scholarly journals Overcoming the Negligence in Laboratory Diagnosis of Mucosal Leishmaniasis

Pathogens ◽  
2021 ◽  
Vol 10 (9) ◽  
pp. 1116
Author(s):  
Lilian Motta Cantanhêde ◽  
Cristiane Batista Mattos ◽  
Ana Karoline Cruz ◽  
Yoda Janaina Ikenohuchi ◽  
Flavia Gonçalves Fernandes ◽  
...  
Keyword(s):  
Rna Virus ◽  
Laboratory Diagnosis ◽  
Northern Region ◽  
Clinical Samples ◽  
Sample Collection ◽  
Cutaneous Lesions ◽  
Blood Samples ◽  
Less Invasive ◽  
Reference Services ◽  
Mucosal Leishmaniasis

The northern region of Brazil, which has the largest number of cases of tegumentary leishmaniasis (TL) in the country, is also the region that has the highest diversity of species of vectors and Leishmania parasites. In this region, cases of mucosal leishmaniasis (ML), a clinical form of TL, exceed the national average of cases, reaching up to 12% of the total annual TL notifications. ML is associated with multiple factors, such as the parasite species and the viral endosymbiont Leishmania RNA virus 1 (LRV1). Being a chronic parasitological disease, laboratory diagnosis of ML poses a challenge for health services. Here, we evaluated more than 700 clinical samples from patients with clinical suspicion of TL, including patients with cutaneous leishmaniasis (CL) and mucosal leishmaniasis, comparing the results of parasitological tests—direct parasitological examination by microscopy (DP) and conventional PCR (cPCR) targeting of both kDNA and hsp70. The DP was performed by collecting material from lesions through biopsies (mucosal lesions) or scarification (cutaneous lesions); for PCR, a cervical brush was used for sample collection. Blood samples were tested employing standardized real-time PCR (qPCR) protocol targeting the HSP70 gene. PCR tests showed higher sensitivity than DP for both CL and ML samples. Considering ML samples only (N = 89), DP showed a sensitivity of 49.4% (N = 44) against 98.8% (N = 88) for kDNA PCR. The qPCR hsp70 for blood samples from patients with ML (N = 14) resulted in superior sensitivity (50%; N = 7) compared to DP (21.4%; N = 3) for samples from the same patients. Our results reinforced the need to implement a molecular test for the diagnosis of ML, in addition to proposing methods less invasive for collecting material from TL patients. Sample collection using a cervical brush in lesions observed in CL and ML patients is easy to perform and less invasive, compared to scarification and biopsies. Blood samples could be a good source for qPCR diagnosis for ML patients. Thus, we propose here a standardized method for collection and for performing of molecular diagnosis of clinical samples from suspicious ML patients that can be applied in reference services for improving ML diagnosis.

scholarly journals Resource optimization in COVID-19 diagnosis

2020 ◽  
Author(s):  
S.A Taniwaki ◽  
S.O.S Silva ◽  
N.F Santana-Clavijo ◽  
J. A Conselheiro ◽  
G.T Barone ◽  
...  
Keyword(s):  
Clinical Sample ◽  
Laboratory Diagnosis ◽  
Resource Optimization ◽  
Confirmatory Test ◽  
Clinical Samples ◽  
Sample Collection ◽  
Reaction Volume ◽  
Rapid Dissemination ◽  
Alternative Type ◽  
Diagnostic Capacity

AbtsractThe emergence and rapid dissemination worldwide of a novel Coronavirus (SARS-CoV-2) results in decrease of swabs availability for clinical samples collection, as well as, reagents for RT-qPCR diagnostic kits considered a confirmatory test for COVID-19 infection. This scenario, showed the requirement of improve de diagnostic capacity, so the aim of this study were to verify the possibility of reducing the reaction volume of RT-qPCR and to test cotton swabs as alternative for sample collection. RT-qPCR volumes and RNA sample concentration were optimized without affecting the sensitivity of assays, using both probe-based and intercalation dyes methods. Although rayon swabs showed better performance, cotton swabs could be used as alternative type for clinical sample collection. COVID-19 laboratory diagnosis is important to isolate and restrict the dissemination of virus, so seek for alternatives to decrease the coast of assays improve the control of disease.

Rapid Radioimmunoassay For Fibrinopeptide A In Human Plasma

1981 ◽  
Author(s):  
B J Woodhams ◽  
P B A Kernoff
Keyword(s):  
Human Plasma ◽  
Degradation Products ◽  
Clinical Samples ◽  
Blood Collection ◽  
Sample Collection ◽  
Normal Range ◽  
Blood Samples ◽  
Fibrin Degradation Products ◽  
Sequential Samples

The originally-described method for assay of fibrino- peptide A (FPA) in human plasma includes alcohol precipitation and dialysis steps which are complex, time consuming, and limit the applicability of the assay. The purpose of this work was to devise an assay for FPA which could be more easily applied to clinical studies, and to use this assay to study some of the factors which might cause artefactual results on blood samples obtained from patients. A rapid method for FPA assay has been developed which is simple, robust and sensitive and allows results to be obtained within 2.5 hrs. of blood collection. The assay depends on the use of bentonite to absorb fibrinogen from plasma, and gives a normal range for plasma FPA (0.3 - 1.5 pmol/ml) which is similar to that measured using the originally-described method. There is an equally good correlation between results obtained by the two methods on samples obtained from patients with various levels of circulating FPA and/or fibrinogen/fibrin degradation products (FDPs). Following venepuncture, FPA levels in sequential samples of blood drawn through 19-gauge ‘Butterfly’ needles became progressively lower in successive samples, indicating that a larger than usual discard of blood is necessary if basal levels are to be accurately assessed. Different antisera showed differences in affinity for FPA and cross reaction with des amino tyrosyl FPA, but such differences are unlikely to cause problems when assaying clinical samples. Generation of FPA from whole blood in vitro is completely suppressed by heparin/Trasylol anticoagulant in conventionally-used concentration, and we therefore find no evidence in favour of the suggestion that this anticoagulant mixture is unsuitable for sample collection for FPA assay.

Isolation of Stenotrophomonas maltophilia from Clinical Samples in Some Hospitals in Shiraz, Iran

2020 ◽  
pp. 1-3
Author(s):  
Majid Baserisalehi ◽  
Samira Zarezadeh ◽  
Majid Baserisalehi ◽  
Saeed Shoa
Keyword(s):  
Stenotrophomonas Maltophilia ◽  
Enterobacter Aerogenes ◽  
Late Summer ◽  
Early Spring ◽  
Clinical Samples ◽  
Bacillus Species ◽  
Microbiological Analysis ◽  
Blood Samples ◽  
Healthcare Facilities ◽  
Phenotypic Methods

Stenotrophomonas maltophilia is an emerging pathogenic non-fermentative Gram-negative Bacillus species. It has caused many nosocomial infections and can be isolated from various hospital wards and healthcare facilities. Research has shown that most of its strains are inherently resistant to many antibiotics and have multidrug resistance. This research intended to determine its occurrence frequency at some Hospitals in shiraz, Iran. The present study was conducted in six months (from early spring to late summer 2019). Clinical samples (Blood, Urine and cerebrospinal fluid (CSF)) collected from 120 patients afflicted with various infections. The samples were transferred to the Laboratory and subjected to microbiological analysis. Identification of the isolates was carried out by phenotypic methods and Stenotrophomonas maltophilia isolates verified using molecular methods. In total, various bacteria were isolated from 84 clinical samples. The isolates were Escherichia coli, Enterobacter aerogenes, Klebsiella pneumoniae, Stenotrophomonas maltophilia, Staphylococcus aureus and Pseudomonas aeruginosa. Stenotrophomonas maltophilia was isolated from 17 (20.2%) positive samples and most of them were isolated from blood samples. Our finding indicated that Stenotrophomonas maltophilia isolated more from blood samples follow by CSF sample. In addition, our finding illustrated that Stenotrophomonas maltophilia can be considered as the common nosocomial agent at hospitals in Shiraz, Iran.

scholarly journals Factors Enhancing Serum Syndecan-1 Concentrations: A Large-Scale Comprehensive Medical Examination

2019 ◽  
Vol 8 (9) ◽  
pp. 1320
Author(s):  
Kazumasa Oda ◽  
Hideshi Okada ◽  
Akio Suzuki ◽  
Hiroyuki Tomita ◽  
Ryo Kobayashi ◽  
...  
Keyword(s):  
Large Scale ◽  
Prospective Observational Study ◽  
Nitrogen Concentration ◽  
Laboratory Data ◽  
University Hospital ◽  
Endothelial Glycocalyx ◽  
Sample Collection ◽  
Blood Samples ◽  
Triglyceride Concentration ◽  
Medical Examinations

Endothelial disorders are related to various diseases. An initial endothelial injury is characterized by endothelial glycocalyx injury. We aimed to evaluate endothelial glycocalyx injury by measuring serum syndecan-1 concentrations in patients during comprehensive medical examinations. A single-center, prospective, observational study was conducted at Asahi University Hospital. The participants enrolled in this study were 1313 patients who underwent comprehensive medical examinations at Asahi University Hospital from January 2018 to June 2018. One patient undergoing hemodialysis was excluded from the study. At enrollment, blood samples were obtained, and study personnel collected demographic and clinical data. No treatments or exposures were conducted except for standard medical examinations and blood sample collection. Laboratory data were obtained by the collection of blood samples at the time of study enrolment. According to nonlinear regression, the concentrations of serum syndecan-1 were significantly related to age (p = 0.016), aspartic aminotransferase concentration (AST, p = 0.020), blood urea nitrogen concentration (BUN, p = 0.013), triglyceride concentration (p < 0.001), and hematocrit (p = 0.006). These relationships were independent associations. Endothelial glycocalyx injury, which is reflected by serum syndecan-1 concentrations, is related to age, hematocrit, AST concentration, BUN concentration, and triglyceride concentration.

Approaching stability challenges for flow cytometry in a regulated bioanalytical environment

Bioanalysis ◽  
2019 ◽  
Vol 11 (20) ◽  
pp. 1845-1858 ◽  
Author(s):  
Anamica Muruganandham ◽  
Carolyne Dumont ◽  
Kuiyi Xing
Keyword(s):  
Flow Cytometry ◽  
Critical Parameter ◽  
Storage Period ◽  
Reliable Data ◽  
Clinical Samples ◽  
Sample Collection ◽  
Testing Laboratory ◽  
Stability Parameters ◽  
Stability Issues ◽  
Limited Period

Stability of samples for flow cytometry is a critical parameter since storage period of samples is restricted to only a limited period after collection. For most studies, clinical samples have to be shipped to a testing laboratory, in contrast to preclinical samples, which can be analyzed on-site or off-site. Therefore, evaluating stability is critical to provide flexibility on testing of samples to obtain reliable data. A wide variety of factors contributes to establishing stability from sample collection through acquisition. We provided suggestions for experimental and stability parameters to be taken into consideration when designing a flow cytometry method. The case studies presented represent how certain stability issues were overcome to perform flow cytometry assays in a regulated bioanalytical environment.

scholarly journals Acanthamoeba spp. as Agents of Disease in Humans

2003 ◽  
Vol 16 (2) ◽  
pp. 273-307 ◽  
Author(s):  
Francine Marciano-Cabral ◽  
Guy Cabral
Keyword(s):  
Cerebrospinal Fluid ◽  
Cutaneous Lesion ◽  
Laboratory Diagnosis ◽  
Immune Defense ◽  
In Vitro Cultivation ◽  
Cutaneous Lesions ◽  
Trophozoite Stage ◽  
Biopsy Material ◽  
Pathogenic Potential

SUMMARY Acanthamoeba spp. are free-living amebae that inhabit a variety of air, soil, and water environments. However, these amebae can also act as opportunistic as well as nonopportunistic pathogens. They are the causative agents of granulomatous amebic encephalitis and amebic keratitis and have been associated with cutaneous lesions and sinusitis. Immuno compromised individuals, including AIDS patients, are particularly susceptible to infections with Acanthamoeba. The immune defense mechanisms that operate against Acanthamoeba have not been well characterized, but it has been proposed that both innate and acquired immunity play a role. The ameba's life cycle includes an active feeding trophozoite stage and a dormant cyst stage. Trophozoites feed on bacteria, yeast, and algae. However, both trophozoites and cysts can retain viable bacteria and may serve as reservoirs for bacteria with human pathogenic potential. Diagnosis of infection includes direct microscopy of wet mounts of cerebrospinal fluid or stained smears of cerebrospinal fluid sediment, light or electron microscopy of tissues, in vitro cultivation of Acanthamoeba, and histological assessment of frozen or paraffin-embedded sections of brain or cutaneous lesion biopsy material. Immunocytochemistry, chemifluorescent dye staining, PCR, and analysis of DNA sequence variation also have been employed for laboratory diagnosis. Treatment of Acanthamoeba infections has met with mixed results. However, chlorhexidine gluconate, alone or in combination with propamidene isethionate, is effective in some patients. Furthermore, effective treatment is complicated since patients may present with underlying disease and Acanthamoeba infection may not be recognized. Since an increase in the number of cases of Acanthamoeba infections has occurred worldwide, these protozoa have become increasingly important as agents of human disease.

scholarly journals Assessment of Plasmodium antigens and CRP in dried blood spots with multiplex malaria array

2021 ◽  
Author(s):  
Ihn Kyung Jang ◽  
Sara Aranda ◽  
Rebecca Barney ◽  
Andrew Rashid ◽  
Muhammad Helwany ◽  
...  
Keyword(s):  
Thermal Stability ◽  
Whole Blood ◽  
Dried Blood Spots ◽  
Careful Consideration ◽  
Clinical Samples ◽  
Sample Type ◽  
Sample Collection ◽  
Malaria Surveillance ◽  
Inflammatory Biomarker ◽  
Blood Spots

AbstractDried blood spots (DBS) typically prepared on filter papers are an ideal sample type for malaria surveillance by offering easy and cost-effective methods in terms of sample collection, storage, and transport. The objective of this study was to evaluate the applicability of DBS with a commercial multiplex malaria assay, developed to concurrently measure Plasmodium antigens, histidine-rich protein 2 (HRP2), Plasmodium lactate dehydrogenase (pLDH), and a host inflammatory biomarker, C-reactive protein (CRP), in whole blood. The assay conditions were optimized for DBS, and thermal stability for measurement of Plasmodium antigens and CRP in dried blood were determined. Performance of the multiplex assay on matched DBS and whole blood pellet samples was also evaluated using the clinical samples. The results indicate the acceptable performance in multiplex antigen detection using DBS samples. At cutoff levels for DBS, with a diagnostic specificity with a lower 95% confidence bound > 92%, diagnostic sensitivities against polymerase chain reaction (PCR)–confirmed malaria for HRP2, Pf LDH, Pv LDH, and Pan LDH were 93.5%, 80.4%, 21.3%, and 55.6%, respectively. The half-life of pLDH was significantly less than that of HRP2 in thermal stability studies. Results with DBS samples collected from Peru indicate that the uncontrolled storage conditions of DBS can result in inaccurate reporting for infection with P. falciparum parasites with hrp2/3 deletions. With careful consideration that minimizing the unfavorable DBS storage environment is essential for ensuring integrity of heat-labile Plasmodium antigens, DBS samples can be used as an alternative to liquid whole blood to detect P. falciparum with hrp2/3 deletions in malaria surveillance.

Techniques' Comparison and Report of the North Carolina Experience

PEDIATRICS ◽  
1989 ◽  
Vol 83 (5) ◽  
pp. 843-848
Author(s):  
Thomas R. Kinney ◽  
Martha Sawtschenko ◽  
Mary Whorton ◽  
Jean Shearin ◽  
Christy Stine ◽  
...  
Keyword(s):  
North Carolina ◽  
Filter Paper ◽  
Screening Program ◽  
Medical Center ◽  
Sample Collection ◽  
Capillary Tubes ◽  
Blood Samples ◽  
Screening Programs ◽  
The North ◽  
Duke University

Controversy still exists as to the best laboratory method to use to screen newborns for sickle cell disease and other hemoglobinopathies. The proposed methods include hemoglobin electrophoresis, column chromatography, isoelectric focusing, and high performance liquid chromatography. There is also debate concerning the preferred method of sample collection. The proposed methods of sample collection include cord blood or blood obtained from the infant collected in a tube with anticoagulant or on filter paper. We compared hemoglobin electrophoresis patterns from infant blood samples collected in heparinized capillary tubes and on filter paper. This comparison was performed because hemoglobin electrophoresis of dried blood samples collected on filter paper has been advocated as a practical, reliable, and inexpensive method for mass screening programs, although the limitations of this technique have not been explored fully. We also summarize data from the North Carolina Newborn Hemoglobinopathy Screening Program, which relates to the advantages and limitations of hemoglobin electrophoresis from filter paper blood specimens. MATERIALS AND METHODS Specimens Four sets of specimens were used for this study: (1) specimens collected at Duke University Medical Center to compare hemoglobin electrophoresis patterns of hemolysates from filter paper and heparinized capillary tubes, (2) specimens collected by the North Carolina program for hemoglobinopathy screening, (3) specimens routinely collected at Duke University in heparinized capillary tubes for newborn hemoglobinopathy screening, and (4) samples for retesting to examine the error rate of the state program and to confirm screening results compatible with a hemoglobinopathy. Samples for Direct Comparison Between Filter Paper and Heparinized Specimens

scholarly journals Recent Advances in Portable Biosensors for Biomarker Detection in Body Fluids

Biosensors ◽  
2020 ◽  
Vol 10 (9) ◽  
pp. 127 ◽  
Author(s):  
Brian Senf ◽  
Woon-Hong Yeo ◽  
Jong-Hoon Kim
Keyword(s):  
Minimally Invasive ◽  
Clinical Intervention ◽  
Body Fluids ◽  
Disease Spread ◽  
Biomarker Detection ◽  
Sample Collection ◽  
Less Invasive ◽  
Recent Advances ◽  
Detection Of Biomarkers

A recent development in portable biosensors allows rapid, accurate, and on-site detection of biomarkers, which helps to prevent disease spread by the control of sources. Less invasive sample collection is necessary to use portable biosensors in remote environments for accurate on-site diagnostics and testing. For non- or minimally invasive sampling, easily accessible body fluids, such as saliva, sweat, blood, or urine, have been utilized. It is also imperative to find accurate biomarkers to provide better clinical intervention and treatment at the onset of disease. At the same time, these reliable biomarkers can be utilized to monitor the progress of the disease. In this review, we summarize the most recent development of portable biosensors to detect various biomarkers accurately. In addition, we discuss ongoing issues and limitations of the existing systems and methods. Lastly, we present the key requirements of portable biosensors and discuss ideas for functional enhancements.

scholarly journals Inactivation of SARS-CoV-2 virus in saliva using a guanidium based transport medium suitable for RT-PCR diagnostic assays

PLoS ONE ◽  
2021 ◽  
Vol 16 (6) ◽  
pp. e0252687
Author(s):  
Sukalyani Banik ◽  
Kaheerman Saibire ◽  
Shraddha Suryavanshi ◽  
Glenn Johns ◽  
Soumitesh Chakravorty ◽  
...  
Keyword(s):  
Limit Of Detection ◽  
Viral Rna ◽  
Clinical Samples ◽  
Sample Collection ◽  
Rt Pcr ◽  
Diagnostic Assays ◽  
Guanidinium Thiocyanate ◽  
Upper Respiratory ◽  
Polymerase Chain ◽  
The Stability

Background Upper respiratory samples used to test for SARS-CoV-2 virus may be infectious and present a hazard during transport and testing. A buffer with the ability to inactivate SARS-CoV-2 at the time of sample collection could simplify and expand testing for COVID-19 to non-conventional settings. Methods We evaluated a guanidium thiocyanate-based buffer, eNAT™ (Copan) as a possible transport and inactivation medium for downstream Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR) testing to detect SARS-CoV-2. Inactivation of SARS-CoV-2 USA-WA1/2020 in eNAT and in diluted saliva was studied at different incubation times. The stability of viral RNA in eNAT was also evaluated for up to 7 days at room temperature (28°C), refrigerated conditions (4°C) and at 35°C. Results SARS-COV-2 virus spiked directly in eNAT could be inactivated at >5.6 log10 PFU/ml within a minute of incubation. When saliva was diluted 1:1 in eNAT, no cytopathic effect (CPE) on VeroE6 cells was observed, although SARS-CoV-2 RNA could be detected even after 30 min incubation and after two cell culture passages. A 1:2 (saliva:eNAT) dilution abrogated both CPE and detectable viral RNA after as little as 5 min incubation in eNAT. SARS-CoV-2 RNA from virus spiked at 5X the limit of detection remained positive up to 7 days of incubation in all tested conditions. Conclusion eNAT and similar guanidinium thiocyanate-based media may be of value for transport, stabilization, and processing of clinical samples for RT-PCR based SARS-CoV-2 detection.

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