate immune mechanisms. Moreover, several molecules resulting from the cleavage of proteins were detected after the fungal infection. Altogether, this first differential proteomic analysis of the immune response of Drosophila paves the way for the study of proteins affected during innate immunity.
Mol Cell Proteomics 2004 Feb
PMID:Proteomic analysis of the systemic immune response of Drosophila. 1464 1

Tinea pedis is a chronic fungal infection of the feet, very often observed in patients who are immuno-suppressed or have diabetes mellitus. The practicing allergist may be called upon to treat this disease for various reasons. Sometimes tinea infection may be mistaken for atopic dermatitis or allergic eczema. In other patients, tinea pedis may complicate allergy and asthma and may contribute to refractory atopic disease. Patients with recurrent cellulitis may be referred to the allergist/immunologist for an immune evaluation and discovered to have tinea pedis as a predisposing factor. From a molecular standpoint, superficial fungal infections may induce a type2 T helper cell response (Th2) that can aggravate atopy. Th2 cytokines may induce eosinophil recruitment and immunoglobulin E (IgE) class switching by B cells, thereby leading to exacerbation of atopic conditions. Three groups of fungal pathogens, referred to as dermatophytes, have been shown to cause tinea pedis: Trychophyton sp, Epidermophyton sp, and Microsporum sp. The disease manifests as a pruritic, erythematous, scaly eruption on the foot and depending on its location, three variants have been described: interdigital type, moccasin type, and vesiculobullous type. Tinea pedis may be associated with recurrent cellulitis, as the fungal pathogens provide a portal for bacterial invasion of subcutaneous tissues. In some cases of refractory asthma, treatment of the associated tinea pedis infection may induce remission in airway disease. Very often, protracted topical and/or oral antifungal agents are required to treat this often frustrating and morbid disease. An evaluation for underlying immuno-suppression or diabetes may be indicated in patients with refractory disease.
Clin Mol Allergy 2004 Mar 29
PMID:Dermatology for the practicing allergist: Tinea pedis and its complications. 1505 29

In the absence of antibodies specific for lymphatic vessels, analysis of lymphatic vessels within different tissues has been widely performed with light microscopic and, most importantly, electron microscopic techniques. In regard to lymphatic vessels in the ocular globe and the periocular structures, controversy remains about the specific distribution of lymphatic channels. It is postulated that bulbar and retrobulbar tissues are devoid of lymphatic vessels, but lymphatic vessels have been demonstrated in lacrimal gland and epibulbar conjunctiva. In this study, we analyzed orbital fat for the presence of lymphatic tissue using D2-40, a monoclonal antibody, specific for lymphatic vessels. We found lymphatic vessels present within bulbar conjunctiva extending to the level of the ciliary apparatus. No lymphatics were identified in healthy anterior orbital adipose tissue. In two cases of orbital mucor-mycosis and one case of panendophthalmitis, significant lymphovascular proliferation was present within granulation tissue associated with the acute inflammation. We conclude that lymph vessel proliferation may be induced in inflammatory conditions in tissues which are normally devoid of lymph channels.
Int J Mol Med 2004 May
PMID:Observation of lymphatic vessels in orbital fat of patients with inflammatory conditions: a form fruste of lymphangiogenesis? 1506 70

Paracoccidioides brasiliensis, a thermodimorphic fungus, is the causative agent of paracoccidioidomycosis (PCM), the most prevalent systemic mycosis in Latin America. Pathogenicity appears to be intimately related to the dimorphic transition from the hyphal to the yeast form, which is induced by a shift from environmental temperature to the temperature of the mammalian host. Little information is available on the P. brasiliensis genes that are necessary during the pathogenic phase. We have therefore undertaken Suppression Subtraction Hybridization (SSH) and macroarray analyses with the aim of identifying genes that are preferentially expressed in the yeast phase. Genes identified by both procedures as being more highly expressed in the yeast phase are involved in basic metabolism, signal transduction, growth and morphogenesis, and sulfur metabolism. In order to test whether the observed changes in gene expression reflect the differences between the growth conditions used to obtain the two morphological forms rather than differences intrinsic to the cell types, we performed real-time RT-PCR experiments using RNAs derived from both yeast cells and mycelia that had been cultured at 37 degrees C and 26 degrees C in either complete medium (YPD or Sabouraud) or minimal medium. Twenty genes, including AGS1 (alpha-1,3-glucan synthase) and TSA1 (thiol-specific antioxidant), were shown to be more highly expressed in the yeast cells than in the hyphae. Although their levels of expression could be different in rich and minimal media, there was a general tendency for these genes to be more highly expressed in the yeast cells.
Mol Genet Genomics 2004 Jul
PMID:Identification of genes preferentially expressed in the pathogenic yeast phase of Paracoccidioides brasiliensis, using suppression subtraction hybridization and differential macroarray analysis. 1513 90

Infections caused by fungi (mycoses) are increasingly reported in many countries owing to greater life expectancy associated with an increase in quality of medical and surgical procedures, as well as the emergence of diseases or infections that affect the immune system such as AIDS. Nosocomial outbreaks of fungal infections are sometimes reported, and typing is then necessary to find the reservoirs, analyze the modes of transmission, study the antifungal susceptibility patterns, and investigate the susceptibility of the host. In addition, the food industry is increasingly demanding typing methods that could help in selection of the best fungal strains, in order to incorporate them in the productive chains and augment the quality and security of food. This is the case for Saccharomyces cerevisiae in the wine industry: the selection and characterization of indigenous or autochthonous strains is an important objective for the production of high-quality certified wines.Several genotyping methods are now widely used for strain delineation of medically or economically important microorganisms belonging to the kingdom Fungi. Most molecular typing methods are comparable to those already described for bacteria, although the peculiarities of their nucleic acids increase the number of available methods. Although typing procedures based on the analysis of nucleic acid sequences have been developed, most genotyping methods currently in use are electrophoretically based, and the procedures include the visual comparison of nucleic acid band profiles or their reading with the help of computerized software. Here we describe some of the most frequently used genotyping methods for fungi, based on polymerase chain reactions (PCR), the isolation of chromosomal or mitochondrial DNA, and their restriction using endonuclease enzymes. The latter methods are exclusive for typing eukaryotic organisms and are based on the expected polymorphism obtained from the separation of large chromosomes using pulsed-field gel electrophoresis (PFGE) and the restriction of mitochondrial or chromosomal DNA. More sophisticated methods, such as those that combine endonuclease restriction with hybridization, are also available, although their use is less extensive and is limited mostly to research laboratories.
Methods Mol Biol 2004
PMID:Typing fungal isolates: molecular methods and computerized analysis. 1515 23


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