Concentrated Urine Samples Research Articles

B-057 Evaluating the Use of Unconcentrated Urine for Protein Electrophoresis and Immunofixation: An Alternative Approach for Efficient Analysis

Abstract Background Protein electrophoresis is a widely employed technique for separating and analyzing proteins based on their charge and size. Traditionally, concentrated urine samples have been utilized to enhance protein detection sensitivity. However, this study explores the feasibility of using unconcentrated urine as an alternative sample type for protein electrophoresis (UEL) and immunofixation (UIF) to streamline the analysis process and reduce costs and turnaround time in clinical laboratories. It offers a more efficient and resource-friendly alternative. Methods Both UEL and UIF were performed on routinely ordered 80 urine specimens (random or 24 Hr.) for patient management and performed by both concentrated (X40) by Helena and unconcentrated using Sebia (HYDRAGEL 9IF gels using HYDRAGEL BJ dynamic mask procedure. The electrophoretic patterns obtained from unconcentrated urine were compared with those from concentrated urine. Various electrophoretic parameters, including resolution, sensitivity, and reproducibility, were evaluated to determine the reliability of unconcentrated urine in protein electrophoresis. In addition, retrospective analysis was performed before and after implementation on all urine specimens ordered to analyze if there were any differences in the monoclonal band distributions in the population. Results All the protein concentrations were correlated well with 95-100% agreement between the methods (Figure 1) clear separation of protein fractions. Serial dilution studies of monoclonal band analyses showed that these bands can be detected at 1.25 mg/dL using an unconcentrated method. Retrospective analysis of patient data before and after the implementation of the new method shows no differences in the identification of monoclonal proteins (Helena [69 of 872 total patients tested (8.0%)] vs. Sebia [90/800 (11.0%)]. Conclusions These results indicate that unconcentrated urine can be successfully utilized for UEL and UIF with comparable outcomes to traditional concentrated samples. Utilizing unconcentrated urine simplifies the sample preparation process and presents a promising alternative.

Acoustofluidic-based microscopic examination for automated and point-of-care urinalysis.

Urinalysis is a heavily used diagnostic test in clinical laboratories; however, it is chronically held back by urine sediment microscopic examination. Current instruments are bulky and expensive to be widely adopted, making microscopic examination a procedure that still relies on manual operations and requires large time and labor costs. To improve the efficacy and automation of urinalysis, this study develops an acoustofluidic-based microscopic examination system. The system utilizes the combination of acoustofluidic manipulation and a passive hydrodynamic mechanism, and thus achieves a high throughput (1000 μL min-1) and a high concentration factor (95.2 ± 2.1 fold) simultaneously, fulfilling the demands for urine examination. The concentrated urine sample is automatically dispensed into a hemocytometer chamber and the images are then analyzed using a machine learning algorithm. The whole process is completed within 3 minutes with detection accuracies of erythrocytes and leukocytes of 94.6 ± 3.5% and 95.1 ± 1.8%, respectively. The examination outcome of urine samples from 50 volunteers by this device shows a correlation coefficient of 0.96 compared to manual microscopic examination. Our system offers a promising tool for automated urine microscopic examination, thus it has potential to save a large amount of time and labor in clinical laboratories, as well as to promote point-of-care urine testing applications in and beyond hospitals.

Is Concentrated Urine PCR Test Valuable to Detect Genital HPV Infection [ID 2683587

INTRODUCTION: Human papillomavirus (HPV) is the leading cause of cervical cancer and early detection is crucial for effective prevention. The standard HPV detection method, cervical smear PCR, entails discomfort and deters many women. Therefore, it is challenging to screen large populations for HPV from cervical smear samples. In this study, we investigated the sensitivity of detecting HPV by PCR in concentrated urine samples. METHODS: Urine and cervical smear samples were collected from 126 patients (ages 20–35) at Acibadem Altunizade Hospital. Urine samples were concentrated by a tool (MyMagiCon). The presence of HPV DNA in cervical smear and urine samples was determined by PCR. RESULTS: Among 126 patients, HPV was detected in 57 either in at least one of the sample types (cervical smear, urine, and concentrated urine). Among all HPV-positive patients, HPV was identified in 87.7% of swab samples, 86% of concentrated urine samples, and only 73.7% of unconcentrated urine samples. The χ2 test comparing cervical smear and concentrated urine samples yielded a Kappa value of 0.78 (95% confidence), indicating strong statistical correlation within this study. Additionally, PCR from concentrated urine samples, identified six HPV-positive samples that were HPV negative in cervical smear samples of the same patients. CONCLUSION: It was possible to identify HPV in concentrated urine samples by PCR with a sensitivity almost equal to cervical smear sampling. Since obtaining urine is much easier than obtaining cervical smear samples, this may enable easy screening of large populations for HPV. This may have a big effect in the prevention of cervical cancer.

Detection of human papillomavirus in fresh and dried urine through an automated system for cervical cancer screening in low- and middle-income countries.

The majority of cervical cancer cases and associated deaths occur in low- and middle-income countries (LMICs), where sociocultural barriers, poor access to prevention and care, and technical and practical difficulties hinder screening coverage improvement. Using urine specimens for human papillomaviruses (HPV) molecular screening through automated testing platforms can help to overcome these problems. We evaluated the high-risk (HR) HPV detection performance of the Xpert® HPV test on GeneXpert® System (Cepheid), on fresh and dried urine (Dried Urine Spot [DUS]) samples as compared to an in-house polymerase chain reaction (PCR) genotyping assay. Forty-five concentrated urine samples collected from women with known cytological and HPV infection status, determined through in-house PCR and genotyping assays, were tested "as is" and as DUS with the Xpert® HPV test. This system detected HR-HPV in 86.4% of fresh and in 77.3% of dried urine samples collected from HPV+ women, correctly identifying HR-HPV infection in 100% of women with low- and high-grade lesions. High concordance (91.4%, k = 0.82) was found between PCR test and Xpert® HPV Test from urine. Urine-based Xpert® HPV test seems to be a suitable screening test for detection of HR-HPV infections associated with low- and high-grade lesions requiring follow-up monitoring or treatment. This methodology, relying on noninvasively collected samples and on available rapid testing platforms, could facilitate large, at-scale screening programs, particularly in LMICs and rural areas, thus reducing adverse outcomes of HPV infection and facilitating achievement of the WHO cervical cancer elimination goal.

Urine specific gravity, pyuria, and neutrophil gelatinase-associated lipocalin for identifying urinary tract infection in young children.

To determine whether urine neutrophil gelatinase-associated lipocalin (uNGAL) might be superior to pyuria for detecting urinary tract infection (UTI) regardless of urine specific gravity (SG) in young children. We conducted a retrospective analysis of children aged < 3 years who were evaluated for UTI with urinalysis, urine culture, and uNGAL measurements during a 5-year period. Sensitivity, specificity, likelihood ratios (LRs), predictive values (PVs), area under the curves (AUCs) of uNGAL cut-off levels, and various microscopic pyuria thresholds for detecting UTI were calculated for dilute (SG < 1.015) and concentrated urine (SG ≥ 1.015). Of 456 children included, 218 had UTI. The diagnostic value of urine white blood cell (WBC) concentration to define UTI changed with urine SG. For detecting UTI, uNGAL cut-off of 68.4 ng/mL had higher AUC values than pyuria ≥ 5 WBCs/high power field (HPF) for dilute and concentrated urine samples (both P < 0.05). Positive LR and PV and specificity of uNGAL were all greater than those of pyuria ≥ 5 WBCs/HPF regardless of urine SG, although the sensitivity of pyuria ≥ 5 WBCs/HPF was higher than that of uNGAL cut-off for dilute urine (93.8% vs. 83.5%) (P < 0.05). At uNGAL ≥ 68.4 ng/mL and ≥ 5 WBCs/HPF, posttest probabilities of UTI were 68.8% and 57.5% for dilute urine and 73.4% and 57.3% for concentrated urine, respectively. Urine SG can affect the diagnostic performance of pyuria for detecting UTI and uNGAL might be helpful for identifying UTI regardless of urine SG in young children. A higher resolution version of the Graphical abstract is available as Supplementary information.

Human Hydration Indices: Spot Urine Sample Reference Values for Urine Concentration Markers in Athletic Populations

Background: Reference values and confidence intervals for the hydration indices of a large athletic population are currently lacking. Methods: Urine indices were gathered from an athletic population (n = 189) based on spot-urine samples. Results: High urine concentration was associated with a low volume and short void duration. When stratifying the data, differences for urine volume were seen for race and ethnicity and for athletic affiliation (p &lt; 0.05), but no differences were found for urine concentration markers or volume for time of day of collection, thirst sensation, or age (p &gt; 0.05). When classifying urine samples for a low vs. a high urine concentration by scoring urine color (Uc), the athletic population reported a slightly lower accuracy (4–7%) compared to investigators (p &lt; 0.02). Subjects scored samples as lighter than the investigators, with a higher misclassification of the more concentrated urine samples. Conclusions: In this convenience sample of a predominantly young athletic population, urinary indices did not differ for subgroups within a large athletic population aside from some difference for race and ethnicity on urine volume. Although well-trained investigators reported better accuracy for Uc scoring, both athletes and investigators reported the highest accuracy for correctly classifying samples with a very low or a very high urine concentration.

Abstract B03: Urine cell-free DNA (cfDNA) concentration and stability test for future clinical use

Abstract Introduction: Urine was shown to be a liquid biopsy source with high sensitivity for cfDNA mutation detection in both early- and late-stage cancer patients, outperforming plasma cfDNA and tissue samples in some patients. In addition, obtaining a urine sample is completely noninvasive. Limitations of using urine samples in the clinical setting include their unknown nucleic acid stability and the large total volume of one deposit, and therefore the presence of highly diluted cfDNA. Urine volume concentration, cfDNA stability, cfDNA quantity, and detection of wild-type (wt) EGFR alleles were explored to determine the future suitability of urine cfDNA as a diagnostic or prognostic biosource. Methods: First-void urine samples, with a higher fraction of nucleic acids due to overnight concentration, were collected from 5 healthy donors and 5 cancer patients. Cell-free urine samples were concentrated using Amicon filter tubes to determine differences in cfDNA concentration. In addition, we tested the ability to detect wild-type EGFR exon 19 in cfDNA from concentrated and unconcentrated urine samples using a peptide nucleic acid (PNA) Taqman Assay. Hereafter, stability of urine cfDNA was explored using variable storage temperatures and times and addition of a urine cfDNA preservative directly after sample collection. cfDNA was extracted using the automated QIAsymphony system with the DSP virus/pathogen kit and quantified using the Qubit dsDNA HS Assay in all experiments. Results: Amicon filter-based concentration of urine samples from 15 mL to 1.2 mL yielded a two-fold increase in cfDNA concentration. Importantly, while wild-type EGFR could not be detected in first-void urine using a PNA Taqman Assay, detection was possible using concentrated urine samples. Therefore, in all following experiments, samples were concentrated unless specified otherwise. Stability tests indicated that storage temperature matters when samples are not processed within two hours, with nearly doubled cfDNA concentrations found in samples stored at four degrees for 26 hours, compared to room temperature (RT). Addition of a cfDNA preservative yielded a six-fold higher cfDNA concentration after 26 hours of storage at RT, compared to samples without a preservative added. For this reason, we tested the use of only cfDNA preservative without concentrating the sample and found a four-fold increase in cfDNA concentration when adding only preservative, versus a 15-fold increase when using both after sample storage at RT for 24 hours. Conclusion: Patient urine samples should be either concentrated and processed within two hours, or a urine cfDNA preservative should be added directly after sample collection to prevent cfDNA degradation. Depending on the downstream application, samples where preservative was added may be concentrated to enhance the cfDNA yield, for example, when downstream NGS analysis will be performed. Citation Format: Jillian W. P. Bracht, Niki Karachaliou, Jordi Berenguer, Manuel Fernandez-Bruno, Mónica Garzón, Ana Gimenez-Capitan, Clara Mayo-de-las-Casas, Miguel Angel Molina-Vila, Rafael Rosell. Urine cell-free DNA (cfDNA) concentration and stability test for future clinical use [abstract]. In: Proceedings of the AACR Special Conference on Advances in Liquid Biopsies; Jan 13-16, 2020; Miami, FL. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(11_Suppl):Abstract nr B03.

Inactivation kinetics modeling of Escherichia coli in concentrated urine for implementing predictive environmental microbiology in sanitation safety planning

Urine concentration (condensation) leads to the inactivation of pathogens in urine owing to a hyperosmotic environment. This study proposed an inactivation kinetic model of Escherichia coli (E. coli), a surrogate of human bacterial pathogens, in concentrated synthetic urine. The model parameters were obtained under an assumption that the inactivation rate of E. coli followed a binomial distribution, which made it possible to accurately simulate the time-course decay of E. coli in synthetic urine. The inactivation rate constant values obtained in concentrated urine samples, ammonium buffer solutions and carbonate buffer solutions indicated that the osmotic pressure was a relatively predominant cause for the inactivation of E. coli. The appropriate storage time was estimated using the approach of quantitative microbial risk assessment, which indicated that the 5-fold concentrated urine could be safely collected after 1-day storage when urea was hydrolyzed, whereas 91-hour storage was required for non-concentrated urine. The occupational risk was not negligible even with 6-month storage at 20 °C when urea was not hydrolyzed, which suggested that the urine storage styles should be clarified more minutely. The present study highlights the importance of “predictive environmental microbiology,” which deals with inactivation kinetic models of microorganisms under varied environmental conditions to fully implement the hazard analysis and critical control point (HACCP) approach for the safe use of human excreta in agriculture.

Getting Specific About Urine Specific Gravity

Research Article| February 01 2020 Getting Specific About Urine Specific Gravity AAP Grand Rounds (2020) 43 (2): 18. https://doi.org/10.1542/gr.43-2-18 Views Icon Views Article contents Figures & tables Video Audio Supplementary Data Peer Review Share Icon Share Facebook Twitter LinkedIn MailTo Tools Icon Tools Get Permissions Cite Icon Cite Search Site Citation Getting Specific About Urine Specific Gravity. AAP Grand Rounds February 2020; 43 (2): 18. https://doi.org/10.1542/gr.43-2-18 Download citation file: Ris (Zotero) Reference Manager EasyBib Bookends Mendeley Papers EndNote RefWorks BibTex toolbar search toolbar search search input Search input auto suggest filter your search All PublicationsAll JournalsAAP Grand RoundsPediatricsHospital PediatricsPediatrics In ReviewNeoReviewsAAP NewsAll AAP Sites Search Advanced Search Topics: urinalysis, urine specific gravity, urinary tract infections, nitrites Source: Shaikh N, Shope MF, Kurs-Lasky M. Urine specific gravity and the accuracy of urinalysis. Pediatrics. 2019; 144(5): e20190467; doi: https://doi.org/0.1542/peds.2019-0467Google Scholar Investigators from the University of Pittsburgh conducted a retrospective analysis to determine whether urine specific gravity (SG) affects the accuracy of other components of the urinalysis. Children <24 months of age with a visit to the study ED between 2007 and 2013 were eligible if they underwent bladder catheterization and had an automated urinalysis and urine culture performed within 3 hours of each other. Children with genitourinary anomalies were excluded. Investigators evaluated the accuracy of urinalysis components used for diagnosis of UTI by comparing the positive likelihood ratios (LRs) in dilute versus concentrated urine samples. Dilute urine samples were defined as those with an SG <1.015, and concentrated samples were defined as those with an SG ≥1.015. UTI was defined as ≥100,000 colony-forming units (CFUs) per milliliter of a uropathogen from specimens obtained via clean catch or ≥50,000 CFU/mL from specimens obtained via catheterization. Urinalysis components evaluated included leukocyte esterase (LE; a positive test defined as ≥1), nitrite, white blood cell count (WBC) per high-power field (hpf; a positive test defined as ≥5), WBC/mm3 (a positive test defined as ≥10), bacteria/hpf (a positive test defined as ≥ few), and bacteria on Gram-stain (a positive test defined as ≥ any). Investigators also used predictive models to calculate how many children would be overtreated or missed with and without SG being used in the interpretation of other urinalysis components. There were 10,078 children included in analysis. Of these, 39.4% (n=3,966) had concentrated urine, and, overall, 6.1% had a UTI (n=617). The positive LRs for LE and bacteria/hpf (ie, their ability to rule in UTI) were similar in dilute and concentrated urine. However, WBC/hpf and WBC/mm3 had lower LRs in concentrated (vs dilute) urine, and nitrite and bacteria on Gram stain had higher LRs. Despite this finding, there was no change in predictive models for the number of children overtreated or missed with or without inclusion of SG in combination with urinalysis components. The investigators conclude that SG influences the accuracy of some components of the urinalysis but that inclusion of SG in decision-making would result in little change to clinical practice. Dr Bechtel has disclosed no financial relationship relevant to this commentary. This commentary does not contain a discussion of an unapproved/investigative use of a commercial product/device. UTIs are common bacterial infections in the pediatric population.1 An uncontaminated sample of urine for urinalysis and culture is generally required before starting any presumptive antimicrobial treatment. The urinalysis serves as the necessary screening test to determine the need to start empiric antibiotics, especially if pyuria and bacteriuria are present.2 Despite the widespread use of urinalyses by clinicians to make the presumptive diagnosis of UTI, urine SG has not previously been incorporated into their interpretation. For example, the urine SG has not been considered... You do not currently have access to this content.

Urine Specific Gravity and the Accuracy of Urinalysis.

A recent study in young infants found that different cutoffs maximized the accuracy of the urine white blood cell count in dilute versus concentrated urine samples. We aimed to confirm this finding and to determine its impact on clinical care. We conducted a retrospective analysis of data gathered on consecutive children <24 months of age with visits to the emergency department during a 5-year period. We evaluated the accuracy of screening tests for urinary tract infection (UTI) in dilute and concentrated urine samples. We also calculated the number of children who would have been treated differently in a hypothetical cohort of 1000 children presenting with fever had urine specific gravity (SG) been taken into consideration. We included 10 078 children. The ability to rule in UTI (as measured by the positive likelihood ratio [LR]) was similar in dilute and concentrated urine for the leukocyte esterase test (11.76 vs 10.71, respectively). The positive LR for urine white blood cell count per high-powered field was higher in dilute urine (9.83 vs 6.12). In contrast, the positive LR for the nitrite test was lower in dilute urine (20.54 vs 47.44). Despite these differences, we found little change in the number of children treated with antibiotics in predictive models that took urine SG into consideration. Although we found that urine SG influences the accuracy of some components of the urinalysis, its inclusion in the decision-making process had negligible effect on the clinical care of children with UTI.

False rifampicin resistant results using Xpert MTB/RIF on urine samples in hospitalised HIV-infected patients.

BackgroundA small proportion of false rifampicin resistant results have previously been reported using GeneXpert MTB/RIF version G4 on sputum samples; however, this has not been investigated for urine samples in HIV-associated tuberculosis (TB).ObjectivesWe sought to determine the proportion of false rifampicin resistant results using Xpert MTB/RIF version G4 on urine samples among HIV-infected inpatients investigated for TB.MethodsHospitalised HIV-infected patients undergoing systematic TB testing from two cohorts in Cape Town, South Africa, were enrolled. All patients with ≥1 urine Xpert result available were included. Rifampicin resistant urine Xpert results were classified into three mutually exclusive groups: (1) true rifampicin resistance, (2) false rifampicin resistance or (3) unknown after review of available microbiologic and clinical data.ResultsOverall, 1171 patients were included, from whom a total of 1704 urine Xpert results were available on unconcentrated and/or concentrated urine samples. There were 416 samples positive for TB (24.4% [95% CI 22.4–26.5]), of which 43/413 (10.4% [95% CI 7.6–13.8]) were rifampicin resistant (after excluding three results that were falsely positive due to contamination). Of 43 rifampicin resistant Xpert results (among 40 patients), 30 were classified as true resistance, 11 as false resistance and 2 could not be classified. Excluding unclassifiable results, 30/41 results were confirmed as true-positive urine Xpert rifampicin resistance (positive predictive value: 73.2% [95% CI 57.1–85.8]).ConclusionUrine Xpert testing showed a high proportion of false rifampicin resistance results. Urine Xpert rifampicin resistant results should be interpreted cautiously and confirmed when possible.

Robust Markers of Coffee Consumption Identified Among the Volatile Organic Compounds in Human Urine.

The human volatilome has gained high interest for the discovery of potential biomarkers of diseases. However, knowledge about the diet as a crucial factor affecting the volatilome is scarce. Therefore, the search for disease biomarkers, as well as the potential use of volatiles as dietary markers is so far limited. The aim of this study is to investigate the association of the diet with the urinary volatilome with the special task to find potential markers of coffee consumption in 24 h urine samples from the Karlsruhe Metabolomics and Nutrition (KarMeN) study. Acidified urine samples are analyzed using an approach combining headspace solid phase microextraction (HS-SPME) sampling with untargeted GC×GC-MS. Overall, 138 reliably occurring volatiles are detected. To account for the unequally concentrated urine samples, results of six different commonly used normalization methods are compared. Statistical analysis evidences six potential markers of coffee consumption, the most promising being 3,4-dimethyl-2,5-furandione. A correlation analysis between the 24 h dietary recall data and the urinary volatilome reveals further promising associations. The human urinary volatilome is highly affected by the diet, enabling access to a high level of information about potential diet-related biomarkers. Therefore, it is a very promising source for further investigations on dietary markers.

Diagnostic accuracy, incremental yield and prognostic value of Determine TB-LAM for routine diagnostic testing for tuberculosis in HIV-infected patients requiring acute hospital admission in South Africa: a prospective cohort

BackgroundWe previously reported that one-third of HIV-positive adults requiring medical admission to a South African district hospital had laboratory-confirmed tuberculosis (TB) and that almost two-thirds of cases could be rapidly diagnosed using Xpert MTB/RIF-testing of concentrated urine samples obtained on the first day of admission. Implementation of urine-based, routine, point-of-care TB screening is an attractive intervention that might be facilitated by use of a simple, low-cost diagnostic tool, such as the Determine TB-LAM lateral-flow rapid test for HIV-associated TB.MethodsSputum, urine and blood samples were systematically obtained from unselected HIV-positive adults within 24 hours of admission to a South African township hospital. Additional clinical samples were obtained during hospitalization as clinically indicated. TB was defined by the detection of Mycobacterium tuberculosis in any sample using Xpert MTB/RIF or liquid culture. The diagnostic yield, accuracy and prognostic value of urine-lipoarabinomannan (LAM) testing were determined, but urine-LAM results did not inform treatment decisions.ResultsConsecutive HIV-positive adult acute medical admissions not already receiving TB treatment (n = 427) were enrolled regardless of clinical presentation or symptoms. TB was diagnosed in 139 patients (TB prevalence 32.6%; median CD4 count 80 cells/μL). In the first 24 hours of admission, sputum (spot and/or induced) samples were obtained from 37.0% of patients and urine samples from 99.5% of patients (P < 0.001). The diagnostic yields from these specimens were 19.4% (n = 27/139) for sputum-microscopy, 26.6% (n = 37/139) for sputum-Xpert, 38.1% (n = 53/139) for urine-LAM and 52.5% (n = 73/139) for sputum-Xpert/urine-LAM combined (P < 0.01). Corresponding yields among patients with CD4 counts <100 cells/μL were 18.9%, 24.3%, 55.4% and 63.5%, respectively (P < 0.01). The diagnostic yield of urine-LAM was unrelated to respiratory symptoms, and LAM assay specificity (using a grade-2 cut-off) was 98.9% (274/277; 95% confidence interval [CI] 96.9–99.8). Among TB cases, positive urine-LAM status was strongly associated with mortality at 90 days (adjusted hazard ratio 4.20; 95% CI 1.50–11.75).ConclusionsRoutine testing for TB in newly admitted HIV-positive adults using Determine TB-LAM to test urine provides major incremental diagnostic yield with very high specificity when used in combination with sputum testing and has important utility among those without respiratory TB symptoms and/or unable to produce sputum. The assay also rapidly identifies individuals with a poor prognosis.

Detection of Mycobacterial Lipoarabinomannan with A Monoclonal Antibody Qualitative ELISA in Urine of Tuberculous Meningitis Patients

BACKGROUND: Tuberculous meningitis is the most severe manifestation of tuberculosis. The diagnostic approach of tuberculous meningitis is difficult. Combination of clinical, laboratory and radiological criteria were used in diagnostic approach of tuberculous meningitis. Urinary mycobacterial lipoarabinomannan (LAM) antigen detection is a promising diagnostic tool. Detection of mycobacterial antigen in concentrated urine sample is predicted to improve the positivity rate of the qualitative enzyme-linked immunosorbent assay (ELISA) diagnostic tool. The purpose of this study is to examine the detection ability of a monoclonal antibody qualitative ELISA in concentrated and unconcentrated urine of tuberculous meningitis patients.METHODS: This research is a descriptive, crosssectionally designed. The study was conducted in the Clinical Pathology Department laboratory of Dr. Hasan Sadikin Hospital, in July-October 2014. A total of 27 patients diagnosed as tuberculous meningitis patients were included and the subjects were classified into possible and probable criteria according to consensus criteria. The subjects were classified as definite if the cerebrospinal fluid culture was positive for Mycobacterial tuberculosis growth. The subjects were examined for the presence of LAM in unconcentrated and concentrated urine with a monoclonal antibody qualitative ELISA method.RESULTS: Unconcentrated urinary LAM examination positivity was 0% while in concentrated urine was 14.8%. The positivity of concentrated urinary LAM were higher among the definite criteria group.CONCLUSION: Concentrating urine sample increase the positivity rate of urinary LAM detection with ELISA method as high as 14.8%. The urinary antigen detection is higher among the definite tuberculous meningitis patients.KEYWORDS: LAM, concentrated urine, tuberculous meningitis, qualitative ELISA

Microbiological diagnosis and molecular typing of Legionella strains during an outbreak of legionellosis in Southern Germany

An explosive outbreak of Legionnaires’ disease with 64 reported cases occurred in Ulm/Neu-Ulm in the South of Germany in December 2009/January 2010 caused by Legionella (L.) pneumophila serogroup 1, monoclonal (mAb) subtype Knoxville, sequence type (ST) 62. Here we present the clinical microbiological results from 51 patients who were diagnosed at the University hospital of Ulm, the results of the environmental investigations and of molecular typing of patients and environmental strains. All 50 patients from whom urine specimens were available were positive for L. pneumophila antigen when an enzyme-linked immunosorbent assay (EIA) was used following concentration of those urine samples that tested initially negative. The sensitivity of the BinaxNow rapid immunographic assay (ICA), after 15min reading and after 60min reading were 70% and 84%, respectively. Direct typing confirmed the monoclonal subtype Knoxville in 5 out of 8 concentrated urine samples. Real time PCR testing of respiratory tract specimens for L. pneumophila was positive in 15 out of 25 (60%) patients. Direct nested sequence based typing (nSBT) in some of these samples allowed partial confirmation of ST62. L. pneumophila serogroup 1, monoclonal subtype Knoxville ST62, defined as the epidemic strain was isolated from 8 out of 31 outbreak patients (26%) and from one cooling tower confirming it as the most likely source of the outbreak. While rapid detection of Legionella antigenuria was crucial for the recognition and management of the outbreak, culture and molecular typing of the strains from patients and environmental specimens was the clue for the rapid identification of the source of infection.

Evaluation of the BinaxNOW® Streptococcus pneumoniae antigen test on fresh, frozen and concentrated urine samples in elderly patients with and without community-acquired pneumonia

We evaluated the BinaxNOW® urine antigen test in elderly. For fresh un-concentrated urine samples, the sensitivity for pneumococcal pneumonia was 63% and specificity 97%. After freezing and concentration, the results comparable to positive control line in intensity at 60min gave high sensitivity (81%) with no loss in specificity (96%).

Urinary protein-to-creatinine ratio in pregnant women after dipstick testing: prospective observational study.

BackgroundThe dipstick test is widely used as a primary screening test for detection of significant proteinuria in pregnancy (SPIP). However, it often shows a false positive test result. This study was performed to determine which pregnant women should be recommended to undergo determination of urinary protein-to-creatinine ratio (mg/mg, P/Cr test) after dipstick test for confirmation of SPIP.MethodsThis was a multicenter, prospective, and observational study of 2212 urine specimens from 1033 pregnant women who underwent simultaneous dipstick and P/Cr tests in the same spot urine samples at least once. SPIP was defined as P/Cr > 0.27. Preeclampsia was diagnosed in women with both hypertension and SPIP.ResultsPreeclampsia, hypertension alone, and SPIP alone developed in 202 (20 %), 73 (7.1 %), and 120 (12 %) women, respectively. Creatinine concentration [Cr] varied greatly, ranging from 8.1 to 831 mg/dL in the 2212 urine samples. Rate of positive dipstick test results increased with increasing [Cr], while SPIP prevalence rate was lower in urine samples with higher [Cr], yielding higher false positive rates in samples with higher [Cr]. Postpartum urine samples had significantly lower [Cr] compared to those obtained antepartum (60 [8.7–297] vs. 100 [10–401] mg/dL, respectively). At the first P/Cr test among women with similar dipstick test results, the risk of having SPIP was consistently and significantly higher for hypertensive women than for normotensive women at any dipstick test result: 18 % (14/77) vs. 3.2 % (8/251), 47 % (26/55) vs. 8.7 % (37/425), 91 % (82/90) vs. 59 % (44/75) for negative/equivocal, 1+, and ≥ 2+ test results, respectively. The risk of SPIP was 16 % (9/55) for normotensive women when two successive antenatal urine samples showed a dipstick test result of 1 + .ConclusionsFor prediction of SPIP, the dipstick test was more likely to show a false positive result in concentrated urine samples with higher [Cr]. Hypertensive women with ≥ 1+ as well as normotensive women with ≥ 2+ on dipstick test should be advised to undergo the P/Cr test.

Urine osmolality in the US population: Implications for environmental biomonitoring

BackgroundFor many environmental chemicals, concentrations in spot urine samples are considered valid surrogates of exposure and internal dose. To correct for urine dilution, spot urine concentrations are commonly adjusted for urinary creatinine. There are, however, several concerns about the use of urine creatinine. While urine osmolality is an attractive alternative; its characteristics and determinants in the general population remain unknown. Our objective was to describe the determinants of urine osmolality and to contrast the difference between osmolality and creatinine in urine. MethodsFrom the National Health and Nutrition Examination Survey (NHANES) (2009–2010), 10,769 participants aged 16 years or older with measured urine osmolality and creatinine were used in the analysis. Very dilute and very concentrated urine was defined as urine creatinine lower than 0.3g/l and higher than 3g/l, respectively. Linear and logistic regression analyses were performed to investigate the associations of interest. ResultsUrine osmolality and creatinine were highly correlated (Pearson correlation coefficient=0.75) and their respective median values were 648mOsm/kg and 1.07g/l. The prevalence of very dilute and very concentrated urine samples was 8.1% and 3.1%, respectively. Factors associated in the same direction with both urine osmolality and urine creatinine included age, sex, race, body mass index (BMI), hypertension, water intake, and blood osmolality. The magnitude of associations expressed as percent change was significantly stronger with creatinine than osmolality. Compared to urine creatinine, urine osmolality did not vary by diabetes status but was affected by daily total protein intake. Participants with chronic kidney disease (CKD) had significantly higher urine creatinine concentrations but lower urine osmolality. Both very dilute and concentrated urine were associated with a diverse array of sociodemographic, medical conditions, and dietary factors. For instance, females were approximately 3.3 times more likely to have urine over-dilution than male [the adjusted odds ratios (95% CI)=3.27 (2.10–5.10)]. ConclusionAlthough the determinants of urine osmolality were generally similar to those of urine creatinine, the relative influence of socio-demographic and medical conditions was less on urine osmolality than on urine creatinine. Protocols for spot urine sample collection could recommend avoiding excessive and insufficient water intake before urine sampling to improve urine adequacy. The feasibility of adopting urine osmolality adjustment and water intake recommendations before providing spot urine samples for environmental biomonitoring merits further investigation.

First confirmation of imported dengue virus serotype 2 complete genome in urine from a Chinese traveler returning from India

Dengue virus (DENV) is a mosquito-borne virus that has four serotypes. Collection of serum from patients is time- and labor- consuming, and presents a high injury risk for infants and children. The genomic and serological diagnosis of imported dengue fever from a urine sample was used as a non-invasive diagnostic method in this study. A serum sample was collected on disease day 5, and a serum and urine sample were collected on disease day 8 and 18. The results of serological tests for DENV IgM revealed that the serum samples were positive for DENV. The results of RT-qPCR assay revealed that the serum sample collected on day 5 was DENV-positive; however, the serum sample collected on day 8 and 18 were negative for DENV. The urine sample collected on day 8 and 18 were DENV-positive. We also sequenced the complete DENV genome (10723 bp) from the urine sample (GenBank KF479233). The results of phylogenetic and epidemiological analysis indicated strong confirmation that the strain was located within the DENV-2 group with a 100% bootstrap value. In this report, we (1) provided the first evidence of a DENV infection that was imported from India to a non-endemic city of China, (2) investigated the DENV genome detection having a longer timeframe for positive detection in urine sample compared to previous studies, (3) provided the sequence results for the complete DENV-2 genome from a concentrated urine sample (4) discussed how virus-typing results could be used to manage the risk of sero-specific and re-infected travel-associated dengue fever.

Abstract 610: A Pepstatin-Sensitive Aspartyl Peptidase as an Early Urinary Marker of Renal Injury in the Diabetic Hypertensive mRen2.Lewis Female Rat

We previously reported that induction of diabetes in estrogen-depleted, hypertensive mRen2.Lewis (mRen2) female rats leads to significant albuminuria, proteinuria and tubulointerstial oxidative stress at 28 days. The current study assessed the time course for the development of proteinuria and peptidase activities in the urine of the diabetic hypertensive mRen2 to begin to establish potential biomarkers of renal injury in this model. Heterozygous mRen2 underwent ovariectomy and diabetes was induced by a single administration of streptozotocin (65 mg/kg; STZ) at 15 weeks of age. Urinary samples were taken prior to (day 0) and 5 and 28 days following STZ. Aspartyl peptidases were assayed with the quenched fluorescent substrate Mca-GWPILFFRLW(Dnp)R in the presence or absence of the inhibitor pepstatin. Enzyme activities were expressed as nmols Mca per 30 min per mg creatinine. Pepstatin-sensitive activity [IC50 ∼ 0.4 nM] was detectable in the pre-diabetic urines [26 ± 9; n=8]; however, the diabetic rats exhibited a significant increase in urinary activity at days 5 and 28 [184 ± 25 and 204 ± 53, respectively; p&lt;0.05 vs. day 0]. In contrast, pepstatin-insensitive activity was markedly lower and decreased between days 0 and 5 [22 ± 2 vs. 12 ± 4; p&lt;0.05], but was unchanged at day 28 [34 ± 12]. Proteinuria was similar at days 0 and day 5, but significantly increased by day 28. Aspartyl peptidases in the kidney and urine that are sensitive to pepstatin include cathepsins D and E, as well as the enzyme napsin. However, immunoblots of concentrated urine samples using selective cathepsin and napsin antibodies failed to match protein expression to the increased activities in the diabetic rats. Finally, we assessed peptidase activity in isolated cortical slices from diabetic mRen2 to determine the extent of enzyme release. Aspartyl peptidase activity was readily detectable in the perfusate of the isolated kidney [6 ± 0.3 pmols Mca per min release, n=3] and completely inhibited by pepstatin suggesting that the urinary activity is of renal origin. We conclude that a pepstatin-sensitive aspartyl peptidase, potentially distinct from cathepsin D/E and napsin may constitute an early marker of renal injury in the diabetic hypertensive mRen2 strain.