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Candida auris in Germany and Previous Exposure to Foreign Healthcare

The emerging yeast Candida auris has disseminated worldwide. We report on 7 cases identified in Germany during 2015–2017. In 6 of these cases, C. auris was isolated from patients previously hospitalized abroad. Whole-genome sequencing and epidemiologic analyses revealed that all patients in Germany were infected with different strains.

Since the worldwide emergence of Candida auris, the National Reference Centre for Fungal Infections (NRZMyk) has issued several alerts on this species and informed diagnostic laboratories in Germany to send all suspicious Candida spp. isolates to the reference laboratory for identification. In total, during November 2015-December 2017, we collected 8 isolates from 6 patients; for 1 case (reliable identification with Bruker MALDI), no isolate was available.

Identification and Susceptibility Testing
We identified all isolates by biochemical methods (Vitek2 Yeast Card, API ID32C V4.0 (bioMérieux) and by MALDI-TOF using 2 different systems (VitekMS, bioMérieux or Biotyper Microflex, Bruker Daltonics) and 2 protein extraction protocols, as previously described (1). In addition, we identified all isolates by PCR and sequencing of the internal transcribed spacer (ITS) (2).

DNA Isolation
We isolated fungal DNA with a phenol-free procedure, as described previously (3). We analyzed the quality of the isolated DNA by Nanodrop and with an Invitrogen Qubit Fluorometer (https://www.thermofisher.com).

Whole-Genome Sequencing
Library preparation and whole-genome sequencing of C. auris isolates was performed by either GATC (Konstanz, Germany) or LGC Genomics (Berlin, Germany) on the Illumina platform using 2x 150-bp paired end reads. Illumina adapters were removed from the GATCsequenced isolates using Trim Galore version 0.4.3.1 (Babraham Institute, https://www.babraham.ac.uk). Further data processing was performed using the CSI Phylogeny Pipeline (4). Briefly, reads were mapped to the B8441 v2 reference genome using BWA version 0.7.12 (5). SNP calling was performed using SAMtools version 0.1.18 (6). Following variant detection, SNPs were filtered for a minimum depth of 10, minimum relative depth of 10, minimum SNP quality of 30, and a Z-score of greater than 1.96. No SNP pruning (removal of closely spaced SNPs) was performed to better separate closely related isolates. SNPs were then concatenated and maximum-likelihood phylogeny inferred using FastTree version 2.1.7 (7). For comparison of the isolates in this study to reference strains from other C. auris clades, raw sequence data from the Indian isolate 6684 (BioSample ID SAMN03200169), the Japanese isolate B11220 (BioSample ID SAMN05379608), the South African isolate B11221 (BioSample ID SAMN05379609), and the Venezuelan isolate B11243 (BioSample ID SAMN05379619) were downloaded from the National Center for Biotechnology Information (NCBI) and processed using the same pipeline.

Data Availability
Raw sequence read files were uploaded to the NCBI Sequence Read Archive and are publicly available under BioProject IDs PRJNA485145, PRJNA485239, PRJNA485259, PRJNA485409, PRJNA485414, and PRJNA485415.

Clinical Case Presentations
Patient 1 was admitted for a prosthetic joint infection, which was first diagnosed and treated with antimicrobial drugs in Oman. After hospital admission in Germany and replacement of the prosthesis, a febrile episode occurred and C. auris was isolated from blood cultures. The patient recovered after treatment with an echinocandin and C. auris has not been isolated in subsequent hospital stays.

Patient 3 had an intracranial hemorrhage that had previously been treated in a Saudi
Arabian healthcare facility. Because of a known colonization with carbapenemase-producing Enterobacteriaceae, the patient was placed in a single room. Culture of the catheter urine revealed growth of 10 5 CFU/mL of C. auris. Even though urinary catheters were changed several times, C. auris was isolated from 2 follow-up urine specimens. Furthermore, C. auris was grown from the groin and the tracheostomy. In the absence of any signs of infection in the later course, the patient was transferred to rehabilitation without further treatment of C. auris. Patient 4 was transferred to Germany from Dubai because of a neurologic disorder. C. auris was isolated from catheter urine. No treatment was initiated, as the patient was asymptomatic and was discharged alive.
Patient 5 had a multiple trauma, which was initially treated in Russia. From an intraabdominal sample taken during surgery, Enterococcus faecalis, E. avium, and C. auris were isolated. Treatment with ampicillin-sulbactam and amphotericin B was initiated. The patient was transferred to another hospital in good condition 10 days after surgery.

Patient 6 was treated in Germany for sequelae of an injury originating in Afghanistan.
Because of systemic signs of infection, blood cultures were drawn, revealing the presence of C.
auris. Infection control measures were implemented for this patient after detection of C. auris.
Because the isolate had elevated MICs for echinocandins, treatment with liposomal amphotericin B and voriconazole was initiated. C. auris was also isolated from the urine, likely as a result of an infection of the renal pelvis and the presence of several large kidney stones. Candiduria has not improved for >6 months despite prolonged therapy with voriconazole and amphotericin B.
However, since >6 months, no C. auris is detectable from this patient's specimen.