16 results on '"Ukey R"'
Search Results
2. Validation of the Diabetomics CovAB SARS-CoV-2 antibody test in children: comparison with serology.
- Author
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Gurumurthy M, Schwab J, Pentakota SR, Ukey R, Gennaro M, Thomas P, and Friedman S
- Subjects
- Humans, Child, Child, Preschool, Female, Male, Cross-Sectional Studies, Infant, Sensitivity and Specificity, Seroepidemiologic Studies, United States epidemiology, COVID-19 diagnosis, COVID-19 epidemiology, COVID-19 immunology, Antibodies, Viral blood, SARS-CoV-2 immunology, SARS-CoV-2 isolation & purification, COVID-19 Serological Testing methods
- Abstract
Monitoring antibody prevalence is a valuable tool to evaluate the burden of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in a community, identify risk factors, and assess the impact of clinical and public health intervention strategies. The antibody prevalence of SARS-CoV-2 in children in the United States in early 2022 was estimated by the Centers for Disease Control and Prevention to be 74.2%, using seroprevalence from a variety of sources. A study by the New Jersey Department of Health in late 2022/early 2023 in unvaccinated children found a lower prevalence, 68% when using a gum swab method to detect antibodies. This study compared the accuracy of the gum swab method to detect antibodies with simultaneously obtained serological samples in additional children. This cross-sectional study recruited well children, not vaccinated for SARS-CoV-2, aged 18 months to 11 years, who were scheduled for routine bloodwork at an inner-city university-based pediatric clinic. With parental consent, an extra 5 cc of blood and a gum swab sample were collected. Results from Diabetomics CovAb SARS-CoV-2 gum swab antibody test and Rutgers New Jersey Medical School enzyme-linked immunosorbent assay serology test for spike protein antibody were compared. The seropositivity of these paired samples was compared using McNemar's test, Cohen's kappa statistic, and other diagnostic accuracy statistics. From June through August 2023, 86 children were recruited. Antibody positivity by gum swab was 70.9% and by serology was 87.2%. The Cohen's kappa statistic was 0.39 indicating minimal agreement and McNemar's test was significant ( P -value of 0.0010). Compared with serology, gum swab was 78.7% sensitive (95% CI 68.7% to 87.3%) and 81.8% specific (95% CI 48.2% to 97.7%). Positive and negative predictive values were 97.5% and 29.9%, respectively, and accuracy was 79.0%. Sensitivity in non-Hispanic versus Hispanic children was 74.2% versus 82.5%, and in children 6-11 years versus 18 months to 5 years, it was 74.2% versus 81.8%. While the gum swab method of antibody detection is not as sensitive or specific as serology, sample collection can be done in settings where phlebotomy is not feasible. This method could be useful in non-clinical settings such as surveillance, for assessing epidemiological trends and associations., Importance: Recently a study determining the prevalence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antibodies in unvaccinated children in NJ (Katic et al. 2023. Pediatric Academic Societies Meeting; Washington, D.C. https://2023.pas-meeting.org/searchbyposterbucket.asp?bm=Public+Health+%26+Prevention&t=Public+Health+%26+Prevention&pfp=Track) was conducted using a gum swab method for antibody detection. The Diabetomics CovAB test, which qualitatively identifies antibodies to SARS-CoV-2 spike protein, is a point-of-care, low-cost test, that is easy to administer in children. While this test provides sensitive and specific results in adults [US Food and Drug Administration (FDA). 2022. Center for devices and radiological health. EUA authorized serology test performance. Available from: https://www.fda.gov/medical-devices/covid-19-emergency-use-authorizations-medical-devices/eua-authorized-serology-test-performance], data on its accuracy in children is lacking. As a follow-up to the above-mentioned study, we compared the results of the gum swab test to a serologic antibody test. We found that the gum swab test was inferior to serology but was fairly sensitive and specific with a high positive predictive value. While the test is not ideal for diagnostic purposes in children it can be a valuable tool for public health officials and pediatricians to understand the extent of past health interventions., Competing Interests: The authors declare no conflict of interest.
- Published
- 2024
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3. Cryptococcosis, tuberculosis, and a kidney cancer fail to fit the atherosclerosis paradigm for foam cell lipid content.
- Author
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Guerrini V, Prideaux B, Khan R, Subbian S, Wang Y, Sadimin E, Pawar S, Ukey R, Singer EA, Xue C, and Gennaro ML
- Abstract
Foam cells are dysfunctional, lipid-laden macrophages associated with chronic inflammation of diverse origin. The long-standing paradigm that foam cells are cholesterol-laden derives from atherosclerosis research. We previously showed that, in tuberculosis, foam cells surprisingly accumulate triglycerides. Here, we utilized bacterial ( Mycobacterium tuberculosis ), fungal ( Cryptococcus neoformans ), and human papillary renal cell carcinoma (pRCC) models to address the need for a new explanation of foam cell biogenesis. We applied mass spectrometry-based imaging to assess the spatial distribution of storage lipids relative to foam-cell-rich areas in lesional tissues, and we characterized lipid-laden macrophages generated under corresponding in vitro conditions. The in vivo data and the in vitro findings showed that cryptococcus-infected macrophages accumulate triglycerides, while macrophages exposed to pRCC- conditioned-medium accumulated both triglycerides and cholesterol. Moreover, cryptococcus- and mycobacterium-infected macrophages accumulated triglycerides in different ways. Collectively, the data show that the molecular events underlying foam cell formation are specific to disease and microenvironment. Since foam cells are potential therapeutic targets, recognizing that their formation is disease-specific opens new biomedical research directions.
- Published
- 2024
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4. A genetically modulated Toll-like-receptor-tolerant phenotype in peripheral blood cells of children with multisystem inflammatory syndrome.
- Author
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Khan R, Ji W, Guzman-Rivera J, Madhvi A, Andrews T, Richlin B, Suarez C, Gaur S, Cuddy W, Singh AR, Bukulmez H, Kaelber D, Kimura Y, Ganapathi U, Michailidis IE, Ukey R, Moroso-Fela S, Kuster JK, Casseus M, Roy J, Kleinman LC, Horton DB, Lakhani SA, and Gennaro ML
- Abstract
Dysregulated innate immune responses contribute to multisystem inflammatory syndrome in children (MIS-C), characterized by gastrointestinal, mucocutaneous, and/or cardiovascular injury occurring weeks after SARS-CoV-2 exposure. To investigate innate immune functions in MIS-C, we stimulated ex vivo peripheral blood cells from MIS-C patients with agonists of Toll-like receptors (TLR), key innate immune response initiators. We found severely dampened cytokine responses and elevated gene expression of negative regulators of TLR signaling. Increased plasma levels of zonulin, a gut leakage marker, were also detected. These effects were also observed in children enrolled months after MIS-C recovery. Moreover, cells from MIS-C children carrying rare genetic variants of lysosomal trafficking regulator ( LYST ) were less refractory to TLR stimulation and exhibited lysosomal and mitochondrial abnormalities with altered energy metabolism. Our results strongly suggest that MIS-C hyperinflammation and/or excessive or prolonged stimulation with gut-originated TLR ligands drive immune cells to a lasting refractory state. TLR hyporesponsiveness is likely beneficial, as suggested by excess lymphopenia among rare LYST variant carriers. Our findings point to cellular mechanisms underlying TLR hyporesponsiveness; identify genetic determinants that may explain the MIS-C clinical spectrum; suggest potential associations between innate refractory states and long COVID; and highlight the need to monitor long-term consequences of MIS-C.
- Published
- 2024
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5. Prevalence and Correlates of SARS CoV-2 Among a Community-Based Sample Recruited Using Randomized Venue-Based Sampling. Essex County, NJ, 2020.
- Author
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Raymond HF, Datta P, Ukey R, Wang P, Martino RJ, Krause KD, Rosmarin-DeStefano C, Pinter A, Halkitis PN, and Gennaro ML
- Subjects
- Humans, Prevalence, New Jersey epidemiology, Patient Selection, Antibodies, Viral, SARS-CoV-2, COVID-19 epidemiology
- Abstract
Disparities in morbidity and mortality related to COVID-19 based on race and ethnicity have been documented in the USA. However, it is unclear if these disparities also exist at the exposure stage. To determine this, studies are needed to document the underlying burden of disease, potential disparities through serologic surveillance. Additionally, such studies can help identify where along the disease spectrum (e.g., exposure, infection, diagnosis, treatment, death) and with regard to the structural factors that necessitate public health and/or clinical interventions. Our objectives in this study were to estimate the true burden of SARS CoV-2 in the community of Essex County, NJ, an early and hard hit area, to determine the correlates of SARS CoV-2 prevalence and to determine if COVID-19 disparities seen by race/ethnicity were also reflected in SARS CoV-2 burden. We utilized venue-based-sampling (VBS) to sample members of the community in Essex County. Participants completed a short electronic survey and provided finger stick blood samples for testing. We sampled 924 residents of Essex County, New Jersey. Testing conducted in this study identified 83 (9.0%) participants as positive for SARS-CoV-2 antibodies. Importantly, our findings suggest that the true burden of SARS-Cov-2 and the pool of persons potentially spreading the virus are slightly more than six times than that suggested by PCR testing Notably, there were no significant differences in odds of testing positive for SARS CoV-2 antibodies in terms of race/ethnicity where we compared Black and Latinx participants to other race participants. Our study suggests that disparities in COVID-19 outcomes stem from potential upstream issues such as underlying conditions, access to testing, and access to care rather than disparities in exposure to the virus., (© 2021. W. Montague Cobb-NMA Health Institute.)
- Published
- 2022
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6. Longitudinal Analysis of Biologic Correlates of COVID-19 Resolution: Case Report.
- Author
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Bruiners N, Guerrini V, Ukey R, Dikdan RJ, Yang JH, Mishra PK, Onyuka A, Handler D, Vieth J, Carayannopoulos M, Guo S, Pollen M, Pinter A, Tyagi S, Feingold D, Philipp C, Libutti SK, and Gennaro ML
- Abstract
While the biomarkers of COVID-19 severity have been thoroughly investigated, the key biological dynamics associated with COVID-19 resolution are still insufficiently understood. We report a case of full resolution of severe COVID-19 due to convalescent plasma transfusion. Following transfusion, the patient showed fever remission, improved respiratory status, and rapidly decreased viral burden in respiratory fluids and SARS-CoV-2 RNAemia. Longitudinal unbiased proteomic analysis of plasma and single-cell transcriptomics of peripheral blood cells conducted prior to and at multiple times after convalescent plasma transfusion identified the key biological processes associated with the transition from severe disease to disease-free state. These included (i) temporally ordered upward and downward changes in plasma proteins reestablishing homeostasis and (ii) post-transfusion disappearance of a subset of monocytes characterized by hyperactivated Interferon responses and decreased TNF-α signaling. Monitoring specific dysfunctional myeloid cell subsets in peripheral blood may provide prognostic keys in COVID-19., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Bruiners, Guerrini, Ukey, Dikdan, Yang, Mishra, Onyuka, Handler, Vieth, Carayannopoulos, Guo, Pollen, Pinter, Tyagi, Feingold, Philipp, Libutti and Gennaro.)
- Published
- 2022
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7. Biologic correlates of beneficial convalescent plasma therapy in a COVID-19 patient reveal disease resolution mechanisms.
- Author
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Bruiners N, Guerrini V, Ukey R, Dikdan R, Yang J, Mishra PK, Onyuka A, Handler D, Vieth J, Carayannopulos M, Guo S, Pollen M, Pinter A, Tyagi S, Feingold D, Philipp C, Libutti S, and Gennaro ML
- Abstract
Background: While the biomarkers of COVID-19 severity have been thoroughly investigated, the key biological dynamics associated with COVID-19 resolution are still insufficiently understood., Main Body: We report a case of full resolution of severe COVID-19 due to convalescent plasma transfusion in a patient with underlying multiple autoimmune syndrome. Following transfusion, the patient showed fever remission, improved respiratory status, and rapidly decreased viral burden in respiratory fluids and SARS-CoV-2 RNAemia. Longitudinal unbiased proteomic analysis of plasma and single-cell transcriptomics of peripheral blood cells conducted prior to and at multiple times after convalescent plasma transfusion identified the key biological processes associated with the transition from severe disease to disease-free state. These included (i) temporally ordered upward and downward changes in plasma proteins reestablishing homeostasis and (ii) post-transfusion disappearance of a particular subset of dysfunctional monocytes characterized by hyperactivated Interferon responses and decreased TNF-α signaling., Conclusions: Monitoring specific subsets of innate immune cells in peripheral blood may provide prognostic keys in severe COVID-19. Moreover, understanding disease resolution at the molecular and cellular level should contribute to identify targets of therapeutic interventions against severe COVID-19.
- Published
- 2022
- Full Text
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8. Dichotomy between the humoral and cellular responses elicited by mRNA and adenoviral vector vaccines against SARS-CoV-2.
- Author
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Ukey R, Bruiners N, Mishra H, Mishra PK, McCloskey D, Onyuka A, Chen F, Pinter A, Weiskopf D, Sette A, Roy J, Gaur S, and Gennaro ML
- Subjects
- Ad26COVS1, Antibodies, Viral, Humans, Immunity, Humoral, RNA, Messenger genetics, SARS-CoV-2, Vaccination, mRNA Vaccines, COVID-19, COVID-19 Vaccines
- Abstract
Background: Protection from severe disease and hospitalization by SARS-CoV-2 vaccination has been amply demonstrated by real-world data. However, the rapidly evolving pandemic raises new concerns. One pertains efficacy of adenoviral vector-based vaccines, particularly the single-dose Ad26.COV2.S, relative to mRNA vaccines., Main Body: We investigated the immunogenicity of Ad26.COV2.S and mRNA vaccines in 33 subjects vaccinated with either vaccine class 5 months earlier on average. After controlling for the time since vaccination, Spike-binding antibody and neutralizing antibody levels were higher in the mRNA-vaccinated subjects, while no significant differences in antigen-specific B cell and T cell responses were observed between the two groups., Conclusions: A dichotomy exists between the humoral and cellular responses elicited by the two vaccine classes. Testing only for humoral responses to compare the durability of SARS-CoV-2 vaccine-induced responses, as typically performed for public health and research purposes, is insufficient., (© 2022. The Author(s).)
- Published
- 2022
- Full Text
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9. Highly versatile antibody binding assay for the detection of SARS-CoV-2 infection and vaccination.
- Author
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Datta P, Ukey R, Bruiners N, Honnen W, Carayannopoulos MO, Reichman C, Choudhary A, Onyuka A, Handler D, Guerrini V, Mishra PK, Dewald HK, Lardizabal A, Lederer L, Leiser AL, Hussain S, Jagpal SK, Radbel J, Bhowmick T, Horton DB, Barrett ES, Xie YL, Fitzgerald-Bocarsly P, Weiss SH, Woortman M, Parmar H, Roy J, Dominguez-Bello MG, Blaser MJ, Carson JL, Panettieri RA Jr, Libutti SK, Raymond HF, Pinter A, and Gennaro ML
- Subjects
- Antibodies, Viral immunology, Binding Sites, COVID-19 blood, COVID-19 immunology, Humans, Vaccination, Antibodies, Viral blood, COVID-19 diagnosis, Dried Blood Spot Testing
- Abstract
Monitoring the burden and spread of infection with the new coronavirus SARS-CoV-2, whether within small communities or in large geographical settings, is of paramount importance for public health purposes. Serology, which detects the host antibody response to the infection, is the most appropriate tool for this task, since virus-derived markers are most reliably detected during the acute phase of infection. Here we show that our ELISA protocol, which is based on antibody binding to the Receptor Binding Domain (RBD) of the S1 subunit of the viral Spike protein expressed as a novel fusion protein, detects antibody responses to SARS-CoV-2 infection and vaccination. We also show that our ELISA is accurate and versatile. It compares favorably with commercial assays widely used in clinical practice to determine exposure to SARS-CoV-2. Moreover, our protocol accommodates use of various blood- and non-blood-derived biospecimens, such as breast milk, as well as dried blood obtained with microsampling cartridges that are appropriate for remote collection. As a result, our RBD-based ELISA protocols are well suited for seroepidemiology and other large-scale studies requiring parsimonious sample collection outside of healthcare settings., (Copyright © 2021. Published by Elsevier B.V.)
- Published
- 2021
- Full Text
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10. Determinants and Dynamics of SARS-CoV-2 Infection in a Diverse Population: 6-Month Evaluation of a Prospective Cohort Study.
- Author
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Horton DB, Barrett ES, Roy J, Gennaro ML, Andrews T, Greenberg P, Bruiners N, Datta P, Ukey R, Velusamy SK, Fine D, Honnen WJ, Yin YS, Pinter A, Brooks A, Tischfield J, Hussain S, Jagpal S, Swaminathan S, Parmar V, Reilly N, Gaur S, Panettieri RA, Carson JL, and Blaser MJ
- Subjects
- Adult, Antibodies, Viral immunology, Asymptomatic Infections epidemiology, COVID-19 blood, COVID-19 epidemiology, COVID-19 transmission, Comorbidity, Female, Humans, Immunoglobulin G immunology, Incidence, Male, Middle Aged, Prospective Studies, Risk Factors, SARS-CoV-2 immunology, Young Adult, Antibodies, Viral blood, COVID-19 diagnosis, Immunoglobulin G blood, SARS-CoV-2 isolation & purification, Severity of Illness Index
- Abstract
Background: We studied risk factors, antibodies, and symptoms of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in a diverse, ambulatory population., Methods: A prospective cohort (n = 831) previously undiagnosed with SARS-CoV-2 infection underwent serial testing (SARS-CoV-2 polymerase chain reaction, immunoglobulin G [IgG]) for 6 months., Results: Ninety-three participants (11.2%) tested SARS-CoV-2-positive: 14 (15.1%) asymptomatic, 24 (25.8%) severely symptomatic. Healthcare workers (n = 548) were more likely to become infected (14.2% vs 5.3%; adjusted odds ratio, 2.1; 95% confidence interval, 1.4-3.3) and severely symptomatic (29.5% vs 6.7%). IgG antibodies were detected after 79% of asymptomatic infections, 89% with mild-moderate symptoms, and 96% with severe symptoms. IgG trajectories after asymptomatic infections (slow increases) differed from symptomatic infections (early peaks within 2 months). Most participants (92%) had persistent IgG responses (median 171 days). In multivariable models, IgG titers were positively associated with symptom severity, certain comorbidities, and hospital work. Dyspnea and neurologic changes (including altered smell/taste) lasted ≥ 120 days in ≥ 10% of affected participants. Prolonged symptoms (frequently more severe) corresponded to higher antibody levels., Conclusions: In a prospective, ethnically diverse cohort, symptom severity correlated with the magnitude and trajectory of IgG production. Symptoms frequently persisted for many months after infection.Clinical Trials Registration. NCT04336215., (© The Author(s) 2021. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)
- Published
- 2021
- Full Text
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11. Dichotomy between the humoral and cellular responses elicited by mRNA and adenoviral vector vaccines against SARS-CoV-2.
- Author
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Ukey R, Bruiners N, Mishra H, Mishra PK, McCloskey D, Onyuka A, Chen F, Pinter A, Weiskopf D, Sette A, Roy J, Gaur S, and Gennaro ML
- Abstract
Protection from severe disease and hospitalization by SARS-CoV-2 vaccination has been amply demonstrated by real-world data. However, the rapidly evolving pandemic raises new concerns. One pertains efficacy of adenoviral vector-based vaccines, particularly the single-dose Ad26.COV2.S, relative to mRNA vaccines. We investigated the immunogenicity of Ad26.COV2.S and mRNA vaccines in 33 subjects vaccinated with either vaccine class five months earlier on average. After controlling for time since vaccination, Spike-binding antibody and neutralizing antibody levels were higher in the mRNA-vaccinated subjects, while no significant differences in antigen-specific B cell and T cell responses were observed between the two groups. Thus, a dichotomy exists between humoral and cellular responses elicited by the two vaccine classes. Our results have implications for the need of booster doses in vaccinated subjects and might explain the dichotomy reported between the waning protection from symptomatic infection by SARS-CoV-2 vaccination and its persisting efficacy in preventing hospitalization and death.
- Published
- 2021
- Full Text
- View/download PDF
12. Highly versatile antibody binding assay for the detection of SARS-CoV-2 infection.
- Author
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Datta P, Ukey R, Bruiners N, Honnen W, Carayannopoulos MO, Reichman C, Choudhary A, Onyuka A, Handler D, Guerrini V, Mishra PK, Dewald HK, Lardizabal A, Lederer L, Leiser AL, Hussain S, Jagpal SK, Radbel J, Bhowmick T, Horton DB, Barrett ES, Xie YL, Fitzgerald-Bocarsly P, Weiss SH, Woortman M, Parmar H, Roy J, Dominguez-Bello MG, Blaser MJ, Carson JL, Panettieri RA Jr, Libutti SK, Raymond HF, Pinter A, and Gennaro ML
- Abstract
Monitoring the burden and spread of infection with the new coronavirus SARS-CoV-2, whether within small communities or in large geographical settings, is of paramount importance for public health purposes. Serology, which detects the host antibody response to the infection, is the most appropriate tool for this task, since virus-derived markers are most reliably detected during the acute phase of infection. Here we show that our ELISA protocol, which is based on antibody binding to the Receptor Binding Domain (RBD) of the S1 subunit of the viral Spike protein expressed as a novel fusion protein, detects antibody responses to SARS-CoV-2 infection and COVID-19 vaccination. We also show that our ELISA is accurate and versatile. It compares favorably with commercial assays widely used in clinical practice to determine exposure to SARS-CoV-2. Moreover, our protocol accommodates use of various blood- and non-blood-derived biospecimens, such as breast milk, as well as dried blood obtained with microsampling cartridges that are appropriate for remote collection. As a result, our RBD-based ELISA protocols are well suited for seroepidemiology and other large-scale studies requiring parsimonious sample collection outside of healthcare settings.
- Published
- 2021
- Full Text
- View/download PDF
13. Vaccination boosts protective responses and counters SARS-CoV-2-induced pathogenic memory B cells.
- Author
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Mishra PK, Bruiners N, Ukey R, Datta P, Onyuka A, Handler D, Hussain S, Honnen W, Singh S, Guerrini V, Yin Y, Dewald H, Choudhary A, Horton DB, Barrett ES, Roy J, Weiss SH, Fitzgerald-Bocarsly P, Blaser MJ, Carson JL, Panettieri RA Jr, Lardizabal A, Chang TL, Pinter A, and Gennaro ML
- Abstract
Much is to be learned about the interface between immune responses to SARS-CoV-2 infection and vaccination. We monitored immune responses specific to SARS-CoV-2 Spike Receptor-Binding-Domain (RBD) in convalescent individuals for eight months after infection diagnosis and following vaccination. Over time, neutralizing antibody responses, which are predominantly RBD specific, generally decreased, while RBD-specific memory B cells persisted. RBD-specific antibody and B cell responses to vaccination were more vigorous than those elicited by infection in the same subjects or by vaccination in infection-naïve comparators. Notably, the frequencies of double negative B memory cells, which are dysfunctional and potentially pathogenic, increased in the convalescent subjects over time. Unexpectedly, this effect was reversed by vaccination. Our work identifies a novel aspect of immune dysfunction in mild/moderate COVID-19, supports the practice of offering SARS-CoV-2 vaccination regardless of infection history, and provides a potential mechanistic explanation for the vaccination-induced reduction of "Long-COVID" symptoms.
- Published
- 2021
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14. The Yarrowia lipolytica PAH1 homologue contributes but is not required for triacylglycerol biosynthesis during growth on glucose.
- Author
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Ukey R, Carmon T, Hardman D, Hill N, and Fakas S
- Subjects
- Fungal Proteins genetics, Mutation, Phosphatidate Phosphatase metabolism, Yarrowia genetics, Fungal Proteins metabolism, Gene Expression Regulation, Fungal, Glucose metabolism, Triglycerides biosynthesis, Yarrowia metabolism
- Abstract
The PAH1-encoded phosphatidate phosphatase (PAP) catalyzes the Mg
2+ -dependent dephosphorylation of phosphatidate to produce diacylglycerol, which can be acylated to form triacylglycerol (TAG). In the model oleaginous yeast Yarrowia lipolytica, TAG is the major lipid produced, and its biosynthesis requires a continuous supply of diacylglycerol, which can be provided by the PAP reaction. However, the regulation of Pah1 has not been studied in detail in Y. lipolytica, and thus its contribution to the biosynthesis of TAG in this yeast is not well understood. In this work, we examined the regulation of the PAH1-mediated PAP activity and Pah1 abundance and localization in cells growing on glucose. We found that Pah1 abundance and localization were regulated in a growth-dependent manner, yet the loss of Pah1 did not have a major effect on PAP activity. We also examined the effects of the Y. lipolytica pah1Δ mutation on cell physiology and lipid biosynthesis. The lack of Pah1 in the pah1Δ mutant resulted in a moderate decrease in TAG levels and an increase in phospholipid levels. These results showed that Pah1 contributed to TAG biosynthesis in Y. lipolytica but also suggested the presence of other activities in the pah1Δ mutant that compensate for the loss of Pah1. Also, the levels of linoleic acid were elevated in pah1Δ cells with a concomitant decrease in the oleic acid levels suggesting that the pah1Δ mutation affected the biosynthesis of fatty acids., (© 2019 John Wiley & Sons, Ltd.)- Published
- 2020
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15. Phosphatidate phosphatase activity is induced during lipogenesis in the oleaginous yeast Yarrowia lipolytica.
- Author
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Hardman D, Ukey R, and Fakas S
- Subjects
- Gene Expression Regulation, Fungal, Glucose, Lipogenesis genetics, Phosphatidate Phosphatase metabolism, Triglycerides biosynthesis, Yarrowia enzymology
- Abstract
Phosphatidate (PA) phosphatase dephosphorylates the membrane phospholipid PA to diacylglycerol (DAG) that can be used for the synthesis of the storage lipid triacylglycerol (TAG). In Yarrowia lipolytica, TAG biosynthesis is induced during the lipogenic phase, which results in the accumulation of this lipid in cells. The accumulation of TAG during lipogenesis requires the supply of DAG, but the source of this DAG is not known in Y. lipolytica. In this study, the regulation of PA phosphatase during lipogenesis and its contribution to TAG biosynthesis was examined in Y. lipolytica. Lipogenesis was triggered by growing cells in high-glucose media, whereas control cultures were grown in low-glucose media. PA phosphatase activity increased in a time-dependent manner as high-glucose cells progressed in the lipogenic phase. In contrast, the activity decreased in low-glucose cells that did not accumulate lipids. An analysis of the subcellular localization of the PA phosphatase activity showed that the membrane-associated activity increased during lipogenesis. The significance of this increase can be explained by the fact that only the membrane-associated PA phosphatase activity is responsible for the production of DAG. Taken together, these results indicate that PA phosphatase is involved in TAG biosynthesis during lipogenesis in Y. lipolytica., (© 2018 John Wiley & Sons, Ltd.)
- Published
- 2018
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16. Evaluation of thioesterases from Acinetobacter baylyi for production of free fatty acids.
- Author
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Ukey R, Holmes WE, Bajpai R, and Chistoserdov AY
- Subjects
- Biofuels, Escherichia coli genetics, Acinetobacter metabolism, Fatty Acids, Nonesterified biosynthesis, Thiolester Hydrolases genetics
- Abstract
Acinetobacter baylyi is one of few Gram-negative bacteria capable of accumulating storage lipids in the form of triacylglycerides and wax esters, which makes it an attractive candidate for production of lipophilic products, including biofuel precursors. Thioesterases play a significant dual role in the triacylglyceride and wax ester biosynthesis by either providing or removing acyl-CoA from this pathway. Therefore, 4 different thioesterase genes were cloned from Acinetobacter baylyi ADP1 and expressed in Escherichia coli to investigate their contribution to free fatty acids (FFAs) accumulation. Overexpression of the genes tesA' (a leaderless form of the gene tesA) and tesC resulted in increased accumulation of FFAs when compared with the host E. coli strain. Overexpression of tesA' showed a 1.87-fold increase in production of long-chain fatty acids (C16 to C18) over the host strain. Unlike TesC and the other investigated thioesterases, the TesA' thioesterase also produced shorter chain FFAs (e.g., myristic acid) and unsaturated FFAs (e.g., cis-vaccenic acid (18:1Δ11)). A comparison of the remaining 3 A. baylyi ADP1 thioesterases (encoded by the tesB, tesC, and tesD genes) revealed that only the strain containing the tesC gene produced statistically higher levels of FFAs over the control, suggesting that it possesses the acyl-ACP thioesterase activity. Both E. coli strains containing the tesB and tesD genes produced levels of FFAs similar to those of the plasmid-free control E. coli strain, which indicates that TesB and TesD lack the acyl-ACP thioesterase activity.
- Published
- 2017
- Full Text
- View/download PDF
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