A new duplex PCR assay for the rapid screening of mating-type idiomorphs of pathogenic Sporothrix species

Carvalho, Jamile Ambrósio de; Pinheiro, Breno Gonçalves; Hagen, Ferry; Gonçalves, Sarah Santos; Negroni, Ricardo; Kano, Rui; Bonifáz, Alexandro; Camargo, Zoilo Pires de; Rodrigues, Anderson Messias

doi:10.1016/j.funbio.2021.05.005

Cited by 20 publications

(27 citation statements)

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“…This paradoxical reproduction system has been observed in Sporothrix ( Teixeira et al. 2015 , de Carvalho et al. 2021b ), Histoplasma ( Rodrigues et al.…”

Section: Discussionmentioning

confidence: 88%

Trends in the molecular epidemiology and population genetics of emergingSporothrixspecies

Roberto¹,

Carvalho²,

Beale³

et al. 2021

Studies in Mycology

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Paracoccidioidomycosis (PCM) is a life-threatening systemic fungal infection acquired after inhalation of Paracoccidioides propagules from the environment. The main agents include members of the P. brasiliensis complex (phylogenetically-defined species S1, PS2, PS3, and PS4) and P. lutzii . DNA-sequencing of protein-coding loci (e.g., GP43 , ARF , and TUB1 ) is the reference method for recognizing Paracoccidioides species due to a lack of robust phenotypic markers. Thus, developing new molecular markers that are informative and cost-effective is key to providing quality information to explore genetic diversity within Paracoccidioides . We report using new amplified fragment length polymorphism (AFLP) markers and mating-type analysis for genotyping Paracoccidioides species. The bioinformatic analysis generated 144 in silico AFLP profiles, highlighting two discriminatory primer pairs combinations (#1 EcoRI-AC/MseI-CT and #2 EcoRI-AT/MseI-CT). The combinations #1 and #2 were used in vitro to genotype 165 Paracoccidioides isolates recovered from across a vast area of South America. Considering the overall scored AFLP markers in vitro (67–87 fragments), the values of polymorphism information content ( PIC = 0.3345–0.3456), marker index ( MI = 0.0018), effective multiplex ratio ( E = 44.6788–60.3818), resolving power ( Rp = 22.3152–34.3152), discriminating power ( D = 0.5183–0.5553), expected heterozygosity ( H = 0.4247–0.4443), and mean heterozygosity ( H avp = 0.00002–0.00004), demonstrated the utility of AFLP markers to speciate Paracoccidioides and to dissect both deep and fine-scale genetic structures. Analysis of molecular variance (AMOVA) revealed that the total genetic variance (65-66 %) was due to variability among P. brasiliensis complex and P. lutzii (PhiPT = 0.651–0.658, P < 0.0001), supporting a highly structured population. Heterothallism was the exclusive mating strategy, and the distributions of MAT1-1 or MAT1-2 idiomorphs were not significantly skewed (1:1 ratio) for P. brasiliensis s. str. (χ 2 = 1.025; P = 0.3113), P. venezuelensis (χ 2 = 0.692; P = 0.4054), and P. lutzii (χ 2 = 0.027; P = 0.8694), supporting random mating within each species. In contrast, skewed distributions were found ...

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“…This paradoxical reproduction system has been observed in Sporothrix ( Teixeira et al. 2015 , de Carvalho et al. 2021b ), Histoplasma ( Rodrigues et al.…”

Section: Discussionmentioning

confidence: 88%

Trends in the molecular epidemiology and population genetics of emergingSporothrixspecies

Roberto¹,

Carvalho²,

Beale³

et al. 2021

Studies in Mycology

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“…(1990) ITS4 TCCTCCGCTTATTGATATGC White et al. (1990) MAT1-1 SPMAT1-1F GATCCCTACAAAAGCAAATGGACCATG de Carvalho et al. (2021) SPMAT1-1R CTGCAATTGGGTTGTGCCTGATG de Carvalho et al.…”

Section: Methodsmentioning

confidence: 99%

“…(1990) MAT1-1 SPMAT1-1F GATCCCTACAAAAGCAAATGGACCATG de Carvalho et al. (2021) SPMAT1-1R CTGCAATTGGGTTGTGCCTGATG de Carvalho et al. (2021) MAT1-2 SPMAT1-2F CCAATTTCCTCTTCCACTATTCGTCGC de Carvalho et al.…”

Section: Methodsmentioning

confidence: 99%

“…(2021) SPMAT1-1R CTGCAATTGGGTTGTGCCTGATG de Carvalho et al. (2021) MAT1-2 SPMAT1-2F CCAATTTCCTCTTCCACTATTCGTCGC de Carvalho et al. (2021) SPMAT1-2R GCTTGATATCCACGGCCATCTTG de Carvalho et al.…”

Section: Methodsmentioning

confidence: 99%

“…(2021) MAT1-2 SPMAT1-2F CCAATTTCCTCTTCCACTATTCGTCGC de Carvalho et al. (2021) SPMAT1-2R GCTTGATATCCACGGCCATCTTG de Carvalho et al. (2021) ATP9-COX2 975–8038F GCTAGAAATCCTTCTTTAAGAGGAC Kawasaki et al.…”

Section: Methodsmentioning

confidence: 99%

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Exploring genetic diversity, population structure, and phylogeography in Paracoccidioides species using AFLP markers

Carvalho¹,

Beale²,

Hagen³

et al. 2021

Studies in Mycology

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Sporothrix ( Ophiostomatales ) comprises species that are pathogenic to humans and other mammals as well as environmental fungi. Developments in molecular phylogeny have changed our perceptions about the epidemiology, host-association, and virulence of Sporothrix . The classical agent of sporotrichosis, Sporothrix schenckii , now comprises several species nested in a clinical clade with S. brasiliensis , S. globosa , and S. luriei . To gain a more precise view of outbreaks dynamics, structure, and origin of genetic variation within and among populations of Sporothrix , we applied three sets of discriminatory AFLP markers (#3 EcoRI-GA/MseI-TT, #5 EcoRI-GA/MseI-AG, and #6 EcoRI-TA/MseI-AA) and mating-type analysis to a large collection of human, animal and environmental isolates spanning the major endemic areas. A total of 451 polymorphic loci were amplified in vitro from 188 samples, and revealed high polymorphism information content ( PIC = 0.1765–0.2253), marker index ( MI = 0.0001–0.0002), effective multiplex ratio ( E = 15.1720–23.5591), resolving power ( Rp = 26.1075–40.2795), discriminating power ( D = 0.9766–0.9879), expected heterozygosity ( H = 0.1957–0.2588), and mean heterozygosity ( H avp = 0.000007–0.000009), demonstrating the effectiveness of AFLP markers to speciate Sporothrix . Analysis using the program structure indicated three genetic clusters matching S. brasiliensis (population 1), S. schenckii (population 2), and S. globosa (population 3), with the presence of patterns of admixture amongst all populations. AMOVA revealed highly structured clusters (PhiPT = 0.458–0.484, P < 0.0001), with roughly equivalent genetic variability within (46–48 %) and between (52–54 %) populations. Heterothallism was the exclusive mating strategy, and the distributions of MAT1-1 or MAT1-2 idiomorphs were not significantly skewed (1:1 ratio) for S. schenckii (χ 2 = 2.522; P = 0.1122), supporting random mating. In contrast, skewed distributions were found for S. globosa (χ 2 = 9.529; P = 0.0020) with a predominance of MAT1-1 isolates, and regional differences were highlighted for S. brasiliensis with the overwhelming occurrence of MAT1-2 in Rio de Janeiro (χ 2 = 14.222; P = 0.0002) and Pernambuco (χ 2 ...

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A Spotlight on Sporothrix and Sporotrichosis

2022

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A new duplex PCR assay for the rapid screening of mating-type idiomorphs of pathogenic Sporothrix species

Cited by 20 publications

References 53 publications

Trends in the molecular epidemiology and population genetics of emergingSporothrixspecies

Trends in the molecular epidemiology and population genetics of emergingSporothrixspecies

Exploring genetic diversity, population structure, and phylogeography in Paracoccidioides species using AFLP markers

A Spotlight on Sporothrix and Sporotrichosis

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