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.

Assessment of antifungal resistance and associated molecular mechanism in Candida albicans isolates from different cohorts of patients in North Indian state of Haryana.

The present study examines the trend in distribution of Candida species and their antifungal resistance patterns in hospitals across Haryana, a North Indian state with poorly addressed epidemiology of fungal infections. In our collection of 228 Candida isolates, Candida albicans dominated in both high vaginal swab (HVS) and urine samples while Candida glabrata and Candida tropicalis were the second-highest non-albicans Candida species (NAC), respectively. Of note, in blood samples, C. tropicalis and C. albicans were present in equal numbers. All 228 isolates were subjected to antifungal susceptibility tests, whereby 51% of C. albicans recovered from HVS samples displayed fluconazole resistance. To understand its mechanistic basis, expression profiling of efflux pump genes CDR1, CDR2, MDR1 and azole drug target, ERG11 was performed in 20 randomly selected resistant isolates, wherein many isolates elicited higher expression. Further, ERG11 gene sequencing suggested that most of the isolates harbored mutations, which are not reported with azole resistance. However, one isolate, RPCA9 (MIC 64 µg/mL) harbored triple mutation (Y132C, F145L, A114V), wherein Y132 and F145 sites were previously implicated in azole resistance. Interestingly, one isolate, (RPCA61) having MIC > 128 µg/mL harbored a novel mutation, G129R. Of note, HVS isolates RPCA 21, RPCA 22, and RPCA 44 (MICs 64 to > 128 µg/mL) did not show any change in alteration in ERG11 or overexpression of efflux pump genes. Together, this study presents a first report of Candida infections in selected hospitals of Haryana State.

Oral health and Candida carriage in socioeconomically disadvantaged US pregnant women.

BACKGROUND: Despite the well-documented associations between poor maternal oral health and increased risk for adverse birth outcomes and dental caries in children after birth, prenatal oral health care is under-utilized, especially among the underserved population. In addition, oral Candida has recently been suggested as a potential culprit for children's dental caries, with evident maternal contributions. Therefore, this study aimed to obtain epidemiological data on the oral health and oral Candida carriage in a cohort of underserved US pregnant women, and reveal factors associated with their oral Candida carriage. METHODS: Demographic-medical-oral hygiene practice data were collected. Comprehensive oral examination was conducted. Caries status and plaque index were recorded. Oral samples (saliva, plaque and swab) were processed to identify Candida species and Streptococcus mutans by culturing-dependent and -independent methods. Multiple logistic regression analyses were used to identify factors associated with oral Candida carriage and caries severity. RESULTS: Eighty-two socioeconomically disadvantaged women (48 pregnant and 34 non-pregnant) were enrolled. More pregnant women (79.1%) had > = 1 untreated decayed tooth when compared to their non-pregnant counterparts (47.1%) (p = 0.01). The average number of decayed teeth in pregnant and non-pregnant women was 3.9 and 3.1 (p > 0.05). Caries severity was positively associated with race (African American vs. white), plaque index and salivary Candida albicans level. C. albicans was the most predominant/abundant Candida strain, with cheek and tonsil as the most common colonized sites. The detection of C. albicans was 56%/56% in saliva and 40%/47% in plaque of the pregnant and non-pregnant groups, respectively. Study women's oral Candida carriage is positively associated with hypertension [p = 0.03, odds ratio = 14.47(1.28, 163.51)], decayed teeth number [p = 0.04, odds ratio = 1.31 (1.01,1.69)] and salivary S. mutans level [p = 0.03, odds ratio = 4.80 (1.18-19.43)]. CONCLUSIONS: Socioeconomically disadvantaged US women are in need of improved prenatal oral health, a large proportion of them have untreated decayed teeth and high carriage of oral Candida. Due to the observed significant association between the decayed teeth number and oral Candida carriage, providing oral health care during pregnancy (including limiting decayed teeth) will not only improve women's oral health, but also present as a promising approach to reduce oral Candida carriage in women.

Population pharmacokinetics of fluconazole in liver transplantation: implications for target attainment for infections with Candida albicans and non-albicans spp.

OBJECTIVES: The study aims to assess the population pharmacokinetics of fluconazole and the adequacy of current dosages and breakpoints against Candida albicans and non-albicans spp. in liver transplant (LT) patients. PATIENTS AND METHODS: Patients initiated i.v. fluconazole within 1 month from liver transplantation (LTx) for prevention or treatment of Candida spp. infections. Multiple assessments of trough and peak plasma concentrations of fluconazole were undertaken in each patient by means of therapeutic drug monitoring. Monte Carlo simulations were performed to define the probability of target attainment (PTA) with a loading dose (LD) of 400, 600, and 800 mg at day 1, 7, 14, and 28 from LTx, followed by a maintenance dose (MD) of 100, 200, and 300 mg daily of the pharmacokinetic/pharmacodynamic target of AUC24h/MIC ratio ≥ 55.2. RESULTS: Nineteen patients were recruited. A two-compartment model with first-order intravenous input and first-order elimination was developed. Patient's age and time elapsed from LTx were the covariates included in the final model. At an MIC of 2 mg/L, a LD of 600 mg was required for optimal PTAs between days 1 and 20 from LTx, while 400 mg was sufficient from days 21 on. A MD of 200 mg was required for patients aged 40-49 years old, while a dose of 100 mg was sufficient for patients aged ≥ 50 years. CONCLUSIONS: Fluconazole dosages of 100-200 mg daily may ensure optimal PTA against C. albicans, C. parapsilosis, and C. tropicalis. Higher dosages are required against C. glabrata. Estimated creatinine clearance is not a reliable predictor of fluconazole clearance in LT patients.

Adherence to the Mediterranean diet is associated with the gut microbiota pattern and gastrointestinal characteristics in an adult population.

This study aimed to explore the potential associations of adherence to the Mediterranean diet with gut microbiota characteristics and gastrointestinal symptomatology in an adult population. Other long-term dietary habits (e.g. consumption of snacks and junk food or stimulant intake) were also evaluated in terms of the gut microbiota profile. Participants (n 120) underwent anthropometric, dietary, physical activity and lifestyle evaluation. Adherence to the Mediterranean diet was assessed using a Mediterranean diet score, the MedDietScore, and subjects were classified into three tertiles according to individual adherence scoring. Gut microbiota composition was determined using quantitative PCR and plate-count techniques, and faecal SCFA were analysed using GC. Gastrointestinal symptoms were also evaluated. Participants with a high adherence to the Mediterranean diet had lower Escherichia coli counts (P=0·022), a higher bifidobacteria:E. coli ratio (P=0·025), increased levels and prevalence of Candida albicans (P=0·039 and P=0·050, respectively), greater molar ratio of acetate (P=0·009), higher defaecation frequency (P=0·028) and a more pronounced gastrointestinal symptomatology compared with those reporting low adherence. A lower molar ratio of valerate was also observed in the case of high adherence to the Mediterranean diet compared with the other two tertiles (P for trend=0·005). Positive correlations of MedDietScore with gastrointestinal symptoms, faecal moisture, total bacteria, bifidobacteria:E. coli ratio, relative share of Bacteroides, C. albicans and total SCFA, as well as negative associations with cultivable E. coli levels and valerate were indicated. Fast food consumption was characterised by suppressed representation of lactobacilli and butyrate-producing bacteria. In conclusion, our findings support a link between adherence to the Mediterranean diet and gut microbiota characteristics.

Comparative assessment of selected intraoral microorganisms ­ potential factors for peri-surgical management complications

In this research, a comparative analysis of results of investigations involving different human populations, in terms of a relation between the oral cavity health and the species composition of mouth microbiota is reported. The purpose of this analysis was to identify and assess microorganisms that could cause health complications in patients with neoplasm requiring dental problem-related surgical management. The patients with the oral cancer surgically treated and those without neoplasm were assessed for their oral health: status of teeth, gingiva, periodontium, and occurrence of inflammatory processes. From each patient, microorganisms isolated of periodontium, dental plaque, and dental pocket swabs were identified in wet and stained microscopic preparations; standard microbiology in vitro techniques were also applied to determine the fungal and bacterial strains. The comparative analysis of results of direct microscopic examinations and in vitro cultures assessment indicated significant differences in prevalences of fungi, parasitic oral protozoans and bacteriae in particular patient's groups. Yeast-like fungi belonging to Candida genus, mostly of C. albicans group, were identified in 93.75% patients with the oral cancer, while in 25% of individuals assessed without neoplasm. E. gingivalis amoebae were only found in 12.5% patients with the serious disease; no trichomonads were detected in all patients analyzed. Among bacteria species, potentially pathogenic Enterobacteriaceae were found in the patients with oral cancer. The pronounced shift in the microbiota species composition in the patients who needed prolonged treatment due to oral cavity cancer, compared to other generally healthy persons has been showed in this analysis.

Optimization of voriconazole dosage regimen to improve the efficacy in patients with invasive fungal disease by pharmacokinetic/pharmacodynamic analysis.

Invasive fungal disease (IFD) is a significant cause of morbidity and mortality in hospitalized patients. To maximize the efficacy of voriconazole treatment, the study established the relationship between voriconazole pharmacokinetic/pharmacodynamic (PK/PD) and probability of response and optimized voriconazole dosage regimen in patients with IFD based on Monte Carlo simulation. Forty-four patients proven with IFD were involved in this study. Among them, the overall cure rate was 75% (33/44) and there was a significant difference between Cmin /MIC values in patients with lack of response (n = 11) and those with successful response (n = 33) (mean value: 1.91 vs. 11.33; P < 0.05). Logistic regression model showed a high correlation between voriconazole Cmin /MIC ratio and clinical response (P = 0.044, OR = 1.349). According to Monte Carlo simulation results under different voriconazole dosing regimens, we could draw a conclusion that 200 mg voriconazole administered intravenously or orally twice daily for Candida infections and 300 mg administered orally or with 200 mg administered intravenously twice daily for Aspergillus infections were rational, which could achieve a value of the cumulative fraction of response >90%. This study built the relationship between voriconazole PK/PD and clinical response and obtained the reasonable empirical dosage regimen, which can be used to customize individual dosage regimen and improve the efficacy of voriconazole treatment.

Adenosine tri-phosphate (ATP)-based cleaning monitoring in health care: how rapidly does environmental ATP deteriorate?

BACKGROUND: Ensuring cleaning compliance of housekeeping staff is critical to ensure adequate application of surface disinfectants. Adenosine triphosphate (ATP) testing has been recommended as a way to monitor cleaning compliance; however, little is known about the stability of ATP on environmental surfaces. AIM: To assess the stability of ATP from various sources to determine if it is stable for sufficient time to be an effective means of assessing environmental cleaning and disinfection in health care. METHODS: Purified ATP, ATP derived from ATS-T (blood-based test soil) and ATP derived from 10(7) colony-forming units/site of micro-organisms (Pseudomonas aeruginosa, Enterococcus faecalis, Candida albicans) were evaluated in liquid suspension and dried on to surfaces to assess stability over 29 days. Cleaners and disinfectants were sprayed on to surface-dried material with no wiping to determine their effect on microbial viability and ATP stability. FINDINGS: Surface-dried P. aeruginosa, E. faecalis and C. albicans retained 65-96% of their original ATP level on Day 29, despite reduced or no viability. Surface-dried ATS-T had 100% and 3% of its original ATP on Days 4 and 29, respectively. Deterioration of the ATP signal was most pronounced for suspensions. Purified ATP was stable over 29 days in suspension or dried on to a surface. CONCLUSIONS: ATP residuals from organic material and micro-organisms (dead or alive) are stable when dried on to surfaces. In the absence of cleaning and disinfection, the relative light unit signal will not deteriorate rapidly, making ATP a good marker to monitor cleaning.

Antibiotic overuse as a risk factor for candidemia in an Indian pediatric ICU.

OBJECTIVE: To identify risk factors and mycological characteristics of candidemia in Pediatric ICU of a tertiary-care hospital. METHODS: Patients were screened for candidemia by blood culture. Recovered isolates were speciated and subjected to antifungal susceptibility testing. For every candidemic patient, three controls were matched for age, underlying diagnosis and period of hospitalization. Premature neonates were also matched for birth-weight. Proportion of cases and controls on specific antibiotics or indwelling devices was compared using Chi-square test, while unpaired t-test was used for comparing the number of antibiotics used and the number of days of antibiotic administration. Concordance between susceptibility testing methods was evaluated using Chi-square test. RESULTS: Significantly wider spectrum of antibiotic coverage was observed among the 28 candidemic patients. While every patient received antibiotic against enteric gram-negative bacilli, antibiotic usage for additional groups of microorganisms was significantly higher among cases. Association of candidemia with increasing use of indwelling devices was also observed. Endogenous colonization was higher in candidemic infants. Candida albicans was the commonest species (n = 18), followed by C. tropicalis (n = 7). Fluconazole and ketoconazole resistance was observed in 10.7 % cases. CONCLUSIONS: This information on pediatric candidemia could be used to devise locally-tailored strategies for identifying at-risk patients, underline the importance of routine antifungal susceptibility testing and formulate appropriate guidelines for management.

Assessment of Candida species colonization and denture-related stomatitis in bar- and locator-retained overdentures.

The aim of this study was to assess the prevalence of denture-related stomatitis (DRS) in different attachment-retained overdenture wearers and its association with particular colonizing Candida species. Thirty-seven edentulous patients with implant-supported maxillary or mandibular overdentures were enrolled. A full clinical history was obtained, including details of patients' oral hygiene practices and the levels of erythema based on Newton's classification scale. Swabs were taken from the palate and investigated mycologically to identify the yeast colonies. Quantitative and qualitative microbiological assessments were performed, which included recording the total numbers of colonies (cfu), their color, and their morphological characteristics. Significant differences were found in cfu values between the attachment and inner surfaces of locator- and bar-retained overdentures (P < .05). Candida albicans was the most common species in both evaluations, being isolated from 81.3% of bar-retained overdentures and 38.1% of locator-retained overdentures. DRS developed in all patients using bar-retained overdentures but in only 71.4% of those using locator-retained overdentures. No statistically significant relationship was found between bar and locator attachments according to smoking habit, overnight removal, or plaque and gingival indices (P > .05).

A microbiological assessment of the oral hygiene of 24-72-month-old kindergarten children and disinfection of their toothbrushes.

BACKGROUND: The objective of this study is to assess the index of decayed, missing and filled teeth (DMF-T), habit of brushing teeth, and the microbiological agents accumulating on the children's toothbrushes for 4 weeks and response of these agents to disinfection via a chlorhexidine solution, then compare those results with the education and income levels of the children's parents. METHOD: Included in the study were 187 children (96 in the control group and 91 in the experiment group - chlorhexidine) chosen randomly from 600 kindergarten children whose ages ranged from 24 months to 72 months. The children selected had not taken any antibiotics, antimicotics for three months and dental treatments during this trial. The distribution of these children to the groups was also done randomly. After performing a survey for the education, occupation, and income status of the parents, the children were examined and the number of decayed teeth was recorded. The children were given toothbrushes, toothpaste (with fluroide), and the solutions (including distilled water and chlorhexidine) for four weeks under the condition that toothbrushes were returned at the end of each week. The 14 different microbiological agents observed as a result of the assessment of the samples taken in the first week were also included in the assessments of the samples taken over the four-week period. RESULTS: The decrease in the DMF-T index was found to be meaningful according to the differences in education, income, and occupation status of the parents. Of all the samples taken from the toothbrushes, the bacteria with the greatest rate of reproduction included Streptococcus mutans, Escherichia Coli, Pseudomonas aeuroginosa, Enterococcus spp, Staphylococcus epidermidis and Candida albicans. Except for Candida albicans, the other microorganisms taken as samples from the toothbrushes reproduced less overall. In the group using the solution with chlorhexidine, a meaningful decrease in bacterial reproduction was discovered compared to the control group. CONCLUSION: The findings of this study show that the education, occupation, and socioeconomic situations of the parents should be considered when discussing children's oral and dental health. Moreover, the study shows that disinfection of toothbrushes in order to prevent reinfection and contamination oral flora with the bacteria again is important in terms of preventive medicine and family-children health.

Evaluation of the Candida albicans removal and mechanical properties of denture acrylics cleaned by a low-cost powered toothbrush.

PURPOSE: To investigate the effects of using a low-cost powered toothbrush for cleaning on dental prostheses made of heat polymerized poly(methyl methacrylate), PMMA. METHODS: Heat cured PMMA specimens beam with the dimensions of 45.0 mm×6.5 mm×4.5mm were fabricated. The specimens were kept in water storage at 37 °C constant temperature for 0, 1, 7, 15, 30 and 60 days and randomly assigned for testing or control. Test specimens underwent brushing by using a powered toothbrush at an applied force of 2.00 N for 22 min with water as medium. Surface roughness measurement (Ra), flexural strength and efficacy of brushing to remove coated Candida albicans biofilm were investigated. RESULTS: The results of the mean surface roughness value and the flexural strength were analysed by using two-way ANOVA and Tukey post hoc test at 5% significance level. In general, the specimens showed no significant changes in flexural strength after brushing. However, the flexural strength and the surface roughness value were significantly lower in specimens group after 7 days in water storage compared to the control. SEM micrographs of post-brushed specimens revealed satisfactory removal of C. albicans biofilm. CONCLUSION: A low-cost powered toothbrush together with a liquid medium successfully removed C. albicans biofilm on dental acrylic PMMA-based prostheses, without compromising the mechanical properties.

Assessment of rose bengal versus riboflavin photodynamic therapy for inhibition of fungal keratitis isolates.

PURPOSE: To compare the in vitro effect of rose bengal and riboflavin as photosensitizing agents for photodynamic therapy (PDT) on fungal isolates that are common causes of fungal keratitis. DESIGN: Experimental study. METHODS: Three isolates (Fusarium solani, Aspergillus fumigatus, Candida albicans) recovered from patients with confirmed fungal keratitis were used in the experiments. Isolates were grown on Sabouraud-Dextrose agar, swabbed, and prepared in suspension, and 1 mL aliquots were inoculated onto test plates in triplicate. Test plates were separated into 5 groups: Group 1, no treatment; Group 2, 0.1% rose bengal alone; Group 3, 518 nm irradiation alone; Group 4, riboflavin PDT (riboflavin + 375 nm irradiation); and Group 5, rose bengal PDT (rose bengal + 518 nm irradiation). Irradiation was performed over a circular area using either a green light-emitting diode (LED) array (peak wavelength: 518 nm) or an ultraviolet-A LED array (peak wavelength: 375 nm). Test plates were irradiated with an energy density of 5.4 J/cm(2). Later, plates were placed in a 30 C incubator and observed for growth. RESULTS: Rose bengal-mediated PDT successfully inhibited the growth of all 3 fungal isolates in the irradiated area. All other groups exhibited unrestricted growth throughout the plate. CONCLUSIONS: Rose bengal-mediated PDT successfully inhibited the growth of 3 types of fungi. No other experimental groups, including riboflavin-mediated PDT, had any inhibitory effect on the isolates. The results might be useful for the treatment of patients suffering from corneal infection.

Antifungal activity of the lipopeptides produced by Bacillus amyloliquefaciens anti-CA against Candida albicans isolated from clinic.

The bacterium Bacillus amyloliquefaciens anti-CA isolated from mangrove system was found to be able to actively kill Candida albicans isolated from clinic. The bacterial strain anti-CA could produce high level of bioactive substance, amylase and protease in the cheap medium containing 2.0 % soybean meal, 2.0 % wheat flour, pH 6.5 within 26 h. After purification, the main bioactive substance was confirmed to be a cyclic lipopeptide containing a heptapeptide, L-Asp→L-Leu→L-Leu→L-Val→L-Val→L-Glu→L-Leu and a 3-OH fatty acid (15 carbons). In addition to C. albicans, the purified lipopeptide can also kill many yeast strains including Metschnikowia bicuspidata, Candida tropicalis, Yarrowia lipolytica and Saccharomyces cerevisiae. After treated by the purified lipopeptide, both the whole cells and protoplasts of C. albicans were destroyed.

Analysis of strain relatedness using high resolution melting in a case of recurrent candiduria.

BACKGROUND: Several genotyping protocols have been described to study Candida albicans strains with different sensitivity values. In this study we have analyzed the genetic relatedness and the antifungal susceptibility of several Candida albicans strains isolated from a patient who from suffered recurrent candiduria for a period of five years. Strains were genotyped using Microsatellite Length Polymorphism (MLP) with three microsatellite markers (HIS 3, EF 3 and CDC 3), and a new method based on high resolution melting (HRM) was developed to analyze the microsatellite region. This method was compared with the conventional technique that uses capillary electrophoresis. RESULTS: MICs of the isolates showed the existence of fluconazole susceptible and resistant strains. An inter-colony test using single concentration (8 and 16 mg/l) of fluconazole revealed the coexistence of both fluconazole susceptible and resistant strains. Both genotyping analysis methods showed that all the patient's isolates had a clonal origin. HRM analysis method developed was able to accurately establish strain relatedness and presented a discriminatory power of 0.77. CONCLUSIONS: Although HRM analysis method presented a lower discriminatory power compared to methods based on capillary electrophoresis, it provided a more cost-effective and suitable alternative for genotyping C. albicans in a clinical laboratory.

Field assessment of the mid winter mass kills of trophic fishes at Mariotteya stream, Egypt: chemical and biological pollution synergistic model.

Pathogenic Candida albicans was isolated from water and fish samples collected during an emergent event of mass mortalities among the juvenile Nile tilapia (Oreochromis niloticus), Sharp toothed catfish (Clarias gariepinus) along the stream of Mariotteya drainage. Investigations indicated that fish mortalities were confined to the area of Shubramant and Aboul Noumros (North to Sakara 7 drainage). C. albicans was isolated from the lesions associated with multiple skin ulcers in both Nile tilapia juveniles and Sharp toothed catfish. Assessment of the field and laboratory data has indicated that Mariotteya environmental disaster was a multifactorial problem. The fish mass kills were initially flared up through the dumping of the improperly treated nasty organic and inorganic chemicals from Elhawamdia sugar factory and municipal sewage. The physical stagnation of the stream, high levels of ammonia, phenol and polycyclic aromatic hydrocarbons (PAHs) and low levels of dissolved oxygen (DO) were all incriminated as the initial stimulus behind biological invasion of pathogenic bacteria (Pseudomonas fluorescence) and yeast (C. albicans). Pathologically, fishes were dying from both respiratory and osmoregulatory failure induced by the severe damage of both gills and skin. It has been implied that such environmental pollutants have direct damaging effects on gills, skin and fins with consequent suppression of the skin's natural innate components. The adversely confronted immunological barriers were further exacerbated by the possible synergistic interactions of P. fluorescence dermotropic toxins followed by the secondary invasion of the pathogenic C. albicans.