Fungal Endocarditis: Pathophysiology, Epidemiology, Clinical Presentation, Diagnosis, and Management.

Fungal endocarditis accounts for 1% to 3% of all infective endocarditis cases, is associated with high morbidity and mortality (>70%), and presents numerous challenges during clinical care. Candida spp. are the most common causes of fungal endocarditis, implicated in over 50% of cases, followed by Aspergillus and Histoplasma spp. Important risk factors for fungal endocarditis include prosthetic valves, prior heart surgery, and injection drug use. The signs and symptoms of fungal endocarditis are nonspecific, and a high degree of clinical suspicion coupled with the judicious use of diagnostic tests is required for diagnosis. In addition to microbiological diagnostics (e.g., blood culture for Candida spp. or galactomannan testing and PCR for Aspergillus spp.), echocardiography remains critical for evaluation of potential infective endocarditis, although radionuclide imaging modalities such as 18F-fluorodeoxyglucose positron emission tomography/computed tomography are increasingly being used. A multimodal treatment approach is necessary: surgery is usually required and should be accompanied by long-term systemic antifungal therapy, such as echinocandin therapy for Candida endocarditis or voriconazole therapy for Aspergillus endocarditis.

Assessment of micafungin dosage regimens against Candida spp. in pediatric patients undergoing hematopoietic stem cell transplantation: a pharmacokinetic/pharmacodynamic analysis using Monte Carlo simulation.

The objective of this study was to evaluate the efficacy of various micafungin dosing regimens against Candida spp. in pediatric patients undergoing hematopoietic stem cell transplantation (HSCT). Monte Carlo simulations were conducted using pharmacokinetic (PK) parameters and pharmacodynamic (PD) data to determine the probabilities of target attainment and cumulative fractions of response in terms of area under the concentration curve/minimum inhibition concentration targets of micafungin. Current standard clinical micafungin dosing regimens of 1 and 2 mg/kg/day were appropriate for the prevention and treatment of Candida glabrata infection in pediatric patients undergoing HSCT, respectively. Moreover, the high-dose prophylactic dosage (2 mg/kg/day) and therapeutic dosage (4 mg/kg/day) should be the preferred option to optimize efficacy against Candida albicans. However, none of the simulated regimens was effective against Candida parapsilosis in pediatric HSCT patients. These PK/PD-based simulations rationalize and optimize the micafungin dosing regimens against Candida spp. in pediatric patients undergoing HSCT.

Assessment of Risk Factors and Clinical Outcomes in Hospitalized COVID-19 Patients with Candida spp. Co-infections: Species Distribution and Antifungal Susceptibility Patterns of Isolates.

INTRODUCTION: Fungal co-infections are considered an important complication in hospitalized patients with SARS-CoV-2 that can be attributed to disease aggravation, increased mortality, and poor outcomes. This study was conducted to determine the species distribution and antifungal susceptibility patterns of Candida isolates from hospitalized COVID-19 patients in Shiraz, Iran, in addition to associated risk factors and outcomes of co-infections with Candida species. MATERIALS AND METHODS: In this single-center study, a total of 106 hospitalized COVID-19 patients were evaluated for clinical characteristics and outcomes. Species identification was performed by ITS1-5.8S-ITS2 gene sequencing. Antifungal susceptibility testing to fluconazole, itraconazole, voriconazole, posaconazole, caspofungin, amphotericin B, and nystatin was determined according to the M27-A3/S4 CLSI protocol. RESULTS: Candida species were recovered from 48% (51/106) of hospitalized COVID-19 patients. Statistical analysis showed that patients who had heart failure, bacterial co-infection, and were receiving empirical antifungal therapy had a higher risk of developing Candida co-infection. In total, 71 Candida isolates were recovered, of which C. albicans (69%) was the most prevalent isolate. The majority of the Candida isolates were susceptible to all classes of tested antifungal drugs. DISCUSSION: Our results elucidate a high rate of Candida co-infections among hospitalized COVID-19 patients. Comorbidities such as heart failure, HTN, COPD, bacterial infections as well as therapeutic interventions including catheterization, mechanical ventilation, and ICU admission increased the risk of Candida spp. isolation from the bloodstream, respiratory tract and urine samples, which led to a higher in-hospital mortality rate. Additionally, obtained data clarified that empirical antifungal therapy was not as successful as anticipated.

Curcumin/H2O2 photodynamically activated: an antimicrobial time-response assessment against an MDR strain of Candida albicans.

OBJECTIVE: Human candidiasis is typically treated with antifungal drugs, but the rise of drug-resistant strains of Candida spp. poses a serious problem, making treatment difficult. At the same time, photodynamic therapy (PDT) has drawn increasing attention from researchers for its potential to effectively inhibit multidrug-resistant pathogenic fungi and for its low tendency to induce drug resistance. This study's goal was to examine how a multidrug-resistant oral isolate of Candida albicans responded to a PDT that used a curcumin/H202 formulation as a photosensitizer and was exposed to various light sources. MATERIALS AND METHODS: A commercial product containing curcumin/H2O2 3% was used as a photosensitizer and evaluated in a PDT treatment that can use two different light sources: traditional irradiation with 7 W light at λ = 460 nm or a new, never evaluated, polarized light source of 25 W with a wavelength range of λ = 380-3,400 nm. The antimicrobial activity of these procedures was assessed on a clinical oral isolate of Candida albicans, in terms of agar susceptibility test, growth curve behavior, and biofilm inhibition. RESULTS: Both light sources were able to activate the photosensitizer formulation, suggesting a fungistatic activity vs. this C. albicans MDR strain. An interesting difference was observed in the cell-generation-time (CGTOD) after PDT treatment, where the polarized light was more active compared to the source of 460 nm. In fact, CGTOD was 16 and 8 hours, respectively. CONCLUSIONS: The PDT evaluated here presented an inhibition window time, a crucial point for clinicians, who could activate an additional prophylactic treatment to resolve the clinical management of Candida infections in the oral cavity.

Epidemiological Analysis And Successful Management Of A Candida Auris Outbreak In A Secondary Care Hospital Setting In Saudi Arabia; An Outbreak Report.

Candida auris has emerged as a major diagnostic and therapeutic challenge in the hospital environment. C. auris is resistant to many antifungals, making it a newer example of one of the world's most problematic and feared health threats. We recently confronted a cluster of C. auris cases at our hospital during the spring of 2020.This outbreak investigation took place at the ICU of King Khalid hospital Al Majmaah Saudi Arabia. Considering its potential to cause an outbreak with serious consequences, strict control measures were implemented thus effectively controlling the outbreak.

Epidemiology, management and outcome of candidaemia in patients with diabetes.

BACKGROUND: Candidaemia is the commonest fungal bloodstream infection in hospitalised patients. Diabetes is one of the risk factors for mortality from candidaemia. METHODS: We compared the epidemiology, clinical characteristics and management of candidaemia in patients with and without diabetes. RESULTS: Over a 10-year period, 200 episodes of Candida bloodstream infection were documented. Patients with diabetes were younger (58.7 vs 65.5 years), less likely to be suffering from cancer (21.8% vs 36%), and had significantly lower 30-day and 90-day crude mortality (17.2% vs 35.6% and 28.4% vs 48.6%, respectively). Candida glabrata was more common in patients with diabetes (39.3% vs 29.7%). Based on European Confederation of Medical Mycology (ECMM) quality indicators, the management of patients with and without diabetes was similar. DISCUSSION: Our study highlights the importance of epidemiological data in relation to candidaemia in patients with diabetes and the growing threat of invasive C. glabrata infection in this subset of patients.

Pharmacokinetic/Pharmacodynamic Target Attainment of Different Antifungal Agents in De-escalation Treatment in Critically Ill Patients: a Step toward Dose Optimization Using Monte Carlo Simulation.

Differences in pharmacokinetics/pharmacodynamics (PK/PD) target attainment are rarely considered when antifungals are switched in critically ill patients. This study intends to explore whether the antifungal de-escalation treatment strategy and the new intermittent dosing strategy of echinocandins in critically ill patients are able to achieve the corresponding PK/PD targets. The published population PK models of antifungals in critically ill patients and a public data set from the MIMIC-III database (n = 662) were employed to evaluate PK/PD target attainment of different dosing regimens of antifungals. Cumulative fraction of response (CFR) was calculated for each dosing regimen. Most guideline-recommended dosing regimens of fluconazole and voriconazole could achieve target exposure as de-escalation treatment in critically ill patients. For initial echinocandin treatment, achievement of the target exposure decreased as body weight increased, and the intermittent dosing strategy had a slightly higher CFR value in most simulations compared to conventional dosing strategy. For Candida albicans and Candida glabrata infection, caspofungin at the lowest dose achieved a CFR of >90%, while micafungin or anidulafungin required almost the highest doses simulated in this study to achieve the same effect. None of the echinocandins other than 150 mg every 24 h (q24h) or 200 mg q48h of caspofungin achieved the target CFR for Candida parapsilosis infection. These findings support the guideline-recommended dose of triazoles for antifungal de-escalation treatment and confirm the insufficient dosage of echinocandins in critically ill patients, indicating that a dosing regimen based on body weight or intermittent dosing of echinocandins may be required.

Risk assessment of malignant transformation of oral leukoplakia in patients with previous oral squamous cell carcinoma.

The aim of this study was to identify the risk factors associated with developing oral squamous cell carcinoma (OSCC) from surgically excised oral leukoplakia (OL) in patients with previous oral cavity cancer. Clinicopathological data of 84 patients who were treated for OL between July 2002 and July 2020 and who had previously received treatment for OSCC were reviewed retrospectively. The follow-up time ranged from 0.69 to 17.99 years (mean 6.78 ± 4.25 years). The overall cumulative malignant transformation rate was 25% and the annual transformation rate was 5.73%. Kaplan-Meier survival analysis and the log-rank test showed that Candida infection (P = 0.010) was a risk factor associated with malignant transformation. In the multivariate Cox regression analysis, tongue and floor of the mouth as the location of the leukoplakia (P = 0.039), multifocal lesions of OL (P = 0.047), and Candida infection (P = 0.018) were the three independent prognostic factors related to the development of OSCC from the treated OL. A cautious approach to OL of the tongue with Candida infection or multifocal disease in this group of patients would be appropriate.

Estimated burden of serious fungal infections in Togo.

BACKGROUND: Over the years, the focus of infectious diseases in many African countries has been mainly on viral, bacterial and parasitic infections. Serious fungal infections (SFIs) with comparable morbidity rate in these countries remain neglected. OBJECTIVES: To estimate the burden of SFI in Togo and to stimulate efforts for improved attention. METHODS: Literature was thoroughly searched for epidemiological data on SFI in Togo. Incidence and/or prevalence of SFI was estimated using socio-demographics, health system's information, risk-groups data and SFI rates obtained from national and international studies. RESULTS: About 5.29% of the 7,265,286 Togolese population is estimated to suffer from SFI annually. Among HIV patients, 1,342, 1,650 and 330 may develop cryptococcal meningitis, Pneumocystis pneumonia and disseminated histoplasmosis respectively per year. Oral and oesophageal candidiasis may annually affect 19,800 and 7,535 persons, respectively, living with HIV. Estimated incidence of invasive aspergillosis (IA) was 283 cases. Prevalence of chronic pulmonary aspergillosis (CPA) was estimated at 191 cases. The annual incidence of allergic bronchopulmonary aspergillosis (ABPA) and severe asthma with fungal sensitization (SAFS) was 4,577 and 6,042 cases, respectively. Tinea capitis and recurrent Candida vaginitis presumably affect 232,271 children and 108,979 women respectively. Candidaemia incidence is estimated at 5 cases per 100, 000 inhabitants and fungal keratitis may affect 981 persons annually. CONCLUSIONS: SFIs in Togo are probably more significant than expected. These findings underscore the need to increase awareness among healthcare professionals, enhance diagnostic and therapeutic capacities and intensify epidemiological studies for effective management of fungal infections in Togo.

Rapid Assessment and Containment of Candida auris Transmission in Postacute Care Settings-Orange County, California, 2019.

BACKGROUND: Candida auris, a multidrug-resistant yeast, can spread rapidly in ventilator-capable skilled-nursing facilities (vSNFs) and long-term acute care hospitals (LTACHs). In 2018, a laboratory serving LTACHs in southern California began identifying species of Candida that were detected in urine specimens to enhance surveillance of C auris, and C auris was identified in February 2019 in a patient in an Orange County (OC), California, LTACH. Further investigation identified C auris at 3 associated facilities. OBJECTIVE: To assess the prevalence of C auris and infection prevention and control (IPC) practices in LTACHs and vSNFs in OC. DESIGN: Point prevalence surveys (PPSs), postdischarge testing for C auris detection, and assessments of IPC were done from March to October 2019. SETTING: All LTACHs (n = 3) and vSNFs (n = 14) serving adult patients in OC. PARTICIPANTS: Current or recent patients in LTACHs and vSNFs in OC. INTERVENTION: In facilities where C auris was detected, PPSs were repeated every 2 weeks. Ongoing IPC support was provided. MEASUREMENTS: Antifungal susceptibility testing and whole-genome sequencing to assess isolate relatedness. RESULTS: Initial PPSs at 17 facilities identified 44 additional patients with C auris in 3 (100%) LTACHs and 6 (43%) vSNFs, with the first bloodstream infection reported in May 2019. By October 2019, a total of 182 patients with C auris were identified by serial PPSs and discharge testing. Of 81 isolates that were sequenced, all were clade III and highly related. Assessments of IPC identified gaps in hand hygiene, transmission-based precautions, and environmental cleaning. The outbreak was contained to 2 facilities by October 2019. LIMITATION: Acute care hospitals were not assessed, and IPC improvements over time could not be rigorously evaluated. CONCLUSION: Enhanced laboratory surveillance and prompt investigation with IPC support enabled swift identification and containment of C auris. PRIMARY FUNDING SOURCE: Centers for Disease Control and Prevention.

Serious fungal diseases in Democratic Republic of Congo - Incidence and prevalence estimates.

A literature review was conducted to assess the burden of serious fungal infections in the Democratic Republic of the Congo (DRC) (population 95,326,000). English and French publications were listed and analysed using PubMed/Medline, Google Scholar and the African Journals database. Publication dates spanning 1943-2020 were included in the scope of the review. From the analysis of published articles, we estimate a total of about 5,177,000 people (5.4%) suffer from serious fungal infections in the DRC annually. The incidence of cryptococcal meningitis, Pneumocystis jirovecii pneumonia in adults and invasive aspergillosis in AIDS patients was estimated at 6168, 2800 and 380 cases per year. Oral and oesophageal candidiasis represent 50,470 and 28,800 HIV-infected patients respectively. Chronic pulmonary aspergillosis post-tuberculosis incidence and prevalence was estimated to be 54,700. Fungal asthma (allergic bronchopulmonary aspergillosis and severe asthma with fungal sensitization) probably has a prevalence of 88,800 and 117,200. The estimated prevalence of recurrent vulvovaginal candidiasis and tinea capitis is 1,202,640 and 3,551,900 respectively.Further work is required to provide additional studies on opportunistic infections for improving diagnosis and the implementation of a national surveillance programme of fungal disease in the DRC.

The fitness costs and benefits of trisomy of each Candida albicans chromosome.

Candida albicans is a prevalent human fungal pathogen. Rapid genomic change, due to aneuploidy, is a common mechanism that facilitates survival from multiple types of stresses including the few classes of available antifungal drugs. The stress survival of aneuploids occurs despite the fitness costs attributed to most aneuploids growing under idealized lab conditions. Systematic study of the aneuploid state in C. albicans has been hindered by the lack of a comprehensive collection of aneuploid strains. Here, we describe a collection of diploid C. albicans aneuploid strains, each carrying one extra copy of each chromosome, all from the same genetic background. We tested the fitness of this collection under several physiological conditions including shifts in pH, low glucose, oxidative stress, temperature, high osmolarity, membrane stress, and cell wall stress. We found that most aneuploids, under most conditions, were less fit than their euploid parent, yet there were specific conditions under which specific aneuploid isolates provided a fitness benefit relative to the euploid parent strain. Importantly, this fitness benefit was attributable to the change in the copy number of specific chromosomes. Thus, C. albicans can tolerate aneuploidy of each chromosome and some aneuploids confer improved growth under conditions that the yeast encounters in its host niches.

Comparative assessment of immune evasion mechanisms in human whole-blood infection assays by a systems biology approach.

Computer simulations of mathematical models open up the possibility of assessing hypotheses generated by experiments on pathogen immune evasion in human whole-blood infection assays. We apply an interdisciplinary systems biology approach in which virtual infection models implemented for the dissection of specific immune mechanisms are combined with experimental studies to validate or falsify the respective hypotheses. Focusing on the assessment of mechanisms that enable pathogens to evade the immune response in the early time course of a whole-blood infection, the least-square error (LSE) as a measure for the quantitative agreement between the theoretical and experimental kinetics is combined with the Akaike information criterion (AIC) as a measure for the model quality depending on its complexity. In particular, we compare mathematical models with three different types of pathogen immune evasion as well as all their combinations: (i) spontaneous immune evasion, (ii) evasion mediated by immune cells, and (iii) pre-existence of an immune-evasive pathogen subpopulation. For example, by testing theoretical predictions in subsequent imaging experiments, we demonstrate that the simple hypothesis of having a subpopulation of pre-existing immune-evasive pathogens can be ruled out. Furthermore, in this study we extend our previous whole-blood infection assays for the two fungal pathogens Candida albicans and C. glabrata by the bacterial pathogen Staphylococcus aureus and calibrated the model predictions to the time-resolved experimental data for each pathogen. Our quantitative assessment generally reveals that models with a lower number of parameters are not only scored with better AIC values, but also exhibit lower values for the LSE. Furthermore, we describe in detail model-specific and pathogen-specific patterns in the kinetics of cell populations that may be measured in future experiments to distinguish and pinpoint the underlying immune mechanisms.

Use of Novasomes as a Vesicular Carrier for Improving the Topical Delivery of Terconazole: In Vitro Characterization, In Vivo Assessment and Exploratory Clinical Experimentation.

PURPOSE: This manuscript aimed at encapsulating an antifungal terconazole (TCZ) into innovative novasomes for improving its penetration into the skin and clinically modulating its therapeutic efficacy. METHODS: Novasomes containing free fatty acid (FFA) as a penetration enhancer were formulated using ethanol injection technique based on 24 full factorial design to explore the impact of various formulation variables on novasomes characteristics regarding entrapment efficiency percent (EE%), particle size (PS), polydispersity index (PDI), and zeta potential (ZP). The optimum formulation was chosen using Design-Expert® software and utilized for further explorations. RESULTS: The chosen formulation (N15; including 100 mg lipid components and Span 80 to oleic acid in a ratio of 2:1 (w/w)) exhibited an EE% = 99.45 ± 0.78%, PS = 623.00 ± 2.97 nm, PDI = 0.40 ± 0.04, and ZP = -73.85 ± 0.64 mV. N15 showed spherical vesicles with a higher deformability index (DI) (9.62 ± 0.15 g) compared to traditional niosomal formulation (0.92 ± 0.12 g). Further, N15 showed superior inhibition of Candida albicans growth relative to TCZ suspension using XTT (2,3-bis-(2-methyloxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide) reduction assay. Moreover, in vivo skin deposition tests revealed a superior TCZ deposition inside the skin from N15 in comparison to traditional niosomal formulation and TCZ suspension. Furthermore, histopathological examination for rats assured the safety of N15 for topical use. A clinical study conducted on infants suffering from napkin candidiasis proved the superiority of N15 to placebo in providing a complete cure of such fungal infections. CONCLUSION: Concisely, the obtained outcomes confirmed the pronounced efficacy of N15 to successfully treat skin fungal infections.

Optimization of Micafungin Dosage for Chinese Patients with Sepsis in the Intensive Care Unit Based on a Population Pharmacokinetic-Pharmacodynamic Analysis.

PURPOSE: This study aimed to identify parameters that influence micafungin pharmacokinetics in Chinese patients with sepsis in the intensive care unit and optimize micafungin dosage by determining the probability of reaching pharmacodynamic targets. METHODS: Blood samples were collected from 32 Chinese patients with sepsis who were treated with micafungin. The samples were analyzed and used to build a population pharmacokinetic model. Monte Carlo simulations were performed to estimate the probability of achieving adequate plasma levels of micafungin against Candida species. RESULTS: Alanine aminotransferase and sequential organ failure assessment score were found to significantly influence the clearance and peripheral distribution volume of micafungin, respectively. Monte Carlo simulations based on area under the plasma concentration-time curve over 24 h showed that patients must be administered at least 200 and 250 mg micafungin daily to reach minimum inhibitory concentration breakpoints of 0.032 and 0.064 mg/L for Candida glabrata and Candida tropicalis, respectively. Additionally, a probability of target attainment of ≥ 90% could not be achieved for Candida krusei or Candida parapsilosis with a 300 mg daily dose. CONCLUSIONS: The recommended daily dose of micafungin (100 mg) may produce low clinical success ratios in non-Candida albicans infections; therefore, higher doses should be administered to improve clinical outcomes.

Impact of Loading Dose of Caspofungin in Pharmacokinetic-Pharmacodynamic Target Attainment for Severe Candidiasis Infections in Patients in Intensive Care Units: the CASPOLOAD Study.

This study evaluated the impact of a high loading dose of caspofungin (CAS) on the pharmacokinetics of CAS and the pharmacokinetic-pharmacodynamic (PK-PD) target attainment in patients in intensive care units (ICU). ICU patients requiring CAS treatment were prospectively included to receive a 140-mg loading dose of CAS. Plasma CAS concentrations (0, 2, 3, 5, 7, and 24 h postinfusion) were determined to develop a two-compartmental population PK model. A Monte Carlo simulation was performed and the probabilities of target attainment (PTAs) were computed using previously published MICs. PK-PD targets were ratios of area under the concentration-time curve from 0 to 24 h (AUC0-24h) divided by the MIC (AUC0-24h/MIC) of 250, 450, and 865 and maximal concentration (Cmax) divided by the MIC (Cmax/MIC) of 5, 10, 15, and 20. Among 13 included patients, CAS clearance was 0.98 ± 0.13 liters/h and distribution volumes were V1 = 9.0 ± 1.2 liters and V2 = 11.9 ± 2.9 liters. Observed and simulated CAS AUC0-24h were 79.1 (IQR 55.2; 108.4) and 81.3 (IQR 63.8; 102.3) mg · h/liter during the first 24 h of therapy, which is comparable to values usually observed in ICU patients at day 3 or later. PTAs were >90% for MICs of 0.19 and 0.5 mg/liter, considering AUC/MIC = 250 and Cmax/MIC = 10 as PK-PD targets, respectively. Thus, a high loading dose of CAS (140 mg) increased CAS exposure in the first 24 h of therapy, allowing early achievement of PK-PD targets for most Candida strains. Such a strategy seems to improve treatment efficacy, though further studies are needed to assess the impact on clinical outcomes. (This study has been registered at ClinicalTrials.gov under identifier NCT02413892.).

Assessment of Silver Levels in a Closed-Incision Negative Pressure Therapy Dressing: In Vitro and In Vivo Study.

Objective: In recent years, reticulated open-cell foam-based closed-incision negative pressure therapy (ROCF-ciNPT) has shown effectiveness in management of various postoperative incisions. These dressings consist of a skin interface layer that absorbs fluid from the skin surface and reduces the potential for microbial colonization within the dressing by means of ionic silver. This study examines the ability of silver to reduce the bioburden within the dressing as well as the localized effect due to potential silver mobility. Approach: Ability of silver to reduce bioburden within the ROCF-ciNPT dressing was assessed using Staphylococcus aureus, Pseudomonas aeruginosa, and Candida spp. Furthermore, silver mobility was assessed using an in vitro skin model to study the zone of inhibition along with released silver quantification. Using a porcine model, diffusion of silver into blood and tissue was studied using emission spectrometry and histology. Results: Microbial growth in the ROCF-ciNPT dressing was significantly reduced (∼2.7-4.9 log reduction) compared to a silver-free negative control. No zone of inhibition was observed for microbial colonies for up to 7 days with minimal localized silver release (<5.5 ppm release). In vivo studies demonstrated no measurable concentration (<0.2 µg/g) of silver in the blood, urine, feces, kidney, and liver tissue biopsy. Innovation: This study provides an important insight into silver concentration and mobility within the ROCF-ciNPT dressing, given emerging concerns associated with potential silver cytotoxicity. Conclusion: These results indicate the concentration of silver (0.019% silver by weight) in the ROCF-ciNPT dressings has been adequate to reduce bioburden within the skin interface layer, while severely limiting the amount of silver leaching out.

An Intact Cell Bioluminescence-Based Assay for the Simple and Rapid Diagnosis of Urinary Tract Infection.

Urinary tract infection (UTI) is one of the most common infections, accounting for a substantial portion of outpatient hospital and clinic visits. Standard diagnosis of UTI by culture and sensitivity can take at least 48 h, and improper diagnosis can lead to an increase in antibiotic resistance following therapy. To address these shortcomings, rapid bioluminescence assays were developed and evaluated for the detection of UTI using intact, viable cells of Photobacterium mandapamensis USTCMS 1132 or previously lyophilized cells of Photobacterium leiognathi ATCC 33981™. Two platform technologies-tube bioluminescence extinction technology urine (TuBETUr) and cellphone-based UTI bioluminescence extinction technology (CUBET)-were developed and standardized using artificial urine to detect four commonly isolated UTI pathogens-namely, Escherichia coli, Proteus mirabilis, Staphylococcus aureus, and Candida albicans. Besides detection, these assays could also provide information regarding pathogen concentration/level, helping guide treatment decisions. These technologies were able to detect microbes associated with UTI at less than 105 CFU/mL, which is usually the lower cut-off limit for a positive UTI diagnosis. Among the 29 positive UTI samples yielding 105-106 CFU/mL pathogen concentrations, a total of 29 urine specimens were correctly detected by TuBETUr as UTI-positive based on an 1119 s detection window. Similarly, the rapid CUBET method was able to discriminate UTIs from normal samples with high confidence (p ≤ 0.0001), using single-pot conditions and cell phone-based monitoring. These technologies could potentially address the need for point-of-care UTI detection while reducing the possibility of antibiotic resistance associated with misdiagnosed cases of urinary tract infections, especially in low-resource environments.

Development and Assessment of a Simulator for in Vivo Confocal Microscopy in Fungal and Acanthamoeba Keratitis.

BACKGROUND AND PURPOSE: In vivo confocal microscopy (IVCM) is a non-invasive imaging technique that allows morphological analysis as a diagnostic approach of the cornea in real time, thus providing a suspected diagnosis of fungal or amoebic keratitis immediately, whereas culture or PCR require several days or even weeks. Since these infections are rare, it is difficult for ophthalmologists to gain the experience necessary to differentiate infection from normal findings or artefacts. The purpose of this project was to establish a simulator, on which physicians could practice as well as acquiring a database of IVCM images of fungal or amoebic keratitis and respective analyses. PATIENTS AND METHODS: An IVCM simulator was set up with cadaver human corneas, infected with either acanthamoeba, candida or aspergillus. Twenty-one ophthalmologists were trained in IVC microscopy first in a Dry Lab, then practically on the simulator. For evaluation, the participants were asked to fill out a standardized questionnaire, with a pre- and post-course self-assessment. RESULTS: The self-assessed theoretical and practical skills in differentiating infectious from non-infectious keratitis in IVCM significantly increased (p = 0.0001, p = 0.0002, respectively). The barrier to use this technique decreased (p = 0.0474). CONCLUSION: A very simple protocol based on a model of ex vivo corneal mycotic and amoebic infections can be used to train novices in the structured approach and diagnostic use of IVCM for corneal infections.