Development of a multiplex PCR short tandem repeat typing scheme for Candida krusei

2021 ◽  
Author(s):  
Merlijn H.I. van Haren ◽  
Theun de Groot ◽  
Bram Spruijtenburg ◽  
Kusum Jain ◽  
Anuradha Chowdhary ◽  
...  
Keyword(s):  
Multiplex Pcr ◽  
Tandem Repeat ◽  
Short Tandem Repeat ◽  
Cost Effective ◽  
Candida Krusei ◽  
Str Typing ◽  
Str Markers ◽  
Typing Scheme ◽  
Snp Calling ◽  
Short Tandem

Candida krusei is a human pathogenic yeast that can cause candidemia with the lowest 90-day survival rate in comparison to other Candida species. Infections occur frequently in immunocompromised patients and several C. krusei outbreaks in health care facilities have been described. Here, we developed a short tandem repeat (STR) typing scheme for C. krusei to allow for fast and cost-effective genotyping of an outbreak and compared identified relatedness of ten isolates to SNP calling from whole-genome sequencing (WGS). From a selection of 14 novel STR markers, six were used to develop two multiplex PCRs. Additionally, three previously reported markers were selected for a third multiplex PCR. In total, 119 C. krusei isolates were typed using these nine markers and 79 different genotypes were found. STR typing correlated well with WGS SNP typing, as isolates with the same STR genotype varied by 8 and 19 SNPs, while isolates that differed in all STR markers varied at least tens of thousands of SNPs. The STR typing assay was found to be specific for C. krusei , stable in 100 subcloned generations, and comparable to SNP calling by WGS. In summary, this newly developed C. krusei STR typing scheme is a fast, reliable, easy-to-interpret and cost-effective method compared to other typing methods. Moreover, the two newly developed multiplexes showed the same discriminatory power as all nine markers combined, indicating that multiplexes M3-1 and M9 are sufficient to type C. krusei .

scholarly journals Resolution of parentage in dogs by examination of microsatellites after death of putative sire: Case report

2001 ◽  
Vol 49 (3) ◽  
pp. 269-273 ◽  
Author(s):  
Z. Pádár ◽  
B. Egyed ◽  
K. Kontadakis ◽  
L. Zöldág ◽  
S. Fekete
Keyword(s):  
Case Report ◽  
Multiplex Pcr ◽  
Tandem Repeat ◽  
Short Tandem Repeat ◽  
Genetic Homogeneity ◽  
Str Loci ◽  
Hair Samples ◽  
Short Tandem

A case of disputed paternity in dogs is reported. DNA examinations were carried out from hair samples of the individuals several months after the death of the putative sire. Ten short tandem repeat (STR) loci were analysed by fluorescence- labelled multiplex PCR using ABI PRISM 310 Genetic Analyser. Based on the results the candidate sire was included in the pedigree records as the biological sire. In spite o f the genetic homogeneity of pedigree dogs due to inbreeding, canine microsatellites can provide an adequate basis for assigning paternity in pure breeds.

Chimerism Monitoring by Short Tandem Repeat (STR) Markers in Allogeneic Stem Cell Transplantation

2018 ◽  
Vol 64 (09/2018) ◽  
Author(s):  
Raluca Dumache ◽  
Alexandra Enache ◽  
Ligia Barbarii ◽  
Carmen Constantinescu ◽  
Andreea Pascalau ◽  
...  
Keyword(s):  
Stem Cell ◽  
Stem Cell Transplantation ◽  
Cell Transplantation ◽  
Allogeneic Stem Cell Transplantation ◽  
Tandem Repeat ◽  
Short Tandem Repeat ◽  
Str Markers ◽  
Short Tandem

Mitochondrial DNA Minisatellites Showed Higher Informativeness and Sensitivity Than the Nuclear DNA Markers for the Quantitative Determination of Chimerism After Allogeneic Stem Cell Transplantation,

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 4004-4004
Author(s):  
Hye Ran Kim ◽  
Eun-Jeong Won ◽  
Hyun-Jung Choi ◽  
Hwan-Young Kim ◽  
James Moon ◽  
...  
Keyword(s):  
Mitochondrial Dna ◽  
Stem Cell ◽  
Stem Cell Transplantation ◽  
Quantitative Determination ◽  
Allogeneic Stem Cell Transplantation ◽  
Tandem Repeat ◽  
Short Tandem Repeat ◽  
Str Markers ◽  
Short Tandem

Abstract Abstract 4004 Background: Mitochondrial DNA (mtDNA) is widely used in forensic identification and anthropologic studies on account of its abundance resulting in preferential amplification, sequencing and inherent variability. We developed mtDNA markers to monitor donor cell engraftment after allogeneic stem cell transplantation(SCT), then compared with nuclear short tandem repeat (STR) markers. Patients and methods: The mtDNA control regions and six mtDNA minisatellites (mtMS) (303 poly C, 16184 poly C, 514 (CA) repeat, 3566 poly C, 12385 poly C and 12418 poly A) from the total DNA samples of 215 cases (donor, recipient and after allogeneic SCT) were amplified using the designated specific primers and PCR. The results were compared with those from the six short tandem repeat (STR) markers (D12S391, D18S51, F13A1, HUM RENA-4, HUM FABP2 and Amelogenin). Results: Polymorphisms in the mtDNA control region identify an informative marker in 88% (189 cases) of all cases. Among the six mtMS markers, the informativeness of 303 poly C and 16184 poly C mtMS was 63% and 67% respectively. A combination of direct sequencing through the mtDNA control region, 303poly C and 16184 poly C mtMS could completely distinguish the donor cells from the recipient cells. The results from a typical mixing experiment to determine the sensitivity revealed a detection limit (DL) of the gene scan analysis in a mtDNA mixture to be visible at 1% heteroplasmy in 303 poly C mtMS marker. However, the DL from D12S391 in the same mixing experiment was 5–10% heteroplasmy. Conclusions: mtMS markers, especially 303 poly C and 16184 poly C markers, can provide a sensitive, accurate and quantitative determination of mixed chimerism after a SCT. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Development of Candida auris Short Tandem Repeat Typing and Its Application to a Global Collection of Isolates

mBio ◽  
2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Theun de Groot ◽  
Ynze Puts ◽  
Indira Berrio ◽  
Anuradha Chowdhary ◽  
Jacques F. Meis
Keyword(s):  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
South Asian ◽  
Tandem Repeat ◽  
Cost Effective ◽  
Whole Genome ◽  
Candida Auris ◽  
Str Typing ◽  
Content Type ◽  
Short Tandem

ABSTRACT Candida auris is a pathogenic yeast that causes invasive infections with high mortality. Infections most often occur in intensive care units of health care facilities. It is crucial to trace the source and prevent further spread of C. auris during an outbreak setting; therefore, genotyping of C. auris is required. To enable fast and cost-effective genotyping, we developed a short tandem repeat (STR) typing assay for C. auris. STRs in C. auris were identified, and from an initial selection of 23 STRs, 12 were used to develop a STR typing assay. Having shown that the STR typing assay was reproducible and specific, a robust set of 444 C. auris isolates was investigated to identify genotypic diversity. In concordance with whole-genome sequencing (WGS) analysis, we identified five major different C. auris clusters of South American, South Asian, African, East Asian, and Iranian origin. Overall, a total of 40 distinct genotypes were identified, with the largest variety in the South Asian clade. Comparison with WGS demonstrated that isolates with <20 single nucleotide polymorphisms (SNPs) are mostly not differentiated by STR analysis, while isolates with 30 or more SNPs usually have differences in one or more STR markers. Altogether, a highly reproducible and specific STR typing assay for C. auris was developed; this assay distinguishes the five different C. auris clades in identical fashion to WGS, while most isolates differing by >30 SNPs, as determined via WGS, are also separated. This new C. auris-specific genotyping technique is a rapid, reliable, and cost-effective alternative to WGS analysis to investigate outbreaks. IMPORTANCE Candida auris is an emerging fungal pathogen now recognized as a threat to public health. The pathogen has spread worldwide and causes mainly hospital-associated outbreaks. To track and trace outbreaks and to relate them to new introductions from elsewhere, whole-genome sequencing and amplified fragment length polymorphism (AFLP) have been used for molecular typing. Whole-genome sequencing is costly and available only at a few centers, and AFLP is a complicated technique and hard to interpret. We describe a novel simple STR genotyping technique based on short tandem repeats in the C. auris genome. We also show that the performance of this STR-based genotyping technique has proven comparable to that of WGS. Overall, this work provides a novel, rapid, reliable, and cost-effective method of molecular outbreak investigations of C. auris.

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