Multilocus microsatellite typing for<i>Cryptococcus neoformans</i>var.<i>grubii</i>

Hanafy, Ahmed M.; Kaocharoen, Sirada; Jover-Botella, Alejandro; Katsu, Masakazu; Iida, Soji; Kogure, Takahisa; Gonoi, Tohru; Mikami, Yuzuru; Meyer, Wieland

doi:10.1080/13693780802027062

Cited by 36 publications

(30 citation statements)

References 43 publications

(35 reference statements)

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“…The greatest genetic diversity among VGII/AFLP6 strains was found in Brazil (42,78) and Colombia (74), where both MATa and MAT␣ strains are present. Environmental studies conducted in South America show that VGII is particularly well adapted to biotopes associated with decaying wood (73,78,90).…”

Section: Global Distribution and Movement Of C Gattii Genotypesmentioning

confidence: 96%

“…2A), AFLP analysis (33) (Fig. 2B), MLMT analysis (42), and MLST analysis (43) are in common use. More recently, WGST has been implemented (52).…”

Section: Standardization Of Molecular Typing Systemsmentioning

confidence: 99%

“…The application of PCR fingerprinting (34,40), AFLP analysis (33), PLB1 restriction fragment length polymorphism (RFLP) analysis (41), URA5 RFLP analysis (40), and multilocus microsatellite type (MLMT) analysis (42,43) revealed intraspecies genetic diversity within C. gattii and led to the discovery of interspecies hybrids of serotypes A and B and serotypes B and D (44,45). C. neoformans-C. gattii is now considered a species complex, with the actual number of scientifically valid species being controversial (43).…”

Section: Phylogenetic Studies and Molecular Subtypingmentioning

confidence: 99%

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Cryptococcus gattii Infections

2014

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SUMMARY Understanding of the taxonomy and phylogeny of Cryptococcus gattii has been advanced by modern molecular techniques. C. gattii probably diverged from Cryptococcus neoformans between 16 million and 160 million years ago, depending on the dating methods applied, and maintains diversity by recombining in nature. South America is the likely source of the virulent C. gattii VGII molecular types that have emerged in North America. C. gattii shares major virulence determinants with C. neoformans , although genomic and transcriptomic studies revealed that despite similar genomes, the VGIIa and VGIIb subtypes employ very different transcriptional circuits and manifest differences in virulence phenotypes. Preliminary evidence suggests that C. gattii VGII causes severe lung disease and death without dissemination, whereas C. neoformans disseminates readily to the central nervous system (CNS) and causes death from meningoencephalitis. Overall, currently available data indicate that the C. gattii VGI, VGII, and VGIII molecular types more commonly affect nonimmunocompromised hosts, in contrast to VGIV. New, rapid, cheap diagnostic tests and imaging modalities are assisting early diagnosis and enabling better outcomes of cerebral cryptococcosis. Complications of CNS infection include increased intracranial pressure, severe neurological sequelae, and development of immune reconstitution syndrome, although the mortality rate is low. C. gattii VGII isolates may exhibit higher fluconazole MICs than other genotypes. Optimal therapeutic regimens are yet to be determined; in most cases, initial therapy with amphotericin B and 5-flucytosine is recommended.

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Section: Global Distribution and Movement Of C Gattii Genotypesmentioning

confidence: 96%

“…2A), AFLP analysis (33) (Fig. 2B), MLMT analysis (42), and MLST analysis (43) are in common use. More recently, WGST has been implemented (52).…”

Section: Standardization Of Molecular Typing Systemsmentioning

confidence: 99%

Section: Phylogenetic Studies and Molecular Subtypingmentioning

confidence: 99%

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Cryptococcus gattii Infections

2014

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“…Techniques employed had been MLEE (Bertout et al 1999), electrophoretic karyotype (Perfect et al 1989), Southern blot (Currie et al 1994), RAPD (Boekhout and van Belkum 1997), multilocus sequence typing (MLST) (Fraser et al 2005), MLMT (Hanafy et al 2008;Illnait-Zaragozí et al 2010), and PCR fingerprinting (Meyer et al 2003). C. neoformans/ C. gattii species complex can be divided in eight molecular types, based on DNA sequence polymorphisms that have been detected by molecular methods such as PCR fingerprinting, RFLP, or MLST: C. neoformans variety grubii [serotype A]: genotype AFLP1/VNI, AFLP1A/VNB/VNII, AFLP1B/VNII, C. neoformans intervariety hybrids [serotype AD]: genotype AFLP3/VNIII, and C. neoformans variety neoformans [serotype D]: genotype AFLP2/VNIV (Meyer et al 2003) and on the other hand, C. gattii [serotype B]: genotypes AFLP4/VGI, AFLP6/VGII, and AFLP10/VGIV and C. gattii [serotype C]: genotypes AFLP5/VGIII and AFLP7/VGIV (Meyer et al 2003;Hagen et al 2012).…”

Section: Introductionmentioning

confidence: 99%

Molecular typing of clinical isolates of Cryptococcus neoformans/Cryptococcus gattii species complex from Northeast Mexico

et al. 2015

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Cryptococcosis is caused by members of the Cryptococcus neoformans/Cryptococcus gattii species complex. Based on molecular identification, these two species have been further differentiated into molecular types. The aim of this work was to characterize clinical cryptococcal isolates recovered from six hospitals in Northeast Mexico from 1995 to 2011. One hundred and sixty-six isolates, which were characterized by biochemical tests and in vitro susceptibility to amphotericin B, fluconazole, and voriconazole, and M13 PCR fingerprinting, were included in this study. Utilizing phenotypic tests, 153 isolates (92.16 %) were identified as C. neoformans and 13 (7.83 %) as C. gattii. All isolates were susceptible to all antifungals tested. Employing M13 PCR fingerprinting, eight molecular types were detected. VNI was the most common genotype (124 cases; 74.6 %), followed by VNII (15 cases; 9 %), VNIII (8 cases; 4.8 %), VNIV (6 cases; 3.6 %), VGI (6 cases; 3.6 %), VGII (3 cases; 1.8 %), and VGIII and VGIV (2 cases, 1.2 % each). We confirm the presence of C. gattii in clinical isolates in Northeast Mexico, and a high clonal diversity in the studied strains of C. neoformans/C. gattii species complex.

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“…Contemporary methods for genotyping C. gattii are PCR-restriction fragment length polymorphism (PCR-RFLP), amplified fragment length polymorphism (AFLP), multilocus microsatellite typing (MLMT), multilocus sequence typing (MLST), and most recent, matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS) [11-14]. High resolution melting (HRM) is a method that has been used to identify the Cryptococcus neoformans-Cryptococcus gattii complex, though it has not been employed for genotyping within either species [15].…”

Section: Introductionmentioning

confidence: 99%

Real-time PCR assays for genotyping of Cryptococcus gattii in North America

et al. 2014

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BackgroundCryptococcus gattii has been the cause of an ongoing outbreak starting in 1999 on Vancouver Island, British Columbia and spreading to mainland Canada and the US Pacific Northwest. In the course of the outbreak, C. gattii has been identified outside of its previously documented climate, habitat, and host disease. Genotyping of C. gattii is essential to understand the ecological and geographical expansion of this emerging pathogen.MethodsWe developed and validated a mismatch amplification mutation assay (MAMA) real-time PCR panel for genotyping C. gattii molecular types VGI-VGIV and VGII subtypes a,b,c. Subtype assays were designed based on whole-genome sequence of 20 C. gattii strains. Publically available multilocus sequence typing (MLST) data from a study of 202 strains was used for the molecular type (VGI-VGIV) assay design. All assays were validated across DNA from 112 strains of diverse international origin and sample types, including animal, environmental and human.ResultsValidation revealed each assay on the panel is 100% sensitive, specific and concordant with MLST. The assay panel can detect down to 0.5 picograms of template DNA.ConclusionsThe (MAMA) real-time PCR panel for C. gattii accurately typed a collection of 112 diverse strains and demonstrated high sensitivity. This is a time and cost efficient method of genotyping C. gattii best suited for application in large-scale epidemiological studies.

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Multilocus microsatellite typing forCryptococcus neoformansvar.grubii

Cited by 36 publications

References 43 publications

Cryptococcus gattii Infections

Cryptococcus gattii Infections

Molecular typing of clinical isolates of Cryptococcus neoformans/Cryptococcus gattii species complex from Northeast Mexico

Real-time PCR assays for genotyping of Cryptococcus gattii in North America

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