Spectrum and management of rare Candida/yeast infections in Kuwait in the Middle East.

Invasive fungal infections (IFIs) are associated with high mortality rates and mostly affect patients with compromised immunity. The incidence of IFIs is increasing worldwide with the expanding population of susceptible patients. Candida and other yeast infections represent a major component of IFIs. Rare Candida/yeast infections have also increased in recent years and pose considerable diagnostic and management challenges as they are not easily recognized by routine phenotypic characteristic-based diagnostic methods and/or by the automated yeast identification systems. Rare Candida/yeasts also exhibit reduced susceptibility to antifungal drugs making proper management of invasive infections challenging. Here, we review the diagnosis and management of 60 cases of rare Candida/yeast IFIs described so far in Kuwait, an Arabian Gulf country in the Middle East. Interestingly, majority (34 of 60, 56.7%) of these rare Candida/yeast invasive infections occurred among neonates or premature, very-low-birth-weight neonates, usually following prior bacteremia episodes. The clinical details, treatment given, and outcome were available for 28 of 34 neonates. The crude mortality rate among these neonates was 32.2% as 19 of 28 (67.8%) survived the infection and were discharged in healthy condition, likely due to accurate diagnosis and frequent use of combination therapy. Physicians treating patients with extended stay under intensive care, on mechanical ventilation, receiving broad spectrum antibiotics and with gastrointestinal surgery/complications should proactively investigate IFIs. Timely diagnosis and early antifungal treatment are essential to decrease mortality. Understanding the epidemiology and spectrum of rare Candida/yeast invasive infections in different geographical regions, their susceptibility profiles and management will help to devise novel diagnostic and treatment approaches and formulate guidelines for improved patient outcome.

Molecular fingerprinting by multi-locus sequence typing identifies microevolution and nosocomial transmission of Candida glabrata in Kuwait.

Backgrounds: Candida glabrata is a frequently isolated non-albicans Candida species and invasive C. glabrata infections in older patients are associated with high mortality rates. Opportunistic Candida infections in critically ill patients may be either endogenous or nosocomial in origin and this distinction is critical for effective intervention strategies. This study performed multi-locus sequence typing (MLST) to study genotypic relatedness among clinical C. glabrata isolates in Kuwait. Methods: Candida glabrata isolates (n = 91) cultured from 91 patients were analyzed by MLST. Repeat isolates (n = 16) from 9 patients were also used. Antifungal susceptibility testing for fluconazole, voriconazole, caspofungin and amphotericin B (AMB) was determined by Etest. Genetic relatedness was determined by constructing phylogenetic tree and minimum spanning tree by using BioNumerics software. Results: Resistance to fluconazole, voriconazole and AMB was detected in 7, 2 and 10 C. glabrata isolates, respectively. MLST identified 28 sequence types (STs), including 12 new STs. ST46 (n = 33), ST3 (n = 8), ST7 (n = 6) and ST55 (n = 6) were prevalent in ≥4 hospitals. Repeat isolates obtained from same or different site yielded identical ST. No association of ST46 with source of isolation or resistance to antifungals was apparent. Microevolution and cross-transmission of infection was indicated in two hospitals that yielded majority (57 of 91, 67%) of C. glabrata. Conclusion: Our data suggest that C. glabrata undergoes microevolution in hospital environment and can be nosocomially transmitted to other susceptible patients. Thus, proper infection control practices during routine procedures on C. glabrata-infected patients may prevent transmission of this pathogen to other hospitalized patients.

Epidemiology of Candidemia in Kuwait: A Nationwide, Population-Based Study.

The Candida species cause a majority of invasive fungal infections. In this article, we describe the nationwide epidemiology of candidemia in Kuwait in 2018. Yeast bloodstream isolates submitted from all major hospitals and identified by phenotypic MALDI-TOF MS and/or by molecular methods were studied. Susceptibility testing was performed by Etest. Out of 313 bloodstream yeasts, 239 Candida spp. isolates (excluding duplicate isolates) were obtained during 234 candidemic episodes among 223 patients. Mixed-species candidemia and re-infection occurred in 5 and 11 patients, respectively. C. albicans (n = 74), C. parapsilosis (n = 54), C. tropicalis (n = 35), C. auris (n = 33), C. glabrata (n = 32), other Candida spp. (n = 11), and other yeasts (n = 9) caused fungemia. Nearly 50% of patients were in intensive care units. Candida spp. isolates (except C. glabrata) were susceptible to caspofungin and 27% of C. auris were amphotericin B-resistant. Resistance to fluconazole was 100% in C. auris, 17% in C. parapsilosis, 12% in C. glabrata, and 1% in C. albicans. Mortality was 47% for other Candida/yeast infections. Nationwide candidemia incidence in 2018 was 5.29 cases/100,000 inhabitants. Changes in species spectrum, increasing fluconazole resistance in C. parapsilosis, and the emergence of C. auris as a major pathogen in Kuwait are noteworthy findings. The data could be of help in informing decisions regarding planning, in the allocation of resources, and in antimicrobial stewardship.

Human Fungal Infections in Kuwait-Burden and Diagnostic Gaps.

Fungal infections are an increasingly important public health issue, yet accurate statistics on fungal burden worldwide and in Kuwait are scarce. Here we estimate the incidence and prevalence of fungal infections in Kuwait. Population statistics from 2018 collected by the Public Authority for Civil Information were used, as well as data from the Ministry of Health. A literature search for Kuwait data on mycotic diseases and population at risk (chronic obstructive pulmonary disease, HIV infection/AIDS, cancer, and transplant patients) was conducted. The population in 2018 was estimated at 4,226,920 million people: 1,303,246 million Kuwaitis and 2,923,674 million expatriates. We determined the annual burden of serious fungal infections number (per 100,000) from high to low based on earlier reported fungal rates for populations at risk: recurrent Candida vaginitis 54,842 (2595); severe asthma with fungal sensitisation 10,411 (246); allergic bronchopulmonary aspergillosis, 7887 (187); chronic pulmonary aspergillosis 995 (21.3); invasive aspergillosis 704 (16.7); fungal keratitis 654 (15.5); candidaemia 288 (6.8); Candida peritonitis 63 (3.5) and oesophageal candidiasis in HIV 33 (0.8). Besides identifying rising new risk groups and expanding reports on antifungal resistance, surveillance programs and further epidemiological studies are needed to achieve more precise assessments of fungal disease epidemiology and correlated morbidity and mortality.

Molecular Epidemiology of Candida Auris Outbreak in a Major Secondary-Care Hospital in Kuwait.

The emerging, often multidrug-resistant Candida auris is increasingly being associated with outbreaks in healthcare facilities. Here we describe the molecular epidemiology of a C. auris outbreak during 18 months, which started in 2018 in the high dependency unit (HDU) of a secondary-care hospital in Kuwait. Demographic and clinical data for candidemia and colonized patients were prospectively recorded. Clinical and environmental isolates were subjected to phenotypic and molecular identification; antifungal susceptibility testing by broth microdilution method; PCR-sequencing of ERG11 and FKS1 for resistance mechanisms to triazoles and echinocandins, respectively; and molecular fingerprinting by short tandem repeat (STR) analyses. Seventy-one (17 candidemic and 54 colonized) patients including 26 with candiduria and seven environmental samples yielded C. auris. All isolates were identified as C. auris by Vitek2, MALDI-TOF MS, PCR amplification and/or PCR-sequencing of rDNA. Twelve candidemia and 26 colonized patients were admitted or exposed to HDU. Following outbreak recognition, an intensive screening program was instituted for new patients. Despite treatment of all candidemia and 36 colonized patients, 9 of 17 candidemia and 27 of 54 colonized patients died with an overall crude mortality rate of ~50%. Nearly all isolates were resistant to fluconazole and contained the Y132F mutation in ERG11 except one patient's isolates, which were also distinct by STR typing. Only urine isolates from two patients developed echinocandin resistance with concomitant FKS1 mutations. The transmission of C. auris in this outbreak was linked to infected/colonized patients and the hospital environment. However, despite continuous surveillance and enforcement of infection control measures, sporadic new cases continued to occur, challenging the containment efforts.

Candida kefyr in Kuwait: Prevalence, antifungal drug susceptibility and genotypic heterogeneity.

OBJECTIVE: Candida kefyr causes invasive candidiasis in immunocompromised patients, particularly among those with oncohematological diseases. This study determined the prevalence of C. kefyr among yeast isolates collected during 2011-2018 in Kuwait. Antifungal susceptibility testing (AST) and genotypic heterogeneity among C. kefyr was also studied. METHODS: Clinical C. kefyr isolates recovered from bloodstream and other specimens during 2011 to 2018 were retrospectively analyzed. All C. kefyr isolates were identified by CHROMagar Candida, Vitek2 and PCR amplification of rDNA. AST was performed by Etest. Molecular basis of resistance to fluconazole and echinocandins was studied by PCR-sequencing of ERG11 and FKS1, respectively. Genotypic heterogeneity was determined with microsatellite-/minisatellite-based primers and for 27 selected isolates by PCR-sequencing of IGS1 region of rDNA. RESULTS: Among 8257 yeast strains, 69 C. kefyr (including four bloodstream) isolates were detected by phenotypic and molecular methods. Isolation from urine and respiratory samples from female and male patients was significantly different (P = 0.001). Four isolates showed reduced susceptibility to amphotericin B and one isolate to all (amphotericin B, fluconazole, voriconazole and caspofungin/micafungin) antifungals tested. Fluconazole-resistant isolate contained only synonymous mutations in ERG11. Echinocandin-resistant isolate contained wild-type hotspot-1 and hotspot-2 of FKS1. Fingerprinting with microsatellite-/minisatellite-based primers identified only three types. IGS1 sequencing identified seven haplotypes among 27 selected isolates. CONCLUSIONS: The overall prevalence of C. kefyr among clinical yeast isolates and among candidemia cases was recorded as 0.83% and 0.32%, respectively. The frequency of isolation of C. kefyr from bloodstream and other invasive samples was stable during the study period. The C. kefyr isolates grown from invasive (bloodstream, bronchoalveolar lavage, abdominal drain fluid, peritonial fluid and gastric fluid) samples and amphotericin B-resistant isolates were genotypically heterogeneous strains.

Molecular identification, genotypic heterogeneity and comparative pathogenicity of environmental isolates of Papiliotrema laurentii.

Introduction. Papiliotrema laurentii, formerly Cryptococcus laurentii, is typically isolated from environmental sources, but also occasionally from clinical specimens. Other close relatives may be misidentified as P. laurentii by phenotypic methods. P. laurentii usually lacks melanin; however, melanin-forming strains have also been isolated.Hypothesis/Gap Statement. Although melanin production by encapsulated budding yeasts is considered a major virulence factor, the comparative pathogenicity of melanin-forming and non-melanized environmental strains of P. laurentii has rarely been studied.Aim. We performed phenotypic and molecular identification and determined the genotypic heterogeneity among P. laurentii isolates. We also studied the pathogenicity of melanin-forming and non-melanized strains in normal and immunosuppressed mice.Methodology. Eleven environmental isolates were tested for their identity by Vitek2 and/or ID32C systems, and by PCR-sequencing of the internal transcribed spacer (ITS) region and D1/D2 domains of ribosomal DNA (rDNA). Genotypic heterogeneity was studied by sequence comparisons. The pathogenicity of melanized and non-melanized P. laurentii strains was studied in intravenously infected normal and immunosuppressed BALB/c mice.Results. Phenotypic methods identified seven of the environmental isolates, while PCR-sequencing of the ITS region and D1/D2 domains of rDNA detected two and five isolates, respectively, as P. laurentii. Sequence comparisons demonstrated genotypic heterogeneity among P. laurentii. The remaining four environmental isolates yielded expected results. None of the normal mice infected with 105 cells of melanized/non-melanized P. laurentii strains died. Infection of immunosuppressed mice with 107 cells caused higher mortality with non-melanized P. laurentii, while viable counts in brain/lung tissue were higher in mice infected with a melanized strain and were detectable for up to 14 days.Conclusion. Phenotypic methods lacked specificity, but PCR-sequencing of D1/D2 domains correctly identified P. laurentii and sequence comparisons demonstrated the genotypic heterogeneity of the isolates. Both melanized and non-melanized strains at a higher dose caused mortality in immunosuppressed mice and persisted in brain/lung tissue up to 14 days post-infection.

Antifungal drug susceptibility, molecular basis of resistance to echinocandins and molecular epidemiology of fluconazole resistance among clinical Candida glabrata isolates in Kuwait.

Candida glabrata readily develops resistance to echinocandins. Identification, antifungal susceptibility testing (AST) and resistance mechanism to echinocandins among C. glabrata was determined in Kuwait. C. glabrata isolates (n = 75) were tested by Vitek2, multiplex PCR and/or PCR-sequencing of rDNA. AST to fluconazole, caspofungin, micafungin and amphotericin B was determined by Etest and to micafungin by broth microdilution (BMD). Mutations in hotspot-1/hotspot-2 of FKS1/FKS2 and ERG11 were detected by PCR-sequencing. All isolates were identified as C. glabrata sensu stricto. Seventy isolates were susceptible and five were resistant to micafungin by Etest and BMD (essential agreement, 93%; categorical agreement, 100%). Three micafungin-resistant isolates were resistant and two were susceptible dose-dependent to caspofungin. Four and one micafungin-resistant isolate contained S663P and ∆659 F mutation, respectively, in hotspot-1 of FKS2. Micafungin-resistant isolates were genotypically distinct strains. Only one of 36 fluconazole-resistant isolate contained nonsynonymous ERG11 mutations. Thirty-four of 36 fluconazole-resistant isolates were genotypically distinct strains. Our data show that micafungin susceptibility reliably identifies echinocandin-resistant isolates and may serve as a surrogate marker for predicting susceptibility/resistance of C. glabrata to caspofungin. All micafungin-resistant isolates also harbored a nonsynonymous/deletion mutation in hotspot-1 of FKS2. Fingerprinting data showed that echinocandin/fluconazole resistance development in C. glabrata is not clonal.

Increasing Trends of Reduced Susceptibility to Antifungal Drugs Among Clinical Candida glabrata Isolates in Kuwait.

Among non-albicans Candida species, Candida glabrata is the leading cause of invasive infections in critically ill patients. It is intrinsically less susceptible to fluconazole/other azoles that limits therapeutic options. This study determined distribution of C. glabrata in clinical specimens and determined their susceptibility to fluconazole, caspofungin, and amphotericin B by E test. During 8-year period (2011-2018), 1,410 isolates were obtained from 1,410 patients including 600, 409, and 131 isolates from respiratory, urine, and bloodstream specimens, respectively. Proportion of C. glabrata isolates was nearly the same during the two 4-year periods. Demographic details were available from 731 patients and susceptibility data for 1,225 isolates. C. glabrata isolation from bloodstream, respiratory, and urine specimens was higher from elderly (>60 years) versus younger patients. More bloodstream and urine isolates were obtained from female patients, however, more respiratory isolates were recovered from male patients (p = <0.05). Resistance to all three drugs increased during 2015-2018 compared with 2011-2014 but was more pronounced for fluconazole (p = 0.001). More isolates with reduced susceptibility to fluconazole/amphotericin B were obtained from elderly patients versus younger subjects and urine versus respiratory samples (p = <0.05). Our data show increasing trends of reduced susceptibility to antifungals, particularly fluconazole, among clinical C. glabrata isolates in Kuwait. Most isolates with reduced susceptibility to fluconazole/amphotericin B were obtained from elderly patients and urine/respiratory samples with urinary tract appearing as the most favorable niche for antifungal drug resistance development. The study also highlights the need for continued surveillance and better antifungal drug stewardship to control resistance development in C. glabrata.

Candida auris in various hospitals across Kuwait and their susceptibility and molecular basis of resistance to antifungal drugs.

BACKGROUND: Candida auris, a multidrug-resistant species, has the propensity of nosocomial transmission despite normal decontamination procedures. Here, we describe the isolation of C auris from patients in various hospitals in Kuwait during 2014-2018. Susceptibility to antifungal drugs and molecular basis of resistance to fluconazole, voriconazole and micafungin were also studied. METHODS: Candida auris (n = 314) obtained from 126 patients in eight hospitals were studied. All isolates were identified by PCR amplification and/or PCR-sequencing of ribosomal DNA (rDNA). Antifungal susceptibility was determined by Etest. Molecular basis of resistance to fluconazole and micafungin was studied by PCR-sequencing of ERG11 and FKS1 genes, respectively. FINDINGS: Bloodstream (n = 58), urine (n = 124), respiratory (n = 98) and other (n = 34) specimens yielded 314 C auris isolates. The proportion of bloodstream C auris among all yeast isolates was higher (42 of 307, 13.7%) in 2018 as compared to 2014-2017 (16 of 964, 1.7%) (P = .001). More bloodstream isolates (42 of 139) were cultured in 2018 than during 2014-2017 (16 of 175) (P = .001). Resistance to amphotericin B, fluconazole, voriconazole and micafungin was detected in 27.1%, 100%, 41.1% and 1.7% isolates, respectively. Fluconazole-resistant isolates contained either Y132F or K143R mutation in ERG11. Isolates with K143R mutation were additionally resistant to voriconazole. Micafungin-resistant isolates contained S639F mutation in hot spot 1 of FKS1. CONCLUSIONS: Our study highlights spreading of C auris in major hospitals across Kuwait and its increasing role as a bloodstream pathogen in 2018. Cross-resistance to voriconazole was also seen in isolates with K143R mutation in ERG11, while micafungin-resistant isolates harboured S639F mutation in hot spot 1 of FKS1.

Lack of detection of Candida nivariensis and Candida bracarensis among 440 clinical Candida glabrata sensu lato isolates in Kuwait.

Occurrence of Candida nivariensis and Candida bracarensis, two species phenotypically similar to Candida glabrata sensu stricto, in human clinical samples from different geographical settings remains unknown. This study developed a low-cost multiplex PCR (mPCR) and three species-specific singleplex PCR assays. Reference strains of common Candida species were used during development and the performance of mPCR and singleplex PCR assays was evaluated with 440 clinical C. glabrata sensu lato isolates. The internal transcribed spacer (ITS) region of rDNA was also sequenced from 85 selected isolates and rDNA sequence variations were used for determining genetic relatedness among the isolates by using MEGA X software. Species-specific amplicons for C. glabrata (~360 bp), C. nivariensis (~250 bp) and C. bracarensis (~180 bp) were obtained in mPCR while no amplicon was obtained from other Candida species. The three singleplex PCR assays also yielded expected results with reference strains of Candida species. The mPCR amplified ~360 bp amplicon from all 440 C. glabrata sensu lato isolates thus identifying all clinical isolates in Kuwait as C. glabrata sensu stricto. The results of mPCR were confirmed for all 440 isolates as they yielded an amplicon only in C. glabrata sensu stricto-specific singleplex PCR assay. The rDNA sequence data identified 28 ITS haplotypes among 85 isolates with 18 isolates belonging to unique haplotypes and 67 isolates belonging to 10 cluster haplotypes. In conclusion, we have developed a simple, low-cost mPCR assay for rapid differentiation of C. glabrata sensu stricto from C. nivariensis and C. bracarensis. Our data obtained from a large collection of clinical C. glabrata sensu lato isolates show that C. nivariensis and C. bracarensis are rare pathogens in Kuwait. Considerable genetic diversity among C. glabrata sensu stricto isolates was also indicated by rDNA sequence analyses.

Magnusiomyces capitatus fungemia: The value of direct microscopy in early diagnosis.

Two cases of fungemia caused by Magnusiomyces capitatus, an arthroconidial yeast-like fungus, in non-hematologic immunocompromised patients are described. Both patients died before definite diagnosis of M. capitatus was made. The report highlights that pending confirmation of the isolate by phenotypic and/or molecular methods, the characteristic morphologic features observed in Gram-stained smears of blood culture positive bottles can lead to early preliminary diagnosis, thus significantly reducing time required for initiating appropriate antifungal therapy.

In vitro Post-Antifungal Effect of Posaconazole and Its Impact on Adhesion-Related Traits and Hemolysin Production of Oral Candida dubliniensis Isolates.

OBJECTIVE: Candidal adherence to denture acrylic surfaces (DAS) and oral buccal epithelial cells (BEC), formation of candidal germ tubes (GT), candidal cell surface hydrophobicity (CSH), and hemolysin production are important pathogenic traits of Candida. The antifungal drug-induced post-antifungal effect (PAFE) also impacts the virulence of Candida. Candida dubliniensis isolates are associated with the causation of oral candidiasis which could be managed with posaconazole. Thus far there is no evidence on posaconazole-induced PAFE and its impact on adhesion-related attributes and production of hemolysin by C. dubliniensis isolates. Hence, the PAFE, adhesion to DAS and BEC, formation of GT, CSH, and hemolysin production of 20 oral C. dubliniensis isolates after brief exposure to posaconazole was ascertained. MATERIALS AND METHODS: The PAFE, adherence to DAS and BEC, formation of GT, candidal CSH, and hemolysin production were investigated by hitherto described in vitro assays. RESULTS: The mean PAFE (h) induced by posaconazole on C. dubliniensis isolates was 1.66. Exposure to posaconazole suppressed the ability of C. dubliniensis to adhere to DAS, BEC, formation of candidal GT, candidal CSH and to produce hemolysin by a reduction of 44, 33, 34, 36, and 15% (p < 0.005 to p < 0.001), respectively. CONCLUSION: Exposure of C. dubliniensis isolates to posaconazole for a brief period induced an antimycotic impact by subduing its growth in addition to suppressing pathogenic adherence-associated attributes, as well as production of hemolysin.

Changing trends in epidemiology and antifungal susceptibility patterns of six bloodstream Candida species isolates over a 12-year period in Kuwait.

Changing trends in incidence and antifungal susceptibility patterns of six Candida species causing candidemia in Kuwait between 2006-2017 are reported. A total of 2075 isolates obtained from 1448 patients were analyzed. Identity of Candida species isolates was determined by phenotypic methods and confirmed by PCR amplification/PCR-sequencing of rDNA and/or MALDI-TOF MS. Antifungal susceptibility was determined by Etest. C. albicans accounted for 539 (37.22%) cases followed by C. parapsilosis (n = 502, 34.67%), C. tropicalis (n = 210, 14.5%), C. glabrata (n = 148, 10.22%), C. krusei (n = 27, 1.81%) and C. dubliniensis (n = 22, 1.5%). The comparative percent distribution of Candida species causing candidemia between 2006-2011 and 2012-2017 was as follows: C. albicans 41.8% and 33.1%, C. parapsilosis complex 32.01% and 37.04%, C. tropicalis 13.59% and 15.31%, and C. glabrata 8.77% and 11.51%, C. krusei 2.0% and 1.7%, and C. dubliniensis 1.75 and 1.3%, respectively. Three of 371 C. albicans isolates during 2006-2011 and five of 363 during 2012-2017 were resistant to fluconazole. Among C. parapsilosis isolates, one of 310 during 2006-2011 and 21 of 446 during 2012-2017 were resistant to this drug. Furthermore, at an epidemiologic cutoff value (ECV) of ≤0.5 µg/ml, 70.1% C. albicans isolates were wild-type for fluconazole during 2006-2011 as compared to 58.1% during 2012-2017. Likewise, at an ECV of ≤2 µg/ml, 98.0% of C. parapsilosis isolates were wild-type during 2006-2011 as compared to 93.4% during 2012-2017. Clonal spread of fluconazole-resistant C. parapsilosis in one major hospital was documented. An 8.8% shift in favor of non-albicans Candida species with concomitant increase in MICs between the two periods preludes emergence of fluconazole-resistant candidemia cases in Kuwait.

Candida lusitaniae in Kuwait: Prevalence, antifungal susceptibility and role in neonatal fungemia.

OBJECTIVES: Candida lusitaniae is an opportunistic yeast pathogen in certain high-risk patient populations/cohorts. The species exhibits an unusual antifungal susceptibility profile with tendency to acquire rapid resistance. Here, we describe prevalence of C. lusitaniae in clinical specimens in Kuwait, its antifungal susceptibility profile and role in neonatal fungemia. METHODS: Clinical C. lusitaniae isolates recovered from diverse specimens during 2011 to 2017 were retrospectively analyzed. All isolates were identified by germ tube test, growth on CHROMagar Candida and by Vitek 2 yeast identification system. A simple species-specific PCR assay was developed and results were confirmed by PCR-sequencing of ITS region of rDNA. Antifungal susceptibility was determined by Etest. Minimum inhibitory concentrations (MICs) were recorded after 24 h incubation at 35°C. RESULTS: Of 7068 yeast isolates, 134 (1.89%) were identified as C. lusitaniae including 25 (2.52%) among 990 bloodstream isolates. Species-specific PCR and PCR-sequencing of rDNA confirmed identification. Of 11 cases of neonatal candidemia, 9 occurred in NICU of Hospital A and are described here. Eight of 9 neonates received liposomal amphotericin B, which was followed by fluconazole in 7 and additionally by caspofungin in 2 cases as salvage therapy. Three of 8 (37.5%) patients died. No isolate exhibited reduced susceptibility to amphotericin B, fluconazole, voriconazole, caspopfungin, micafungin and anidulafungin. The MIC ± geometric mean values for amphotericin B, fluconazole, voriconazole, and caspofungin were as follows: 0.072 ± 0.037 µg/ml, 2.32 ± 0.49 µg/ml, 0.09 ± 0.01 µg/ml and 0.16 ± 0.08 µg/ml, respectively. Only two isolates exhibited reduced susceptibility to fluconazole. CONCLUSIONS: This study describes the prevalence and antifungal susceptibility profile of clinical C. lusitaniae isolates in Kuwait. No isolate showed reduced susceptibility to amphotericin B. The study highlights the emerging role of C. lusitaniae as a healthcare-associated pathogen capable of causing fungemia in preterm neonates and causing significant mortality.

High-resolution fingerprinting of Candida parapsilosis isolates suggests persistence and transmission of infections among neonatal intensive care unit patients in Kuwait.

Candida parapsilosis causes ~35% of all candidemia cases in neonates. High-resolution fingerprinting of C. parapsilosis isolates from neonatal intensive care unit (NICU) patients in Maternity Hospital (MH) was performed to identify epidemiologically related strains. Sixty-eight bloodstream/colonizing strains isolated from 59 NICU patients, two isolates from health care workers (HCWs) from MH and 18 bloodstream isolates from two other hospitals were used. Six microsatellite markers were employed, isolates were assigned a numerical microsatellite genotype (MSG), dendrogram was constructed and similarities between genotypes were visualized by minimum spanning tree. Fifty bloodstream isolates from MH yielded 37 MSGs with 20 isolates clustering in 7 MSGs. Duplicate isolates and colonizing strains yielded same/highly similar MSG as bloodstream isolates. Colonizing strains from two non-candidemia patients yielded unique MSGs while others belonged to a cluster. All isolates from HCWs and from two other hospitals belonged to unique MSGs. Cluster isolates came from patients in NICU-1 or from neonates in NICU-1 and other NICUs. Clonal complexes comprising closely related genotypes indicative of microevolution were also detected. Our data show that some C. parapsilosis strains have persisted in MH environment over several years and these endemic genotypes were transmitted to other patients in NICU-1 and/or other nearby NICUs.

Demonstration of Adventitious Sporulation in Fusarium Petroliphilum Onychomycosis.

Fusarium petroliphilum is a recently described species within the Fusarium solani species complex. Some strains of Fusarium species are capable of forming yeast-like structures in tissue as well as in culture through a process known as "adventitious sporulation." Here, we describe the formation of these yeast-like reproductive structures in infected nail tissue obtained from a case of onychomycosis. These structures were also observed in culture grown on RPMI 1640 agar supplemented with 2% glucose. The isolate was resistant to azoles and echinocandins. To the best of our knowledge, this is the first report describing adventitious sporulation in F. petroliphilum and its etiologic role in onychomycosis.

Novel multiplex real-time quantitative PCR detecting system approach for direct detection of Candida auris and its relatives in spiked serum samples.

The multidrug-resistant opportunistic yeast species of Candida auris, Candida haemulonii, Candida duobushaemulonii and Candida pseudohaemulonii continue to endanger the healthcare settings around the globe. Due to the lack of a specific qPCR assay for detection of these species from clinical samples, we developed a multiplex qPCR assay. Analytical specificity and sensitivity showed 100% specificity and the sensitivity of up to ten genomes of target species with a high value of reproducibility (R2 >0.99). Subsequently, from spiked serum samples, our qPCR specifically could detect up to ten genomes of C. auris and one genome of C. haemulonii, C. duobushaemulonii and C. pseudohaemulonii (R2 >0.98). Lack of cross reaction with the human DNA, a high degree of specificity and sensitivity, showed the potential of our multiplex PCR for direct detection of C. auris and closely related species from serum samples of suspected patients. Future studies are warranted to assure its applicability in clinical settings.

Cyberlindnera fabianii fungaemia outbreak in preterm neonates in Kuwait and literature review.

BACKGROUND: Cyberlindnera fabianii has rarely been reported as a human pathogen. Here, we describe an outbreak of C. fabianii fungaemia involving 10 preterm neonates during a seven-month period in Kuwait and review the published reports. METHODS: Blood cultures were processed, and yeast isolates were initially identified by ID 32 C and/or VITEK 2. Molecular identification was done by PCR sequencing of internally transcribed spacer (ITS) region and D1/D2 domains of rDNA. Fingerprinting was performed with microsatellite-based and minisatellite-based primers to examine genetic relatedness among the isolates. Antifungal susceptibility testing of the isolates was done by Etest. FINDINGS: All infected neonates were preterm, received prior antibiotics and had an intravascular catheter in place. All bloodstream isolates were initially identified as Candida utilis by ID 32 C and/or VITEK 2 and showed reduced susceptibility to triazoles. PCR sequencing of rDNA identified all isolates as Cyberlindnera fabianii. Fingerprinting studies yielded identical patterns indicating clonality. One neonate died before treatment, one died during treatment, and eight neonates survived treatment with amphotericin B with/without fluconazole or caspofungin. Source of infection remained unknown despite surveillance cultures. CONCLUSION: The outbreak highlights emergence of C. fabianii as a neonatal pathogen and reinforces importance of molecular methods in its accurate identification.