Population structure and molecular genetic characterization of clinical Candida tropicalis isolates from a tertiary-care hospital in Kuwait reveal infections with unique strains

Abstract

Candida tropicalis is a frequently isolated yeast species causing bloodstream, urinary tract and other infections particularly in patients admitted to intensive care units (ICUs) and those requiring prolonged urinary catheterization (UC) or receiving broad-spectrum antibiotics (BSA). This study investigated clinical characteristics and genetic relatedness among C. tropicalis strains isolated from patients at Al-Amiri Hospital in Kuwait. C. tropicalis strains (n = 63) isolated from blood, genito-urinary, respiratory (RT) and digestive (GIT) tracts and wound sites from 54 patients were used. All isolates were phenotypically identified and tested against six antifungal drugs by using Vitek 2 system. Molecular identification was performed by PCR amplification of rDNA. Fingerprinting was achieved by 6-loci-based multilocus sequence typing (MLST) and data were analyzed by BioNumerics software for phylogenetic relationships. Patients mean age was >65 years and >20% patients were hospitalized in ICUs. Most patients had underlying conditions that included UC, BSA, diabetes and RT/GIT abnormalities. Most candiduria cases had UC, ureteric stent or suprapubic catheters. All isolates were identified as C. tropicalis by Vitek 2 and by species-specific PCR. Sixty-two isolates were susceptible to all tested antifungal drugs. MLST identified 59 diploid sequence types (DSTs) including 54 newly-identified DSTs. C. tropicalis isolates from multiple sites of same patient usually belonged to different DSTs. Interestingly, 56 of 57 isolates from 48 patients belonged to unique genotypes. Only six isolates from six patients belonged to three DSTs (clusters), however, C. tropicalis strains in each cluster were isolated >3 months apart. Our data show diverse origins of C. tropicalis infections in Kuwait as most isolates were unique strains. There was no obvious correlation between cluster isolates with time of isolation and/or hospital ward of their origin. This study presents the first MLST analysis of C. tropicalis isolates from Middle East and may be useful for studying genetic relationships among global C. tropicalis strains.

Multilocus Sequence Typing Reveals a New Cluster of Closely Related Candida tropicalis Genotypes in Italian Patients With Neurological Disorders

“…In fact all new Italian genotypes (DSTs 747-51 and 759) were not present in the C. tropicalis MLST database before and the unique deposited blood isolate with DST333 (the second most common genotype of this study) was previously reported from a Chinese patient hospitalized in a neurosurgery unit ( Fan et al, 2014 ). However, a recent global analysis of the MLST genotypes confirmed a lack of association between different genotypes and antifungal resistance, specimen type, or geographical locations ( Al-Obaid et al, 2017 ; Wu et al, 2017 ) even if the existence of an exclusive MLST clonal cluster from Italy suggests the occurrence of independent geographical evolution. Nevertheless, although the number of the isolates included in this study make up ∼20% (28/143) of the European C. tropicalis deposited in the MLST database, the amount of the examined Italian isolates was still too small to draw any solid conclusion and therefore further studies will be needed in order to corroborate our observations.…”

“…Candida tropicalis is a diploid yeast that together with C. albicans, C. parapsilosis , and other less relevant Candida pathogens forms part of the so-called Candida CTG clade in which the CUG codon is translated as serine rather than leucine ( Papon et al, 2013 ). Genetically, the population structure of this fungus appears to be quite heterogeneous, and more complex ( Wu et al, 2012 ; Al-Obaid et al, 2017 ; Wu et al, 2017 ) than the one previously described by Tavanti et al (2005) using multilocus sequence typing (MLST) technique. However, compared to the C. albicans MLST database 2 , which contains genetic data from over 4000 isolates, the number of C. tropicalis isolates genotyped so far is still too small to provide a good estimate of the pattern of the genetic variation occurring in this species.…”

Development of Echinocandin Resistance in Candida tropicalis following Short-Term Exposure to Caspofungin for Empiric Therapy

“…Both isolates (Kw3707/16 and Kw107/17) were initially identified by growth on CHROMagar Candida and Vitek 2 yeast identification systems. Identification was confirmed by Vitek MS, species-specific amplification, and PCR sequencing of the internally transcribed spacer (ITS) region of ribosomal DNA (rDNA), performed as described previously (15). Antifungal drug susceptibility testing (AST) was performed, and MICs were determined by Etest as described previously (16).…”

“…The hot spot 1 (HS-1) and HS-2 regions of the FKS1 gene were amplified by using FKS1-1F (5=-GGTTTAGATATGTGGATGTCCTA-3=) plus FKS1-1R (5=-AAATGTTTCTCCATGG AGTCAAG-3=) and FKS1-2F (5=-CTACCAAGATTGGTGCTG-3=) plus FKS1-2R (5=-TATCTAG CACCACCAACAG-3=) primers, respectively, and PCR thermal and cycling conditions as described previously (15,25). Both strands of HS-1 and HS-2 amplicons were sequenced by using internal FKS1-1FS (5=-TTAGATATGTGGATGTCCTACTT-3=) or FKS1-1RS (5=-GTTTCTCCA TGGAGTCAAGATG-3=) and FKS1-2FS (5=-ACCAAGATTGGTGCTGGTA-3=) or FKS1-2R (5=-CTA GCACCACCAACAGTCA-3=) primers, respectively, and other sequencing reaction conditions as described previously (15,25). DNA sequence data were compared to the reference sequence from C. tropicalis (GenBank accession no.…”

“…EU676168) by Clustal Omega (https:// www.ebi.ac.uk/Tools/msa/clustalo/). Genetic relatedness between the two isolates and with five other echinocandin-susceptible isolates collected from the same hospital, including three isolates from the same intensive care unit, was determined by multilocus sequence typing based on six loci (ICL1, MDR1, SAPT2, SAPT4, XYR1, and ZWF1a) (15). The allele number for each gene and allele combinations (diploid sequence type [DST]) for the six loci for each isolate were assigned, or new allele and DST numbers were obtained from the C. tropicalis MLST database (https://pubmlst.org/ctropicalis/).…”