<i>MP1</i>
            Homologue-Based Multilocus Sequence System for Typing the Pathogenic Fungus
            <i>Penicillium marneffei</i>
            : a Novel Approach Using Lineage-Specific Genes

Woo, Patrick C. Y.; Lau, Candy C. Y.; Chong, Ken T. K.; Tse, Herman; Tsang, Dominic N.C.; Lee, Rodney A.; Tse, Cindy Wing-Sze; Que, Tak-Lun; Chung, Liliane M. W.; Ngan, Antonio H. Y.; Hui, Wai-Ting; Wong, Samson Sai-Yin; Lau, Susanna K. P.; Yuen, Kwok‐Yung

doi:10.1128/jcm.00619-07

Cited by 28 publications

(37 citation statements)

References 46 publications

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“…In addition to a high degree of discrim- on April 2, 2019 by guest http://jcm.asm.org/ ination, reproducibility, and typeability, microsatellite analysis has several other advantages over other DNA-based typing assays. Because the assay is PCR based, microsatellite analysis requires relatively small amounts (ϳ1 ng per reaction) of template DNA, and it can be simplified by using multiplex assays with primers labeled with different fluorescent dyes, resulting in higher throughput (6,23). Finally, microsatellite analysis allows the detection of mixed cultures, which would appear as a PCR profile harboring two or more microsatellite alleles from a single DNA sample of the haploid genome of A. flavus.…”

Section: Discussionmentioning

confidence: 99%

Microsatellite Typing To Trace Aspergillus flavus Infections in a Hematology Unit

et al. 2010

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Assessing the relatedness of strains isolated from patients and their environment is instrumental in documenting the source of preventable health care-associated life-threatening Aspergillus flavus human infection clusters. The present study aimed at identifying and selecting suitable microsatellite markers for A. flavus typing. This typing scheme was then applied to investigate the A. flavus epidemiology within a hematology unit in Sfax, Tunisia. Use of a combination of five markers made it possible to discern clusters of isolates and to substantiate the genetic diversity of A. flavus within clusters. Isolates from Tunisia and Marseille, France, displayed distinct haplotypes, indicating a highly significant geographical structuring of A. flavus. The typing of clinical and environmental A. flavus isolates in a hematology unit provided insights into its hospital epidemiology. From a heterogeneous genetic background, a cluster indicative of a clonal propagation episode within the unit could be identified. In two patients with invasive aspergillosis, the same genotype was found in clinical and environmental isolates, indicating hospital-acquired colonization and infection. In further studies, this novel microsatellite typing scheme might be instrumental in illuminating important epidemiological issues about A. flavus population genetics or epidemiology, including tracing the sources and routes of transmission.

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Section: Discussionmentioning

confidence: 99%

Microsatellite Typing To Trace Aspergillus flavus Infections in a Hematology Unit

et al. 2010

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“…Fungal DNA extraction was performed as described in our previous publications (24,26). Briefly, DNA was extracted from 1 g of fungal cells in 10 ml of distilled water using a DNeasy plant minikit according to the manufacturer's instructions (Qiagen, Hilden, Germany).…”

Section: Methodsmentioning

confidence: 99%

Internal Transcribed Spacer Region Sequence Heterogeneity in Rhizopus microsporus : Implications for Molecular Diagnosis in Clinical Microbiology Laboratories

Woo¹,

Leung²,

To³

et al. 2010

J Clin Microbiol

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Although internal transcribed spacer region (ITS) sequence heterogeneity has been reported in a few fungal species, it has very rarely been reported in pathogenic fungi and has never been described in Mucorales, causes of the highly fatal mucormycosis. In a recent outbreak investigation of intestinal mucormycosis due to Rhizopus microsporus infection in patients with hematological malignancies, PCR of the ITS of four of the 28 R. microsporus strains, P11, P12, D3-1, and D4-1, showed thick bands at about 700 bp. Direct sequencing of the purified bands showed frequent double peaks along all of the sequence traces and occasional triple peaks for P12, D3-1, and D4-1. The thick bands of the four R. microsporus strains were purified and cloned. Sequencing of 10 clones for each strain revealed two different ITS sequences for P11 and three different ITS sequences for P12, D3-1, and D4-1. Variations in ITS sequence among the different ribosomal DNA (rDNA) operons in the same strain were observed in only ITS1 and ITS2 and not the 5.8S rDNA region. One copy of P11, P12, and D4-1, respectively, and one copy of P11, P12, D3-1, and D4-1, respectively, showed identical sequences. This represents the first evidence of ITS sequence heterogeneity in Mucorales. ITS sequence heterogeneity is an obstacle to molecular identification and genotyping of fungi in clinical microbiology laboratories. When thick bands and double peaks are observed during PCR sequencing of a gene target, such a strain should be sent to reference laboratories proficient in molecular technologies for further identification and/or genotyping.Genes and intergenic regions of ribosomal DNA (rDNA) operons are the most widely used targets for molecular identification of bacteria and fungi in clinical microbiology laboratories. For bacterial identification, the 16S rDNA gene is the primary target to amplify and sequence (28), whereas for fungi, the 18S rDNA gene and internal transcribed spacer region (ITS) comprising the ITS1-5.8S-ITS2 rDNA gene cluster are commonly used, depending on the group of fungi being identified (4,10,23,27). Irrespective of the target, such a molecular identification technique usually involves PCR amplification of the target and purification and direct sequencing of the PCR product. Since most bacterial and fungal genomes contain more than one rDNA operon, the success of using this technology relies on sequence homogeneity in the various copies of targets in the rDNA operons within the genome of the bacterium or fungus.Interoperon heterogeneities for 16S rDNA genes have been reported in a number of bacteria (3,12). Recently, we reported rDNA operon heterogeneity in a novel genus and species of bacterium, Anaerospora hongkongensis, isolated from an intravenous drug user (25). When present, such rDNA operon heterogeneity will pose difficulties for direct sequencing of the PCR product for bacterial identification as double or multiple nucleotide peaks will be present in the sequence traces. Although ITS sequence heterogeneity has been reported ...

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“…All meiotic genes (except HOP1) and genes encoding putative pheromone processing enzymes, pheromone receptors, and pheromone response pathway proteins in Aspergillus fumigatus and Aspergillus nidulans and a putative MAT-1␣ box mating-type gene were present, suggesting that P. marneffei can potentially be a heterothallic fungus that does not switch mating type (8). Genes encoding Mp1p homologues were identified and used for construction of a highly discriminative multilocus sequence typing scheme for P. marneffei (9). Twenty-three putative polyketide synthase (PKS) genes and two putative PKS-nonribosomal peptide synthase hybrid genes were identified, a diversity much higher than those of other pathogenic thermal dimorphic fungi, such as Histoplasma capsulatum (one PKS gene) and Coccidioides immitis (10 PKS genes) (11).…”

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confidence: 99%

Draft Genome Sequence of Penicillium marneffei Strain PM1

et al. 2011

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Penicillium marneffei is the most important thermal dimorphic, pathogenic fungus endemic in China and Southeast Asia and is particularly important in HIV-positive patients. We report the 28,887,485-bp draft genome sequence of P. marneffei , which contains its complete mitochondrial genome, sexual cycle genes, a high diversity of Mp1p homologues, and polyketide synthase genes.

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MP1 Homologue-Based Multilocus Sequence System for Typing the Pathogenic Fungus Penicillium marneffei : a Novel Approach Using Lineage-Specific Genes

Cited by 28 publications

References 46 publications

Microsatellite Typing To Trace Aspergillus flavus Infections in a Hematology Unit

Microsatellite Typing To Trace Aspergillus flavus Infections in a Hematology Unit

Internal Transcribed Spacer Region Sequence Heterogeneity in Rhizopus microsporus : Implications for Molecular Diagnosis in Clinical Microbiology Laboratories

Draft Genome Sequence of Penicillium marneffei Strain PM1

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