Microscopy and culture of fungal disease

2017 ◽  
Author(s):  
Gillian S. Shankland
Keyword(s):  
Fungal Infection ◽  
Primary Culture ◽  
Culture Media ◽  
Clinical Information ◽  
Fungal Disease ◽  
Rapid Diagnosis ◽  
Clinical Samples ◽  
Presumptive Diagnosis ◽  
Selection Of ◽  
Mycological Laboratory

The clinical mycological laboratory strives to provide an accurate and rapid diagnosis of suspected fungal disease, or exclude its possibility. The most common and most traditional methods for demonstrating fungi in tissue and body fluids are microscopy and culture. The techniques discussed in this chapter are not exhaustive but have proved to be efficient for the visualization and recovery of fungi from clinical samples. In general, these methods do not require expensive or specialist equipment. The clinician’s presumptive diagnosis may help with the selection of the most appropriate specimen and aid the laboratory in the method of processing and selection of the primary culture media. However, all too often there is scant clinical information provided to the laboratory, and indeed a fungal infection may initially not have been suspected.

scholarly journals Prevalence, species distribution, and risk factors of fungal colonization and infection in patients at a burn intensive care unit in Vietnam

2020 ◽  
Author(s):  
Bang Nguyen ◽  
Nguyen Thanh Xuan ◽  
Dinh Xuan Quang ◽  
Cao Ba Loi ◽  
Nguyen Thai N Minh ◽  
...  
Keyword(s):  
Intensive Care Unit ◽  
Intensive Care ◽  
Fungal Infection ◽  
Candida Species ◽  
Clinical Information ◽  
Common Species ◽  
Clinical Samples ◽  
Fungal Colonization ◽  
Burn Patients ◽  
Severe Injuries

Background and Purpose: Burn patients are at a higher risk of infections caused by different organisms. This study aimed to address the prevalence, causative species, and factors related to fungal colonization or infection in patients with acute severe injuries admitted to the intensive care unit (ICU) of a burn hospital in northern Vietnam. Materials and Methods: This prospective study was conducted on 400 patients in a burn ICU between 2017 and 2019. Clinical samples were weekly collected and screened for fungi, and relevant clinical information was obtained from medical records. Results: According to the results, 90% of the patients were colonized with fungi. Out of this group, 12.75% of the cases had invasive fungal infection (IFI). Eleven yeasts and six mold species were isolated from the patients, with the most common species being Candida tropicalis (45.56%) and C. albicans (41.94%). Among the eleven species causing fungal wound infection (FWI), the most common agents were Candida (66.7% of FWI patients) and Aspergillus (38.5%) species. Three Candida species isolated from blood were C. tropicalis (66.7%), C. albicans (20.0%), and C. parapsilosis (14.3%). No factors were found to expose the patients to a higher risk of fungal colonization. However, hyperglycemia, prolonged ICU stay, and heavy Candida species colonization were found to be independently predictive of IFI. Conclusion: Burn patients are at the risk of fungal infection with Candida species (especially C. tropicalis) and Aspergillus as the most frequently responsible agents. Continuous surveillance of fungi and appropriate management of pathophysiological consequences are essential to prevent fungal infection in burn patients.

Identification Pseudomonas aeruginosa by OprD Gene for Differentiation from Other Pseudomonas Species that Isolated from Clinical Samples

2019 ◽  
Vol 9 (01) ◽  
pp. 46-50
Author(s):  
Ashwak B Al-Hashimy ◽  
Huda S Alagely ◽  
Akeel K Albuaji ◽  
Khalid R Majeed
Keyword(s):  
Pseudomonas Aeruginosa ◽  
General Hospital ◽  
Culture Media ◽  
Final Diagnosis ◽  
Clinical Samples ◽  
Bacterial Isolates ◽  
Molecular Method ◽  
Biochemical Tests ◽  
Pseudomonas Species ◽  
Specific Sequences

The present study included the collection of 100 samples from various clinical sources for investigating the presence of P. aeruginosa in those sources, the samples have been collected from some hospitals in Baghdad and Hillah city (Al-qassim General Hospital, ,Al-hillah teaching hospital,and Al-hashimya General hospital ) which included wounds, burns, ear and sputum infections. The study was carried out through October 2017 till the end of March 2018. The samples were identified based on the morphological and microscopically characteristics of the colonies when they were culturing or number of culture media as well as biochemical tests, molecular identification were also used as a final diagnostic test for isolates that were positive as they belong to P.aeruginosa bacteria during previous tests based on the OprD gene which has specific sequences for P.aeruginosa bacteria as a detection gene and also consider as virulence factor so it have a synonyms mechanism to antibiotic resistance . The results of the final diagnosis showed that 38 isolates belong to target bacteria were distributed as 18 of burns, 11 isolates of wounds, 6 isolates of ear infection and 3 isolates of sputum, The examination of the sensitivity of all bacterial isolates was done for elected 38 isolation towards the 9 antibiotic by a Bauer - Kirby and the isolates were resistant for a number of antibiotics used such as Ciprofloxacin 65.7%, Norflaxacin 71%, Imipenem 63.1% Meropenem 68.4%, Gentamicin 65.7%, Amikacin 26.3%, Cefepime 68.4%, Ceftazidime 65.7% and Piperacillin 57.8%.Molecular method , All isolates (38) of P. aeruginosa positive for the diagnostic special gene (OprD) genes (100%).

scholarly journals Immunomagnetic sequential ultrafiltration (iSUF) platform for enrichment and purification of extracellular vesicles from biofluids

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Jingjing Zhang ◽  
Luong T. H. Nguyen ◽  
Richard Hickey ◽  
Nicole Walters ◽  
Xinyu Wang ◽  
...  
Keyword(s):  
Extracellular Vesicles ◽  
Culture Media ◽  
Metastatic Breast ◽  
Clinical Samples ◽  
Clinical Use ◽  
Immunomagnetic Beads ◽  
Liquid Biopsies ◽  
Cell Culture Media ◽  
Non Invasive ◽  
Healthy Donors

AbstractExtracellular vesicles (EVs) derived from tumor cells have the potential to provide a much-needed source of non-invasive molecular biomarkers for liquid biopsies. However, current methods for EV isolation have limited specificity towards tumor-derived EVs that limit their clinical use. Here, we present an approach called immunomagnetic sequential ultrafiltration (iSUF) that consists of sequential stages of purification and enrichment of EVs in approximately 2 h. In iSUF, EVs present in different volumes of biofluids (0.5–100 mL) can be significantly enriched (up to 1000 times), with up to 99% removal of contaminating proteins (e.g., albumin). The EV recovery rate by iSUF for cell culture media (CCM), serum, and urine corresponded to 98.0% ± 3.6%, 96.0% ± 2.0% and 94.0% ± 1.9%, respectively (p > 0.05). The final step of iSUF enables the separation of tumor-specific EVs by incorporating immunomagnetic beads to target EV subpopulations. Serum from a cohort of clinical samples from metastatic breast cancer (BC) patients and healthy donors were processed by the iSUF platform and the isolated EVs from patients showed significantly higher expression levels of BC biomarkers (i.e., HER2, CD24, and miR21).

scholarly journals Azole Resistance in Clinical and Environmental Aspergillus Isolates from the French West Indies (Martinique)

2021 ◽  
Vol 7 (5) ◽  
pp. 355
Author(s):  
Lorra Monpierre ◽  
Nicole Desbois-Nogard ◽  
Isabel Valsecchi ◽  
Marielle Bajal ◽  
Cécile Angebault ◽  
...  
Keyword(s):  
West Indies ◽  
Culture Media ◽  
Antifungal Susceptibility ◽  
Resistance Mechanisms ◽  
Clinical Samples ◽  
Azole Resistance ◽  
Environmental Isolates ◽  
French West Indies ◽  
First Time ◽  
Microdilution Broth

The emergence of azole resistant Aspergillus spp., especially Aspergillus fumigatus, has been described in several countries around the world with varying prevalence depending on the country. To our knowledge, azole resistance in Aspergillus spp. has not been reported in the West Indies yet. In this study, we investigated the antifungal susceptibility of clinical and environmental isolates of Aspergillus spp. from Martinique, and the potential resistance mechanisms associated with mutations in cyp51A gene. Overall, 208 Aspergillus isolates were recovered from clinical samples (n = 45) and environmental soil samples (n = 163). They were screened for resistance to azole drugs using selective culture media. The Minimum Inhibitory Concentrations (MIC) towards voriconazole, itraconazole, posaconazole and isavuconazole, as shown by the resistant isolates, were determined using the European Committee on Antimicrobial Susceptibility Testing (EUCAST) microdilution broth method. Eight isolates (A. fumigatus, n = 6 and A. terreus, n = 2) had high MIC for at least one azole drug. The sequencing of cyp51A gene revealed the mutations G54R and TR34/L98H in two A. fumigatus clinical isolates. Our study showed for the first time the presence of azole resistance in A. fumigatus and A. terreus isolates in the French West Indies.

Continuous Cultures of Plasmodium Falciparum Established in Tanzania From Patients With Acute Malaria

2021 ◽  
Author(s):  
Florence Humphrey Urio ◽  
Matilda Mkombachepa ◽  
Gration Rwegasira ◽  
Twilumba Makene ◽  
Billy Ngasala ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Continuous Culture ◽  
Drug Sensitivity ◽  
Culture Media ◽  
Experimental System ◽  
Dar Es Salaam ◽  
Clinical Samples ◽  
Host Cell Interactions ◽  
Set Up

Abstract BackgroundMalaria morbidity and mortality, almost entirely from Plasmodium falciparum, are still rampant in Africa: therefore, it is important to study the biology of the parasite and the parasite-host cell interactions. In vitro cultivation of Plasmodium falciparum is most useful for this purpose, as well as for investigating drug resistance and possible new therapies. Here we report that the Trager & Jensen continuous culture of P. falciparum can be established in a laboratory in Tanzania with minimal facilities and with modest expenditure.MethodsAn in vitro set-up of continuous culture of P. falciparum was carried out in 2016 to 2020 at Muhimbili university of health and allied sciences, Dar-es salaam. Parasite samples were obtained from patients with acute malaria, frozen parasites and live cultures. Data was collected and analyzed using GraphPad Prism version 8.ResultsWe have successfully achieved exponential growth of existing strains that are used worldwide, as well as of parasites in clinical samples from patients with acute malaria. In the aim to optimize growth we have compared human serum and bovine serum albumin as components of the culture media. In addition, culture synchronization has been achieved using sorbitol.ConclusionThis experimental system is now available to our institution and to researchers aiming at investigating drug sensitivity and mechanisms of protection against Plasmodium falciparum that accrue from various genes expressed in red cells.

scholarly journals Viral Entry Properties Required for Fitness in Humans Are Lost through Rapid Genomic Change during Viral Isolation

mBio ◽  
2018 ◽  
Vol 9 (4) ◽  
Author(s):  
Sho Iketani ◽  
Ryan C. Shean ◽  
Marion Ferren ◽  
Negar Makhsous ◽  
Dolly B. Aquino ◽  
...  
Keyword(s):  
Primary Culture ◽  
Parainfluenza Virus ◽  
Clinical Samples ◽  
Genomic Change ◽  
Viral Isolation ◽  
Dimer Interface ◽  
Genome Wide ◽  
A Genome ◽  
Immortalized Cells ◽  
Human Parainfluenza Virus

ABSTRACTHuman parainfluenza viruses cause a large burden of human respiratory illness. While much research relies upon viruses grown in cultured immortalized cells, human parainfluenza virus 3 (HPIV-3) evolves in culture. Cultured viruses differ in their properties compared to clinical strains. We present a genome-wide survey of HPIV-3 adaptations to culture using metagenomic next-generation sequencing of matched pairs of clinical samples and primary culture isolates (zero passage virus). Nonsynonymous changes arose during primary viral isolation, almost entirely in the genes encoding the two surface glycoproteins—the receptor binding protein hemagglutinin-neuraminidase (HN) or the fusion protein (F). We recovered genomes from 95 HPIV-3 primary culture isolates and 23 HPIV-3 strains directly from clinical samples. HN mutations arising during primary viral isolation resulted in substitutions at HN’s dimerization/F-interaction site, a site critical for activation of viral fusion. Alterations in HN dimer interface residues known to favor infection in culture occurred within 4 days (H552 and N556). A novel cluster of residues at a different face of the HN dimer interface emerged (P241 and R242) and imply a role in HPIV-3-mediated fusion. Functional characterization of these culture-associated HN mutations in a clinical isolate background revealed acquisition of the fusogenic phenotype associated with cultured HPIV-3; the HN-F complex showed enhanced fusion and decreased receptor-cleaving activity. These results utilize a method for identifying genome-wide changes associated with brief adaptation to culture to highlight the notion that even brief exposure to immortalized cells may affect key viral properties and underscore the balance of features of the HN-F complex required for fitness by circulating viruses.IMPORTANCEHuman parainfluenza virus 3 is an important cause of morbidity and mortality among infants, the immunocompromised, and the elderly. Using deep genomic sequencing of HPIV-3-positive clinical material and its subsequent viral isolate, we discover a number of known and novel coding mutations in the main HPIV-3 attachment protein HN during brief exposure to immortalized cells. These mutations significantly alter function of the fusion complex, increasing fusion promotion by HN as well as generally decreasing neuraminidase activity and increasing HN-receptor engagement. These results show that viruses may evolve rapidly in culture even during primary isolation of the virus and before the first passage and reveal features of fitness for humans that are obscured by rapid adaptation to laboratory conditions.

scholarly journals Performance of the 4Kscore Test in Plasma and Serum and Stability of the Component Analytes in Clinical Samples

2018 ◽  
Vol 3 (2) ◽  
pp. 185-199 ◽  
Author(s):  
Christina E Higgins ◽  
Patricia Neybold ◽  
Marcella B Holdridge ◽  
Catherine R Barnes ◽  
Yan Dong ◽  
...  
Keyword(s):  
Dynamic Range ◽  
Specific Antigen ◽  
Clinical Information ◽  
Target Population ◽  
Clinical Samples ◽  
Free Psa ◽  
Total Prostate Specific Antigen ◽  
Edta Plasma ◽  
The Stability ◽  
Intact Psa

Abstract Background The 4Kscore Test determines a personalized risk score for aggressive prostate cancer by combining the blood sample measurements of total prostate-specific antigen (tPSA), free PSA (fPSA), intact PSA (iPSA), and human kallikrein-related peptidase 2 (hK2) with patient clinical information to generate the patient risk's score; thus, accuracy and precision of the 4Kscore depend on the reliability of these measurements. Although tPSA and fPSA are measured on a Food and Drug Administration (FDA)-approved platform, the performance of the iPSA and hK2 assays in the clinical setting has not previously been reported. Methods Analytical performance was determined for the iPSA and hK2 assays in both serum and EDTA plasma, according to Clinical and Laboratory Standards Institute guidelines. Equivalence of the 4Kscore in both sample matrices was demonstrated in a 353-patient clinical cohort, and the stability of endogenous iPSA and hK2 for at least 3 days was demonstrated in a smaller subset. Results Intralaboratory and interlaboratory precision of the iPSA and hK2 assays in both matrices was comparable with that of FDA-approved tPSA and fPSA assays (<18% for iPSA; <8% for hK2). The picogram per milliliter sensitivity and wide dynamic range of the iPSA and hK2 assays allowed for accurate measurements in the target population. The 4Kscore generated in either matrix up to 3 days after collection is equivalent to that measured within 24 h of collection (Passing–Bablok slope 95% CI: plasma, 0.999–1.034; serum, 0.997–1.040). Conclusions The robust performance of component assays and reliable stability of the endogenous analytes in clinical samples proven here ensures an accurate 4Kscore Test result.

scholarly journals Antifungal Susceptibility and Biofilm Production of Candida spp. Isolated from Clinical Samples

2018 ◽  
Vol 2018 ◽  
pp. 1-5 ◽  
Author(s):  
Munmun B. Marak ◽  
Biranthabail Dhanashree
Keyword(s):  
Antifungal Susceptibility ◽  
Clinical Information ◽  
Antibiotic Use ◽  
Microtiter Plate ◽  
Vaginal Swab ◽  
Diffusion Method ◽  
Clinical Samples ◽  
Candida Spp ◽  
Disk Diffusion Method ◽  
Biofilm Production

Objective. The study aims to speciate clinical Candida isolates and detect their biofilm-forming ability and antifungal resistance. Methods. All the Candida spp. isolated from different clinical samples like pus, urine, blood, and body fluid were included in the study. Biofilm production was tested by the microtiter plate method. Antifungal susceptibility was studied by the disk diffusion method. Patient’s demographic details such as age, sex, and clinical information were collected. Presence of other risk factors such as diabetes mellitus, history of antibiotic use, and any urinary tract instrumentations was also recorded. Results. Among 90 Candida species isolated, most predominant species was found to be C. albicans (45.5%) followed by C. tropicalis (28.88%), C. krusei (20%), C. glabrata (3.33%), and C. parapsilosis (2.22%). Candida spp. were isolated from urine (43%), BAL/sputum (18.88%), high vaginal swab (8.88%), suction tips (7.77%), blood and wound swabs (6.66%), pus (3.33%), bile aspirate (2.22%), and deep tissue (1.11%). A larger number of females were affected than males, and the age group of 51 to 60 years was more susceptible to candidiasis. A higher number of C. albicans isolates produced biofilm followed by C. parapsilosis, C. tropicalis, and C. krusei. However, C. glabrata showed no biofilm production in our study. All Candida isolates were 100% sensitive to amphotericin B. Voriconazole was the next effective drug with 81.11% susceptibility. 24.44% of strains were resistant to fluconazole. Conclusion. Speciation of Candida isolates, detection of ability to form the biofilm, and monitoring of antifungal susceptibility testing are necessary for appropriate treatment.

scholarly journals Selection of filamentous fungi that are resistant to the herbicides atrazine, glyphosate and pendimethalin

2021 ◽  
Vol 43 ◽  
pp. e51656
Author(s):  
Nara Priscila Barbosa Bravim ◽  
Anatércia Ferreira Alves ◽  
José Fábio França Orlanda ◽  
Patricia Barbosa Rodrigues Silva
Keyword(s):  
Filamentous Fungi ◽  
Mycelial Growth ◽  
Culture Medium ◽  
Agricultural Soils ◽  
Culture Media ◽  
Fusarium Verticillioides ◽  
Fungal Growth ◽  
Relative Growth ◽  
Maximum Inhibition ◽  
Selection Of

The objective of the present study was to isolate fungi from agricultural soils and evaluate fungal growth in culture medium contaminated with atrazine, glyphosate and pendimethalin. Filamentous fungi were isolated from agricultural soils and cultured in a modified culture medium containing 0, 10, 20, 50, and 100 μg mL-1 atrazine, glyphosate and pendimethalin for 14 days at 28°C. The fungi that presented optimal and satisfactory growth were plated in Sabouraud culture medium with 4% dextrose and containing the herbicides at concentrations of 0, 10, 20, 50, and 100 μg mL-1 for seven days at 28°C. The mean mycelial growth values were submitted to analysis of variance and the Tukey test (p < 0.05%) for comparison and relative growth determination, and maximum inhibition rates were calculated. The isolated fungi Aspergillus fumigatus, Fusarium verticillioides and Penicillium citrinum were shown to be resistant to atrazine, glyphosate and pendimethalin. F. verticillioides showed higher mean mycelial growth in the culture media contaminated with atrazine and glyphosate than the other two fungi. In the culture medium contaminated with pendimethalin, F. verticillioides, and A. fumigatus presented the highest mean mycelial growth values.

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