Flow Cytometry: An Important Diagnostic Tool in Critically Ill Preterm Neonates with Suspected Sepsis

2020 ◽  
Author(s):  
Sneha Goswami ◽  
Richa Gupta ◽  
Siddarth Ramji
Keyword(s):  
Flow Cytometry ◽  
Blood Culture ◽  
Clinical Signs ◽  
Human Leukocyte ◽  
Preterm Neonates ◽  
Suspected Sepsis ◽  
Predictive Values ◽  
Flow Cytometric ◽  
Time Flow ◽  
Culture Positive

Objective Sepsis is a major cause of neonatal mortality. The gold standard for diagnosis is blood culture which suffers from low sensitivity and huge turn-around time. Flow cytometry has been extensively applied to malignant disorders and is an upcoming tool for diagnosis of nonmalignant disorders due to its rapidity and accuracy in detecting cells, cell products, and their functional states. The aim of this study was to investigate the utility of flow cytometric expression of neutrophil CD64, monocyte human leukocyte antigen (HLA-DR) and CD16 in diagnosis in suspected preterm neonates. Study Design In total, 100 preterm neonates with clinical signs of sepsis were enrolled in the study. Blood culture, C-reactive protein (CRP) and flow cytometry for nCD64, mHLA-DR, and mCD16 were performed. The neonates were divided into two groups: culture positive and culture negative and CRP and flow cytometric findings compared. ROC analysis was performed to determine the best cut-off for nCD64, mHLA-DR, and mCD16 values along with estimation of sensitivity, specificity, and predictive values. Probability of <0.05 was taken as significant. Results Out of the 100 enrolled neonates, 34 (34%) were culture positive. CRP was not found to be significantly different in the two groups. Expression of nCD64 (p = 0.03) was significantly upregulated in the blood culture positive cases with a cut-off mean fluorescence intensity (MFI) value = 4.72 and sensitivity of 92% and specificity of 52%. Expression of mCD16 (p = 0.02) was also upregulated in the blood culture positive cases with a cut-off MFI value = 4.9, with sensitivity of 41%, specificity of 83%. Conclusion The study concluded that nCD64 and mCD16 can be potential biomarkers for early diagnosis of neonatal sepsis with a high sensitivity and specificity. Key Points

scholarly journals Study of procalcitonin in neonatal sepsis with blood culture in suspected sepsis in North India

2021 ◽  
Vol 8 (7) ◽  
pp. 1241
Author(s):  
Munna Lal Jaipal ◽  
Ajit Kumar Shrivastava ◽  
Prema Ram Choudhary
Keyword(s):  
Blood Culture ◽  
Positive Blood Culture ◽  
Tertiary Care ◽  
Clinical Signs ◽  
North India ◽  
Care Hospital ◽  
Negative Blood Culture ◽  
Suspected Sepsis ◽  
Cross Sectional ◽  
Number Of Patients

Background: Sepsis can occasionally be difficult to demonstrate, and its difference from non-infectious conditions in critically ill patients is often a challenge. Serum procalcitonin (PCT) assay is one of the biomarkers of sepsis. The aim of the study was to investigate the value of procalcitonin, in the early diagnosis of neonatal sepsis.Methods: A cross-sectional study was conducted at tertiary care hospital in New Delhi. It included all neonates with clinical signs of sepsis. The neonates were divided into two groups as sepsis, and healthy neonates. The PCT level was measured by using ELISA technique and compared between the two groups. Statistical analysis was performed using SPSS windows version 20.0 software.Results: In this study total number of patients included 350, out of which 175 were clinically suspected sepsis cases and 175 were healthy controls. 68 (39%) neonates were show positive blood culture and 107 (61%) neonates were representing negative blood culture report in study group. The mean serum value of PCT was significantly (p<0.001) higher in sepsis neonates. The serum PCT value was significantly increased in neonate’s sepsis with positive blood culture (p<0.001) and negative blood culture (p<0.001) as compared to healthy neonates.Conclusions: It is concluded from this study that the PCT assay was established to be a valuable biomarker of sepsis in this study. The assay might be performed and reported quickly and gave precious information before availability of culture results. This might assist in avoiding unnecessary antibiotic therapy.

Clinical signs and CRP values associated with blood culture results in neonates evaluated for suspected sepsis

2010 ◽  
Vol 99 (11) ◽  
pp. 1635-1640 ◽  
Author(s):  
Andreas Ohlin ◽  
Maria Björkqvist ◽  
Scott M Montgomery ◽  
Jens Schollin
Keyword(s):  
Blood Culture ◽  
Clinical Signs ◽  
Suspected Sepsis

scholarly journals Value of a Single-Tube Widal Test in Diagnosis of Typhoid Fever in Vietnam

1999 ◽  
Vol 37 (9) ◽  
pp. 2882-2886 ◽  
Author(s):  
Christopher M. Parry ◽  
Nguyen Thi Tuyet Hoa ◽  
To Song Diep ◽  
John Wain ◽  
Nguyen Tran Chinh ◽  
...  
Keyword(s):  
Blood Culture ◽  
Typhoid Fever ◽  
False Negative ◽  
Febrile Illness ◽  
Laboratory Diagnosis ◽  
Widal Test ◽  
Predictive Values ◽  
Single Tube ◽  
Culture Positive ◽  
Agglutinin Titer

The diagnostic value of an acute-phase single-tube Widal test for suspected typhoid fever was evaluated with 2,000 Vietnamese patients admitted to an infectious disease referral hospital between 1993 and 1998. Test patients had suspected typhoid fever and a blood culture positive for Salmonella typhi (n= 1,400) orSalmonella paratyphi A (n = 45). Control patients had a febrile illness for which another cause was confirmed (malaria [n = 103], dengue [n = 76], or bacteremia due to another microorganism [n = 156] or tetanus (n = 265). An O-agglutinin titer of ≥100 was found in 18% of the febrile controls and 7% of the tetanus patients. Corresponding values for H agglutinins were 8 and 1%, respectively. The O-agglutinin titer was ≥100 in 83% of the blood culture-positive typhoid fever cases, and the H-agglutinin titer was ≥100 in 67%. The disease prevalence in investigated patients in this hospital was 30.8% (95% confidence interval, 26.8 to 35.1%); at this prevalence, an elevated level of H agglutinins gave better positive predictive values for typhoid fever than did O agglutinins. With a cutoff titer of ≥200 for O agglutinin or ≥100 for H agglutinin, the Widal test would diagnose correctly 74% of the blood culture-positive cases of typhoid fever. However, 14% of the positive results would be false-positive, and 10% of the negative results would be false-negative. The Widal test can be helpful in the laboratory diagnosis of typhoid fever in Vietnam if interpreted with care.

ANTIMICROBIAL SUSCEPTIBILITY IN CULTURE POSITIVE ENTERIC FEVER

2019 ◽  
Vol 3 (7) ◽  
Author(s):  
Dr. Manish Kulshrestha ◽  
Dr. Anjali Kulshrestha
Keyword(s):  
Bone Marrow ◽  
Blood Culture ◽  
Nalidixic Acid ◽  
Antimicrobial Susceptibility ◽  
Salmonella Enterica ◽  
Enteric Fever ◽  
Multidrug Resistant ◽  
Bone Marrow Culture ◽  
Diffusion Method ◽  
Culture Positive

INTRODUCTION: Enteric fever includes typhoid and paratyphoid fever. Peak incidence is seen in children 5–15 years of age; but in regions where the disease is highly endemic, as in India, children younger than 5 years of age may have the highest infection rates. There are about 22 million new typhoid cases occur each year. Young children in poor, resource limited areas, who make up the majority of the new cases and there is a mortality figures of 215,000 deaths annually. A sharp decline in the rates of complications and mortality due to typhoid fever is observed as a result of introduction of effective antibiotic therapy since 1950s. MDR-ST became endemic in many areas of Asia, including India soon after multidrug-resistant strains of Salmonella enterica serotype typhi (MDR-ST) that were resistant to all the three first-line drugs then in use, namely chloramphenicol, amoxycillin and co-trimoxazole emerged in early 1990s. MATERIAL AND METHODS: Only blood culture or bone marrow culture positive cases were included. The patients with culture isolated enteric fever were included in the study. Antimicrobial susceptibility testing was carried out by disk diffusion method using antibiotic discs. The analysis of the antimicrobial susceptibility was carried out as per CLSI interpretative guidelines. RESULTS: A total of 82 culture positive cases were included in the present study. 80 culture isolates were from blood culture and 2 from the bone marrow culture. Salmonella entericasubspecies enterica serovartyphi (S typhi) was isolated from 67 (81.70%) patients while Salmonella enterica subspecies entericaserovarparatyphi (S paratyphi A) was isolated from 13 (15.85%) cases and 2 (2.44%) were Salmonella enterica subspecies entericaserovarschottmuelleri (S paratyphi B). Of the 82 cases 65(79.3%) isolates were resistant to ciprofloxacin, 17 (20.7%) were resistant to nalidixic acid, one (1.2%) case each was resistant to Cefotaxime and ceftriaxone, 2 (2.4%) were resistant to chloramphenicol, 10 (12.2%) were resistant and to cotrimoxazole 3 (3.7%) were resistant. CONCLUSION: In a culture positive cases 65(79.3%) isolates were resistant to ciprofloxacin and 17 (20.7%) were resistant to nalidixic acid. Multidrug resistant isolates were 65(79.3%).

scholarly journals Interleukin 8 Elicits Rapid Physiological Changes in Neutrophils That Are Altered by Inflammatory Conditions

2021 ◽  
pp. 1-17
Author(s):  
Stefan Bernhard ◽  
Stefan Hug ◽  
Alexander Elias Paul Stratmann ◽  
Maike Erber ◽  
Laura Vidoni ◽  
...  
Keyword(s):  
Systemic Inflammation ◽  
Severe Trauma ◽  
Induced Response ◽  
Neutrophil Granulocytes ◽  
Inflammatory Conditions ◽  
Flow Cytometric ◽  
Time Flow ◽  
Flow Cytometric Measurement ◽  
Detailed Kinetics

A sufficient response of neutrophil granulocytes stimulated by interleukin (IL)-8 is vital during systemic inflammation, for example, in sepsis or severe trauma. Moreover, IL-8 is clinically used as biomarker of inflammatory processes. However, the effects of IL-8 on cellular key regulators of neutrophil properties such as the intracellular pH (pH<sub>i</sub>) in dependence of ion transport proteins and during inflammation remain to be elucidated. Therefore, we investigated in detail the fundamental changes in pH<sub>i</sub>, cellular shape, and chemotactic activity elicited by IL-8. Using flow cytometric methods, we determined that the IL-8-induced cellular activity was largely dependent on specific ion channels and transporters, such as the sodium-proton exchanger 1 (NHE1) and non-NHE1-dependent sodium flux. Exposing neutrophils in vitro to a proinflammatory micromilieu with N-formyl-Met-Leu-Phe, LPS, or IL-8 resulted in a diminished response regarding the increase in cellular size and pH. The detailed kinetics of the reduced reactivity of the neutrophil granulocytes could be illustrated in a near-real-time flow cytometric measurement. Last, the LPS-mediated impairment of the IL-8-induced response in neutrophils was confirmed in a translational, animal-free human whole blood model. Overall, we provide novel mechanistic insights for the interaction of IL-8 with neutrophil granulocytes and report in detail about its alteration during systemic inflammation.

scholarly journals Diagnostic Value of Clinical Features to Distinguish Enteric Fever From Other Febrile Illnesses in Bangladesh, Nepal, and Pakistan

2020 ◽  
Vol 71 (Supplement_3) ◽  
pp. S257-S265 ◽  
Author(s):  
Kristen Aiemjoy ◽  
Dipesh Tamrakar ◽  
Shampa Saha ◽  
Shiva R Naga ◽  
Alexander T Yu ◽  
...  
Keyword(s):  
Blood Culture ◽  
Clinical Features ◽  
Diagnostic Performance ◽  
Enteric Fever ◽  
Random Effect ◽  
Clinical Signs ◽  
Febrile Illness ◽  
Diagnostic Value ◽  
Case Definition ◽  
Mixed Effect

Abstract Background Enteric fever, a bacterial infection caused by Salmonella enterica serotypes Typhi and Paratyphi A, frequently presents as a nonlocalizing febrile illness that is difficult to distinguish from other infectious causes of fever. Blood culture is not widely available in endemic settings and, even when available, results can take up to 5 days. We evaluated the diagnostic performance of clinical features, including both reported symptoms and clinical signs, of enteric fever among patients participating in the Surveillance for Enteric Fever in Asia Project (SEAP), a 3-year surveillance study in Bangladesh, Nepal, and Pakistan. Methods Outpatients presenting with ≥3 consecutive days of reported fever and inpatients with clinically suspected enteric fever from all 6 SEAP study hospitals were eligible to participate. We evaluated the diagnostic performance of select clinical features against blood culture results among outpatients using mixed-effect regression models with a random effect for study site hospital. We also compared the clinical features of S. Typhi to S. Paratyphi A among both outpatients and inpatients. Results We enrolled 20 899 outpatients, of whom 2116 (10.1%) had positive blood cultures for S. Typhi and 297 (1.4%) had positive cultures for S. Paratyphi A. The sensitivity of absence of cough was the highest among all evaluated features, at 65.5% (95% confidence interval [CI], 55.0–74.7), followed by measured fever at presentation at 59.0% (95% CI, 51.6–65.9) and being unable to complete normal activities for 3 or more days at 51.0% (95% CI, 23.8–77.6). A combined case definition of 3 or more consecutive days of reported fever and 1 or more of the following (a) either the absence of cough, (b) fever at presentation, or (c) 3 or more consecutive days of being unable to conduct usual activity--yielded a sensitivity of 94.6% (95% CI, 93.4–95.5) and specificity of 13.6% (95% CI, 9.8–17.5). Conclusions Clinical features do not accurately distinguish blood culture–confirmed enteric fever from other febrile syndromes. Rapid, affordable, and accurate diagnostics are urgently needed, particularly in settings with limited or no blood culture capacity.

scholarly journals O-GlcNAcylation in early stages of chronic lymphocytic leukemia; protocol development for flow cytometry

2021 ◽  
pp. 1-10
Author(s):  
Viktória Temesfői ◽  
Kinga Molnár ◽  
Péter Kaltenecker ◽  
Barbara Réger ◽  
Árpád Szomor ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Lymphocyte Count ◽  
Blood Parameters ◽  
Cell Number ◽  
Metabolic Changes ◽  
Protein Glycosylation ◽  
Lymphocytic Leukemia ◽  
Tween 20 ◽  
Protocol Development ◽  
Flow Cytometric

BACKGROUND: Recent studies proved that metabolic changes in malignant disorders have an impact on protein glycosylation, however, only a few attempts have been made so far to use O-GlcNAc analysis as a prognostic tool. Glucose metabolism is reported to be altered in hematological malignancies thus, we hypothesized that monitoring intracellular O-GlcNAc levels in Rai stage 0-I (Binet A) CLL patients could give deeper insights regarding subtle metabolic changes of progression which are not completely detected by the routine follow-up procedures. OBJECTIVE: In this proof of concept study we established a flow cytometric detection method for the assessment of O-GlcNAcylation as a possible prognostic marker in CLL malignancy which was supported by fluorescence microscopy. METHODS: Healthy volunteers and CLL patients were recruited for this study. Lymphocytes were isolated, fixed and permeabilised by various methods to find the optimal experimental condition for O-GlcNAc detection by flow cytometry. O-GlcNAc levels were measured and compared to lymphocyte count and various blood parameters including plasma glucose level. RESULTS: The protocol we developed includes red blood cell lysis, formalin fixation, 0.1% Tween 20 permeabilisation and employs standardized cell number per sample and unstained controls. We have found significant correlation between O-GlcNAc levels and WBC (R2= 0.8535, p< 0.0029) and lymphocyte count (R2= 0.9225, p< 0.0006) in CLL patients. Interestingly, there was no such correlation in healthy individuals (R2= 0.05664 for O-GlcNAc vs WBC and R2= 0.04379 for O-GlcNAc vs lymphocytes). CONCLUSION: Analyzing O-GlcNAc changes in malignant disorders, specifically in malignant hematologic diseases such as CLL, could be a useful tool to monitor the progression of the disease.

scholarly journals Cord Blood Extracellular Vesicles Analyzed by Flow Cytometry with Thresholding Using 405 nm or 488 nm Laser Leads to Concurrent Results

Diagnostics ◽  
2021 ◽  
Vol 11 (8) ◽  
pp. 1320
Author(s):  
Kristýna Pekárková ◽  
Jakub Soukup ◽  
Marie Kostelanská ◽  
Jan Širc ◽  
Zbyněk Straňák ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Cord Blood ◽  
Extracellular Vesicles ◽  
Correlation Coefficients ◽  
Flow Cytometer ◽  
Liquid Biopsies ◽  
Physiological Conditions ◽  
Flow Cytometric ◽  
Term Births ◽  
And Control

Extracellular vesicles (EVs) from liquid biopsies are extensively analyzed by flow cytometry, a technology that is continuously evolving. Thresholding utilizing a violet 405 nm laser side scatter (VSSC) has recently been implemented. Here, we collected set of large EV (lEV) samples from cord blood, which we analyzed using a standard flow cytometer improved via a 405 nm laser side scatter. Samples were analyzed using two distinct thresholding methods—one based on VSSC, and one based on VSSC combined with fluorescence thresholding on stained phosphatidylserine. Through these thresholding methods, we compared lEVs from pre-term births and control cord blood. Double-labeled lEVs with platelet CD36+/CD41+, activated platelet CD41+/CD62P+ and endothelial CD31+/CD105+ antibodies were used. Apart from comparing the two groups together, we also correlated measured lEVs with the thresholding methods. We also correlated the results of this study with data analyzed in our previous study in which we used a conventional 488 nm laser SSC. We did not find any difference between the two cord blood groups. However, we found highly concurrent data via our correlation of the thresholding methods, with correlation coefficients ranging from 0.80 to 0.96 even though the numbers of detected lEVs differed between thresholding methods. In conclusion, our approaches to thresholding provided concurrent data and it seems that improving the cytometer with the use of a VSSC increases its sensitivity, despite not being particularly critical to the validity of flow cytometric studies that compare pathological and physiological conditions in liquid biopsies.

scholarly journals Flow cytometric measurements as a proxy for sporulation intensity in the cultured macroalga Ulva (Chlorophyta)

2021 ◽  
Vol 0 (0) ◽  
Author(s):  
Omri Nahor ◽  
Cristina F. Morales-Reyes ◽  
Gianmaria Califano ◽  
Thomas Wichard ◽  
Alexander Golberg ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Life Cycle ◽  
Abiotic Factors ◽  
Physiological Status ◽  
Seaweed Cultivation ◽  
Reproductive Growth ◽  
Biotic And Abiotic Factors ◽  
Flow Cytometric ◽  
Abiotic Stimuli ◽  
Fundamental Shift

Abstract Controlling the life cycle of the green macroalga Ulva (Chlorophyta) is essential to maintain its efficient aquaculture. A fundamental shift in cultivation occurs by transforming the thallus cells into gametangia and sporangia (sporulation), with the subsequent release of gametes and zoids. Sporulation occurrence depends on algal age and abiotic stimuli and is controlled by sporulation inhibitors. Thus, quantification of sporulation intensity is critical for identifying the biotic and abiotic factors that influence the transition to reproductive growth. Here, we propose to determine the sporulation index by measuring the number of released gametes using flow cytometry, in proportion to the total number of thallus cells present before the occurrence of the sporulation event. The flow cytometric measurements were validated by manually counting the number of released gametes. We observed a variation in the autofluorescence levels of the gametes which were released from the gametangia. High autofluorescence level correlated to phototactically active behaviour of the gametes. As autofluorescence levels varied between different groups of gametes related to their mobility, flow cytometry can also determine the physiological status of the gametes used as feedstock in seaweed cultivation.

Sign in / Sign up

Export Citation Format

Share Document