The majority of SARS-CoV-2-specific antibodies in COVID-19 patients with obesity are autoimmune and not neutralizing.

BACKGROUND/OBJECTIVES: Obesity decreases the secretion of SARS-CoV-2-specific IgG antibodies in the blood of COVID-19 patients. How obesity impacts the quality of the antibodies secreted, however, is not understood. Therefore, the objective of this study is to evaluate the presence of neutralizing versus autoimmune antibodies in COVID-19 patients with obesity. SUBJECTS/METHODS: Thirty serum samples from individuals who tested positive for SARS-CoV-2 infection by RT-PCR were collected from inpatient and outpatient settings. Of these, 15 were lean (BMI < 25) and 15 were obese (BMI ≥ 30). Control serum samples were from 30 uninfected individuals, age-, gender-, and BMI-matched, recruited before the current pandemic. Neutralizing and autoimmune antibodies were measured by ELISA. IgG autoimmune antibodies were specific for malondialdehyde (MDA), a marker of oxidative stress and lipid peroxidation, and for adipocyte-derived protein antigens (AD), markers of virus-induced cell death in the obese adipose tissue. RESULTS: SARS-CoV-2 infection induces neutralizing antibodies in all lean but only in few obese COVID-19 patients. SARS-CoV-2 infection also induces anti-MDA and anti-AD autoimmune antibodies more in lean than in obese patients as compared to uninfected controls. Serum levels of these autoimmune antibodies, however, are always higher in obese versus lean COVID-19 patients. Moreover, because the autoimmune antibodies found in serum samples of COVID-19 patients have been correlated with serum levels of C-reactive protein (CRP), a general marker of inflammation, we also evaluated the association of anti-MDA and anti-AD antibodies with serum CRP and found a positive association between CRP and autoimmune antibodies. CONCLUSIONS: Our results highlight the importance of evaluating the quality of the antibody response in COVID-19 patients with obesity, particularly the presence of autoimmune antibodies, and identify biomarkers of self-tolerance breakdown. This is crucial to protect this vulnerable population at higher risk of responding poorly to infection with SARS-CoV-2 than lean controls.

DETECTION OF GLIOTOXIN BUT NOT BIS(METHYL)GLIOTOXIN IN PLASMA FROM BIRDS WITH CONFIRMED AND PROBABLE ASPERGILLOSIS.

Aspergillosis remains a difficult disease to diagnose antemortem in many species, especially avian species. In the present study, banked plasma samples from various avian species were examined for gliotoxin (GT), which is a recognized key virulence factor produced during the replication of Aspergillus species hyphae and a secondary metabolite bis(methyl)gliotoxin (bmGT). Initially, liquid chromatography-tandem mass spectrometry methods for detecting GT and bmGT were validated in a controlled model using sera obtained from rats experimentally infected with Aspergillus fumigatus. The minimum detection level for both measurements was determined to be 3 ng/ml, and the assay was found to be accurate and reliable. As proof of concept, GT was detected in 85.7% (30/35) of the samples obtained from birds with confirmed aspergillosis and in 60.7% (17/28) of samples from birds with probable infection but only in one of those from clinically normal birds (1/119). None of the birds were positive for bmGT. Repeated measures from birds under treatment suggests results may have prognostic value. Further studies are needed to implement quantitative methods and to determine the utility of this test in surveillance screening in addition to its use as a diagnostic test in birds with suspected aspergillosis.

SARS-CoV-2 infection induces autoimmune antibody secretion more in lean than in obese COVID-19 patients.

BACKGROUND/OBJECTIVES: Obesity decreases the secretion of SARS-CoV-2-specific IgG antibodies in the blood of COVID-19 patients. How obesity impacts the secretion of autoimmune antibodies in COVID-19 patients, however, is not understood. The serum of adult COVID-19 patients contains autoimmune antibodies generated in response to virus-induced tissue damage and cell death leading to the release of intracellular antigens not known to be immunogenic autoantigens. The objective of this study is to evaluate the presence of autoimmune antibodies in COVID-19 patients with obesity. SUBJECTS/METHODS: Thirty serum samples from individuals who tested positive for SARS-CoV-2 infection by RT-PCR were collected from inpatient and outpatient settings. Of these, 15 were lean (BMI<25), and 15 were obese (BMI ≥30). Control serum samples were from 30 uninfected individuals, age-gender- and BMI-matched, recruited before the current pandemic. Serum IgG antibodies against two autoimmune specificities, as well as against SARS-CoV-2 Spike protein, were measured by ELISA. IgG autoimmune antibodies were specific for malondialdehyde (MDA), a marker of oxidative stress and lipid peroxidation, and for adipocyte-derived protein antigens (AD), markers of virus-induced cell death in the obese AT. RESULTS: Our results show that SARS-CoV-2 infection induces anti-MDA and anti-AD autoimmune antibodies more in lean than in obese patients as compared to uninfected controls. Serum levels of these autoimmune antibodies, however, are always higher in obese versus lean COVID-19 patients. Moreover, because the autoimmune antibodies found in serum samples of COVID-19 patients have been correlated with serum levels of C-reactive protein (CRP), a general marker of inflammation, we also evaluated the association of anti-MDA and anti-AT antibodies with serum CRP and found a significant association between CRP and autoimmune antibodies in our cohort of lean and obese COVID-19 patients. CONCLUSIONS: Our results highlight the importance of evaluating the quality of the antibody response in COVID-19 patients with obesity, particularly the presence of autoimmune antibodies, and identify biomarkers of self-tolerance breakdown. This is crucial to protect this vulnerable population that is at higher risk of responding poorly to infection with SARS-CoV-2 compared to lean controls.

Influence of obesity on serum levels of SARS-CoV-2-specific antibodies in COVID-19 patients.

SARS-CoV-2 (Severe Acute Respiratory Syndrome Corona Virus-2), cause of COVID-19 (Coronavirus Disease of 2019), represents a significant risk to people living with pre-existing conditions associated with exacerbated inflammatory responses and consequent dysfunctional immunity. In this paper, we have evaluated the influence of obesity, a condition associated with chronic systemic inflammation, on the secretion of SARS-CoV-2-specific IgG antibodies in the blood of COVID-19 patients. Our hypothesis is that obesity is associated with reduced amounts of specific IgG antibodies. Results have confirmed our hypothesis and have shown that SARS-CoV-2 IgG antibodies are negatively associated with Body Mass Index (BMI) in COVID-19 obese patients, as expected based on the known influence of obesity on humoral immunity. Antibodies in COVID-19 obese patients are also negatively associated with serum levels of pro-inflammatory and metabolic markers of inflammaging and pulmonary inflammation, such as SAA (serum amyloid A protein), CRP (C-reactive protein), and ferritin, but positively associated with NEFA (nonesterified fatty acids). These results altogether could help to identify an inflammatory signature with strong predictive value for immune dysfunction. Inflammatory markers identified may subsequently be targeted to improve humoral immunity in individuals with obesity and in individuals with other chronic inflammatory conditions.

Effects of obesity on serum levels of SARS-CoV-2-specific antibodies in COVID-19 patients.

SARS-CoV-2 (Severe Acute Respiratory Syndrome Corona Virus-2), cause of COVID-19 (Coronavirus Disease of 2019), represents a significant risk to people living with pre-existing conditions associated with exacerbated inflammatory responses and consequent dysfunctional immunity. In this paper, we have evaluated the effects of obesity, a condition associated with chronic systemic inflammation, on the secretion of SARS-CoV-2-specific IgG antibodies in the blood of COVID-19 patients. Results have shown that SARS-CoV-2 IgG antibodies are negatively associated with Body Mass Index (BMI) in COVID-19 obese patients, as expected based on the known effects of obesity on humoral immunity. Antibodies in COVID-19 obese patients are also negatively associated with serum levels of pro-inflammatory and metabolic markers of inflammaging and pulmonary inflammation, such as SAA (serum amyloid A protein), CRP (C-reactive protein) and ferritin, but positively associated with NEFA (nonesterified fatty acids). These results altogether could help to identify an inflammatory signature with strong predictive value for immune dysfunction that could be targeted to improve humoral immunity in individuals with obesity as well as with other chronic inflammatory conditions.

LC-MS-MS vs ELISA: Validation of a Comprehensive Urine Toxicology Screen by LC-MS-MS and a Comparison of 100 Forensic Specimens.

Toxicology laboratories commonly employ immunoassay methodologies to perform an initial drug screen on urine specimens to direct confirmatory testing. Due to limitations of immunoassay testing and the need to screen for a broader range of drugs with lower limits of detection at a lower cost, mass spectrometry screening techniques have gained favor in the toxicology field. A liquid chromatography-tandem mass spectrometry (LC-MS-MS) urine screening panel was developed and validated for 52 drugs and metabolites. A simple dilute-and-shoot with enzymatic hydrolysis technique was utilized to prepare the urine specimens for analysis. Limit of detection, interference, ionization suppression/enhancement, carryover and stability of processed specimens were assessed during validation. To evaluate the toxicological results obtained from utilizing the LC-MS-MS in comparison with the laboratory's current enzyme-linked immunosorbent assay (ELISA) panel, 100 authentic urine specimens from suspected driving under the influence and drug-facilitated crime cases were analyzed using both methodologies and the results were compared. In addition, the cost of each methodology was evaluated and compared. The validated LC-MS-MS method had limits of detection that were equal to or lower than the concentrations validated for ELISA cutoffs, had fewer exogenous interferences, and the cost of screening per specimen was reduced by ~70% when compared to ELISA. Comparing the toxicology results of forensic urine specimens demonstrated that by only using ELISA, the laboratory was unable to detect benzoylecgonine in 26%, lorazepam in 33% and oxymorphone in 60% of the positive specimens. Additional analytes detected using the LC-MS-MS method were zolpidem and/or metabolite, gabapentin, tramadol and metabolite, methadone and metabolite, meprobamate and phentermine. The results of the validation, the toxicological result comparison and the cost comparison showed that the LC-MS-MS screening method is a simple, sensitive and cost-effective alternative to ELISA screening methods for urine specimens.