Biochemical and structural impact of two novel missense mutations in cystathionine ß-synthase gene associated with homocystinuria.

Homocystinuria is a rare disease caused by mutations in the CBS gene that results in a deficiency of cystathionine ß-synthase (CBS). CBS is an essential pyridoxal 5'-phosphate (PLP)-dependent enzyme in the transsulfuration pathway, responsible for combining serine with homocysteine to produce cystathionine, whose activity is enhanced by the allosteric regulator S-adenosylmethionine (SAM). CBS also plays a role in generating hydrogen sulfide (H2S), a gaseous signaling molecule with diverse regulatory functions within the vascular, nervous, and immune systems. In this study, we present the clinical and biochemical characterization of two novel CBS missense mutations that do not respond to pyridoxine treatment, namely c.689T > A (L230Q) and 215A > T (K72I), identified in a Chinese patient. We observed that the disease-associated K72I genetic variant had no apparent effects on the spectroscopic and catalytic properties of the full-length enzyme. In contrast, the L230Q variant expressed in Escherichia coli did not fully retain heme and when compared with the wild-type enzyme, it exhibited more significant impairments in both the canonical cystathionine-synthesis and the alternative H2S-producing reactions. This reduced activity is consistent with both in vitro and in silico evidence, which indicates that the L230Q mutation significantly decreases the overall protein's stability, which in turn, may represent the underlying cause of its pathogenicity.

SARS-CoV-2 infection triggers more potent antibody-dependent cellular cytotoxicity (ADCC) responses than mRNA-, vector-, and inactivated virus-based COVID-19 vaccines.

Neutralizing antibodies (NAbs) are elicited after infection and vaccination and have been well studied. However, their antibody-dependent cellular cytotoxicity (ADCC) functionality is still poorly characterized. Here, we investigated ADCC activity in convalescent sera from infected patients with wild-type (WT) severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) or omicron variant compared with three coronavirus disease 2019 (COVID-19) vaccine platforms and postvaccination breakthrough infection (BTI). We analyzed ADCC activity targeting SARS-CoV-2 spike (S) and nucleocapsid (N) proteins in convalescent sera following WT SARS-CoV-2-infection (n = 91), including symptomatic and asymptomatic infections, omicron-infection (n = 8), COVID-19 vaccination with messenger RNA- (mRNA)- (BNT162b2 or mRNA-1273, n = 77), adenovirus vector- (n = 41), and inactivated virus- (n = 46) based vaccines, as well as post-mRNA vaccination BTI caused by omicron (n = 28). Correlations between ADCC, binding, and NAb titers were reported. ADCC was elicited within the first month postinfection and -vaccination and remained detectable for ≥3 months. WT-infected symptomatic patients had higher S-specific ADCC levels than asymptomatic and vaccinated individuals. Also, no difference in N-specific ADCC activity was seen between symptomatic and asymptomatic patients, but the levels were higher than the inactivated vaccine. Notably, omicron infection showed reduced overall ADCC activity compared to WT SARS-CoV-2 infection. Although post-mRNA vaccination BTI elicited high levels of binding and NAbs, ADCC activity was significantly reduced. Also, there was no difference in ADCC levels across the four vaccines, although NAbs and binding antibody titers were significantly higher in mRNA-vaccinated individuals. All evaluated vaccine platforms are inferior in inducing ADCC compared to natural infection with WT SARS-CoV-2. The inactivated virus-based vaccine can induce N-specific ADCC activity, but its relevance to clinical outcomes requires further investigation. Our data suggest that ADCC could be used to estimate the extra-neutralization level against COVID-19 and provides evidence that vaccination should focus on other Fc-effector functions besides NAbs. Also, the decreased susceptibility of the omicron variant to ADCC offers valuable guidance for forthcoming efforts to identify the specific targets of antibodies facilitating ADCC.

Seroprevalence of herpes simplex virus type 1 and type 2 among the migrant workers in Qatar.

BACKGROUND: Limited data exists on herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2) infections in migrant populations. This study investigated HSV-1 and HSV-2 seroprevalences and associations among craft and manual workers (CMWs) in Qatar who constitute 60% of Qatar's population. METHODS: A national population-based cross-sectional seroprevalence survey was conducted on the CMW population, all men, between July 26 and September 9, 2020. 2,612 sera were tested for anti-HSV-1 IgG antibodies using HerpeSelect 1 ELISA IgG kits and for anti-HSV-2 IgG antibodies using HerpeSelect 2 ELISA IgG kits (Focus Diagnostics, USA). Univariable and multivariable logistic regression analyses were conducted to identify associations with HSV-1 and HSV-2 infections. RESULTS: Serological testing identified 2,171 sera as positive, 403 as negative, and 38 as equivocal for HSV-1 antibodies, and 300 sera as positive, 2,250 as negative, and 62 as equivocal for HSV-2 antibodies. HSV-1 and HSV-2 seroprevalences among CMWs were estimated at 84.2% (95% CI 82.8-85.6%) and 11.4% (95% CI 10.1-12.6%), respectively. HSV-1 infection was associated with nationality, educational attainment, and occupation. HSV-2 infection was associated with age, nationality, and educational attainment. CONCLUSIONS: Over 80% of CMWs are infected with HSV-1 and over 10% are infected with HSV-2. The findings highlight the need for sexual health programs to tackle sexually transmitted infections among the CMW population.

Low Risk of Serological Cross-Reactivity between the Dengue Virus and SARS-CoV-2-IgG Antibodies Using Advanced Detection Assays.

Several studies have reported serological cross-reactivity of the immune responses between SARS-CoV-2 and DENV. Most of the available studies are based on the point-of-care rapid testing kits. However, some rapid test kits have low specificity and can generate false positives. Hence, we aimed to investigate the potential serological cross-reactivity between SARS-CoV-2 and DENV-IgG antibodies using advanced assays including chemiluminescence immunoassay (CLIA) and enzyme-linked immunosorbent assay (ELISA) test. A total of 90 DENV-IgG-ELISA-positive and 90 DENV-IgG-ELISA-negative prepandemic sera were tested for anti-SARS-CoV-2-IgG using the automated CL-900i CLIA assay. Furthermore, a total of 91 SARS-CoV-2-IgG-CLIA-positive and 91 SARS-CoV-2-IgG-CLIA-negative postpandemic sera were tested for anti-DENV-IgG using the NovaLisa ELISA kit. The DENV-IgG-positive sera resulted in five positives and 85 negatives for SARS-CoV-2-IgG. Similarly, the DENV-IgG-negative sera also resulted in 5 positives and 85 negatives for SARS-CoV-2-IgG. No statistically significant difference in specificity between the DENV-IgG-positive and DENV-IgG-negative sera was found (p value = 1.00). The SARS-CoV-2-IgG-positive sera displayed 43 positives, 47 negatives, and 1 equivocal for DENV-IgG, whereas the SARS-CoV-2-IgG-negative sera resulted in 50 positives, 40 negatives, and 1 equivocal for DENV-IgG. No statistically significant difference in the proportion that is DENV-IgG positive between the SARS-CoV-2-IgG-positive and SARS-CoV-2-IgG-negative sera (p value = 0.58). In conclusion, there is a low risk of serological cross-reactivity between the DENV and SARS-CoV-2-IgG antibodies when using advanced detection assays.

The prevalence of HEV among non-A-C hepatitis in Qatar and efficiency of serological markers for the diagnosis of hepatitis E.

BACKGROUND: The rapid growth of Qatar in the last two decades has attracted a large influx of immigrant workers who mostly come from HEV-hyperendemic countries. Thus, we aim to investigate the prevalence of HEV among acute non-A-C hepatitis patients in Qatar; and to evaluate the performance of four dominant commercial serological assays for HEV diagnosis. METHODS: 259 patients with non-A-C hepatitis were tested using the Wantai HEV-IgM, HEV-IgG, HEV-Ag ELISA kits, and the MP Biomedical HEV-Total Ab ELISA kit. ALT levels were tested and HEV RNA (viral loads) was performed using Taqman AmpliCube HEV RT-PCR kit (Mikrogen, Neuried, Germany). The performance of each kit was assessed according to the RT-PCR results. RESULTS: HEV-RNA was detected in 23.1% of the samples. Most of these HEV-RNA-positive cases belonged to non-Qatari residents from the Indian subcontinent; India, Pakistan, etc. HEV-Ag, HEV-IgM, HEV-IgG, HEV-Total Ab were detected in 5.56%, 8.65%, 32.1%, and 34.2% of all tested samples, respectively. Elevated ALT levels were highly correlated with the HEV-Ag, HEV-IgM, HEV-RNA but not with the HEV-IgG and HEV-Total Ab. Although HEV-Ag was very specific (100%), yet its sensitivity was poor (36.7%). HEV-IgM demonstrated the best second marker for diagnosis of acute HEV after RT-PCR as jugged by the overall performance parameters: specificity (96.2%), sensitivity (71.4%), PPV (83.3%), NPP (92.7%), agreement with RT-PCR (91.0%), and Kappa-value (0.71). CONCLUSION: Our study demonstrated a high prevalence of HEV virus in Qatar, mostly among immigrants from the Indian subcontinent. The HEV-IgM represents the best marker for detecting the acute HEV infection, where RT-PCR cannot be performed.

Epidemiology of SARS-CoV2 in Qatar's primary care population aged 10 years and above.

BACKGROUND: There is an urgent need to elucidate the epidemiology of the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV2) and characterize its potential impact. Investing in characterising the SARS-CoV2 will help plan and improve the response to the pandemic. Furthermore, it will help identify the most efficient ways of managing the pandemic, avoiding public health policies and interventions that may be unduly restrictive of normal activity or unnecessarily costly. This paper describes the design and reports findings of a population based epidemiological study undertaken to characterise SARS-CoV2 in Qatar using limited resources in a timely manner. METHODS: Asymptomatic individuals ≥10 years registered with Qatar's publicly funded primary health provider were eligible. A stratified random sampling technique was utilized to identify the study sample. Participants were invited to an appointment where they completed a questionnaire and provided samples for polymerase chain reaction and Immunoglobulin M and G immunoassay tests. Data collected were analyzed to calculate point and period prevalence by sociodemographic, lifestyle and clinical characteristics. RESULTS: Of 18,918 individuals invited for the study, 2084 participated (response rate 10.8%). The overall point prevalence and period prevalence were estimated to be 1.6% (95% CI 1.1-2.2) and 14.6% (95% CI 13.1-16.2) respectively. Period prevalence of SARS-CoV2 infection was not considerably different across age groups (9.7-19.8%). It was higher in males compared to females (16.2 and 12.7% respectively). A significant variation was observed by nationality (7.1 to 22.2%) and municipalities (6.9-35.3%). CONCLUSIONS: The study provides an example of a methodologically robust approach that can be undertaken in a timely manner with limited resources. It reports much-needed epidemiological data about the spread of SARS-CoV2. Given the low prevalence rates, majority of the population in Qatar remains susceptible. Enhanced surveillance must continue to be in place, particularly due to the large number of asymptomatic cases observed. Robust contact tracing and social distancing measures are key to prevent future outbreaks.

Seroprevalence and incidence of hepatitis E virus among blood donors: A review.

Hepatitis E virus (HEV) is an RNA virus with 4 main genotypes. HEV-1 and HEV-2 infect solely humans, while HEV-3 and HEV-4 infect humans and various animals such as pigs, deer, and rabbits. HEV-5 and HEV-6 infect mainly wild boar. Recently, new genotypes, known as HEV-7 and HEV-8, were found to infect camels and humans. HEV is globally distributed into different epidemiological patterns based on socioeconomic factors and ecology. Although HEV is mainly transmitted through the fecal-oral route, it was also recognized as a transfusion-transmitted virus. Transmission through blood donation was documented worldwide with rising annual observations, accounting for more than 2.5% of all transmissions. HEV infection is usually asymptomatic or subclinical in immunocompetent individuals, so it remains questionable whether there is an urgent need to screen for HEV prior to blood transfusion. Moreover, recent studies conducted in the Middle East and North Africa (MENA) region indicate that HEV is highly endemic. Here, we provide a review on HEV epidemiology, transmission, and laboratory diagnosis, giving special emphasis to the newly discovered genotypes, HEV-7 and HEV-8. Furthermore, we underscore the findings of recent HEV seroprevalence and viremia studies among blood donors worldwide. We also shed light on similar studies performed among blood donors in the MENA region.

Pathogenic Homocystinuria-Associated T236N Mutation Dramatically Alters the Biochemical Properties of Cystathionine Beta-Synthase Protein.

BACKGROUND: Cystathione beta-synthase (CBS) T236N is a novel mutation associated with pyridoxine non-responsiveness, which presents a significant difficulty in the medical treatment of homocystinuria. Reported severe phenotypes in homocystinuria patients highlight the urgent requirement to comprehend the molecular mechanisms underlying mutation pathogenicity for the advancement of the disease. METHODOLOGY: In this study, we used a multidisciplinary approach to investigate the molecular properties of bacterially expressed and purified recombinant CBST236N protein, which we directly compared to those of the wild-type (CBSWT) protein. RESULTS: Our data revealed a profound impact of the p.T236N mutation on CBS enzymatic activity, with a dramatic reduction of ~96% compared to the CBSWT protein. Circular dichroism (CD) experiments indicated that the p.T236N mutation did not significantly alter the secondary structure of the protein. However, CD spectra unveiled distinct differences in the thermal stability of CBSWT and CBST236N mutant protein species. In addition, chemical denaturation experiments further highlighted that the CBSWT protein exhibited greater thermodynamic stability than the CBST236N mutant, suggesting a destabilizing effect of this mutation. CONCLUSIONS: Our findings provide an explanation of the pathogenicity of the p.T236N mutation, shedding light on its role in severe homocystinuria phenotypes. This study contributes to a deeper understanding of CBS deficiency and may improve the development of targeted therapeutic strategies for affected individuals.

Comparable antibody levels in heterologous and homologous mRNA COVID-19 vaccination, with superior neutralizing and IgA antibody responses in mRNA homologous boosting.

BACKGROUND: Priming with two doses of AZD1222 (Oxford-AstraZeneca; ChAd) followed by a third mRNA vaccine boosting is considered in several countries, yet comparisons between heterologous and homologous booster efficacy remain unexplored. AIM: To evaluate and contrast the immunogenicity of homologous and heterologous boosting regimens. METHOD: The study examined antibody responses in 1113 subjects, comprising 895 vaccine-naïve individuals across different vaccination strategies (partial, primary series, heterologous booster, homologous booster) and 218 unvaccinated, naturally infected individuals. Assessments included neutralizing total antibodies (NTAbs), total antibodies (TAbs), anti-S-RBD IgG, and anti-S1 IgA levels. RESULTS: The study found mRNA vaccines to exhibit superior immunogenicity in primary series vaccination compared to ChAd, with mRNA-1273 significantly enhancing NTAbs, TAbs, anti-S-RBD IgG, and anti-S1 IgA levels (p < 0.001). Both booster types improved antibody levels beyond primary outcomes, with no significant difference in TAbs and anti-S-RBD IgG levels between regimens. However, homologous mRNA boosters significantly outperformed heterologous boosters in enhancing NTAbs and anti-S1 IgA levels, with the BNT/BNT/BNT regimen yielding particularly higher enhancements (p < 0.05). CONCLUSION: The study concludes that although TAbs and anti-S-RBD IgG antibody levels are similar for both regimens, homologous mRNA boosting outperform heterologous regimen by enhancing anti-S1 IgA and neutralizing antibody levels.

Follow-Up and Comparative Assessment of SARS-CoV-2 IgA, IgG, Neutralizing, and Total Antibody Responses After BNT162b2 or mRNA-1273 Heterologous Booster Vaccination.

BACKGROUND: Priming with ChAdOx1 followed by heterologous boosting is considered in several countries. Nevertheless, analyses comparing the immunogenicity of heterologous booster to homologous primary vaccination regimens and natural infection are lacking. In this study, we aimed to conduct a comparative assessment of the immunogenicity between homologous primary vaccination regimens and heterologous prime-boost vaccination using BNT162b2 or mRNA-1273. METHODS: We matched vaccinated naïve (VN) individuals (n = 673) with partial vaccination (n = 64), primary vaccination (n = 590), and primary series plus mRNA vaccine heterologous booster (n = 19) with unvaccinated naturally infected (NI) individuals with a documented primary SARS-CoV-2 infection (n = 206). We measured the levels of neutralizing total antibodies (NTAbs), total antibodies (TAbs), anti-S-RBD IgG, and anti-S1 IgA titers. RESULTS: Homologous primary vaccination with ChAdOx1 not only showed less potent NTAb, TAb, anti-S-RBD IgG, and anti-S1 IgA immune responses compared to primary BNT162b2 or mRNA-1273 vaccination regimens (p < 0.05) but also showed ~3-fold less anti-S1 IgA response compared to infection-induced immunity (p < 0.001). Nevertheless, a heterologous booster led to an increase of ~12 times in the immune response when compared to two consecutive homologous ChAdOx1 immunizations. Furthermore, correlation analyses revealed that both anti-S-RBD IgG and anti-S1 IgA significantly contributed to virus neutralization among NI individuals, particularly in symptomatic and pauci-symptomatic individuals, whereas among VN individuals, anti-S-RBD IgG was the main contributor to virus neutralization. CONCLUSION: The results emphasize the potential benefit of using heterologous mRNA boosters to increase antibody levels and neutralizing capacity particularly in patients who received primary vaccination with ChAdOx1.

Prevalence of hepatitis B and C viruses among migrant workers in Qatar.

Limited data exist on viral hepatitis among migrant populations. This study investigated the prevalence of current hepatitis B virus (HBV) infection and lifetime hepatitis C virus (HCV) infection among Qatar's migrant craft and manual workers (CMWs), constituting 60% of the country's population. Sera collected during a nationwide COVID-19 population-based cross-sectional survey on CMWs between July 26 and September 9, 2020, underwent testing for HBsAg and HCV antibodies. Reactive samples underwent confirmatory testing, and logistic regression analyses were employed to explore associations with HBV and HCV infections. Among 2528 specimens tested for HBV infection, 15 were reactive, with 8 subsequently confirmed positive. Three samples lacked sufficient sera for confirmatory testing but were included in the analysis through multiple imputations. Prevalence of current HBV infection was 0.4% (95% CI 0.2-0.7%). Educational attainment and occupation were significantly associated with current HBV infection. For HCV infection, out of 2607 specimens tested, 46 were reactive, and 23 were subsequently confirmed positive. Prevalence of lifetime HCV infection was 0.8% (95% CI 0.5-1.2%). Egyptians exhibited the highest prevalence at 6.5% (95% CI 3.1-13.1%), followed by Pakistanis at 3.1% (95% CI 1.1-8.0%). Nationality, geographic location, and occupation were significantly associated with lifetime HCV infection. HBV infection is relatively low among CMWs, while HCV infection falls within the intermediate range, both compared to global and regional levels.

Screening and diagnostic testing protocols for HIV and Syphilis infections in health care setting in Qatar: Evaluation and recommendations.

BACKGROUND: HIV and Syphilis are common STIs, which have become a concern and burden on healthcare systems, as many infections go untreated and lead to potentially serious complications. HIV is usually diagnosed with Western blot, PCR, and p24 antigen testing. Whereas, Syphilis is mainly diagnosed through clinical findings and serologic testing. The Medical Commission Department (MC) under MOPH is responsible for screening all newcomers to Qatar, aiming to keep the country free from serious infectious diseases. OBJECTIVE: We aimed to evaluate the diagnostic efficiency of the protocols used in the MC for screening HIV and Syphilis infections. METHODS: We conducted a retrospective study of samples analyzed by 4th Generation ARCHITECT® HIV Ag/Ab Combo and Rapid Plasma Reagin (RPR) between January to December 2019. ARCHITECT® HIV Ag/Ab Combo positive samples were confirmed by INNO-LIA™ HIVI/II and RT-PCR. RPR-reactive samples were confirmed by ARCHITECT® Syphilis Treponema pallidium Antibody (Syphilis TPA) assay. RESULTS: For HIV, data were collected from 585,587 individuals, of which 595 (0.1%) were positive by the ARCHITECT® HIV Ag/Ab Combo (Analyzer A). When all initially positive sera were re-tested on newly collected blood samples using different ARCHITECT® HIV Ag/Ab Combo analyzer (analyzer B), 99.8% (594/595) of samples were also positive, suggesting high reproducibility. The positive predictive value (PPV) between ARCHITECT® HIV Ag/Ab Combo and the INNO-LIA™ HIVI/II confirmatory assay was 31.8%. The PPV between ARCHITECT® HIV Ag/Ab Combo and HIV-PCR assay was 26.8%. Retrospective data for Syphilis were collected from a total of 97,298 individuals who visited the MC, of which 198 (0.20%) were initially positive by RPR. The PPV between RPR and Syphilis TPA confirmatory assay was 36.6%. CONCLUSION: Despite the high rate of false positivity using ARCHITECT® HIV Ag/Ab Combo and RPR screening assays, both assays have proven to be highly effective as screening testing methods.

Follow up and comparative assessment of IgG, IgA, and neutralizing antibody responses to SARS-CoV-2 between mRNA-vaccinated naïve and unvaccinated naturally infected individuals over 10 months.

BACKGROUND: Evidence on the effectiveness of vaccination-induced immunity compared to SARS-CoV-2 natural immunity is warranted to inform vaccination recommendations. AIM: In this study, we aimed to conduct a comparative assessment of antibody responses between vaccinated naïve (VN) and unvaccinated naturally infected individuals (NI) over 10 Months. METHOD: The study comprised fully-vaccinated naïve individuals (VN; n = 596) who had no history of SARS-CoV-2 infection, and received two doses of either BNT162b2 or mRNA-1273, and naturally infected individuals who had a documented history of SARS-CoV-2 infection and no vaccination record (NI cohort; n = 218). We measured the levels of neutralizing total antibodies (NtAbs), anti-S-RBD IgG, and anti-S1 IgA titers among VN and NI up to ∼10 months from administration of the first dose, and up to ∼7 months from SARS-CoV-2 infection, respectively. To explore the relationship between the antibody responses and time, Spearman's correlation coefficient was computed. Furthermore, correlations between the levels of NtAbs/anti-S-RBD IgG and NtAbs/anti-S1 IgA were examined through pairwise correlation analysis. RESULTS: Up to six months, VN individuals had a significantly higher NtAb and anti-S-RBD IgG antibody responses compared to NI individuals. At the 7th month, there was a significant decline in antibody responses among VN individuals, but not NI individuals, with a minimum decrease of 3.7-fold (p < 0.001). Among VN individuals, anti-S1 IgA levels began to decrease significantly (1.4-fold; p = 0.007) after two months, and both NtAb and S-RBD IgG levels began to decline significantly (NtAb: 2.0-fold; p = 0.042, S-RBD IgG: 2.4-fold; p = 0.035) after three months. After 10 months, the most significant decline among VN individuals was observed for S-RBD-IgG (30.0-fold; P < 0.001), followed by NtAb (15.7-fold; P < 0.001) and S-IgA (3.7-fold; P < 0.001) (most stable). Moreover, after 5 months, there was no significant difference in the IgA response between the two groups. CONCLUSION: These findings have important implications for policymakers in the development of vaccination strategies, particularly in the consideration of booster doses to sustain long-lasting protection against COVID-19.

Adiponectin Ameliorates Hyperglycemia-Induced Retinal Endothelial Dysfunction, Highlighting Pathways, Regulators, and Networks.

Background: The pathophysiology of diabetic retinopathy (DR) is multifaced. A low level of circulating adiponectin (APN) in type 2 diabetes is associated with microvasculature complications, and its role in the evolution of DR is complex. Aim: This study is designed to explore the potential impact of APN in the pathogenesis of DR, linking the changes in cellular and biological processes with the pathways, networks, and regulators involved in its actions. Methods: Human microvascular retinal endothelial cells (HMRECs) were exposed to 30mM glucose (HG) and treated with globular adiponectin (30µg/mL) for 24 hours. The cells were evaluated for reactive oxidative stress (ROS) and apoptosis. RT-PCR profile arrays were utilized to evaluate the profile of genes involved in endothelial functions, angiogenesis, extracellular matrix, and adhesion molecules for hyperglycemic HMRECs treated with adiponectin. In addition, the barrier function, leukocyte migration, and angiogenesis were evaluated. The differential expressed genes (DEGs) were outlined, and bioinformatic analysis was applied. Results: Adiponectin suppresses ROS production and apoptosis in HMRECs under HG conditions. Adiponectin improved migration and barrier functions in hyperglycemic cells. The bioinformatic analysis highlighted that the signaling pathways of integrin, HMGB1, and p38 AMPK, are mainly involved in the actions of APN on HMRECs. APN significantly affects molecular functions, including the adhesion of cells, chemotaxis, migration of WBCs, and angiogenesis. STAT3, NFKB, IKBKB, and mir-8 are the top upstream regulators, which affect the expressions of the genes of the data set, while TNF and TGFB1 are the top regulators. Conclusion: Adiponectin significantly counteracts hyperglycemia at various cellular and molecular levels, reducing its impact on the pathophysiological progression towards DR in vitro using HMRECs. Adiponectin ameliorates inflammatory response, oxidative stress, and endothelial barrier dysfunction using a causal network of NFBk complex, TNF, and HMGB1 and integrin pathways.

Performance evaluation of novel fluorescent-based lateral flow immunoassay (LFIA) for rapid detection and quantification of total anti-SARS-CoV-2 S-RBD binding antibodies in infected individuals.

BACKGROUND: A vast majority of the commercially available lateral flow immunoassay (LFIA) is used to detect SARS-CoV-2 antibodies qualitatively. Recently, a novel fluorescence-based lateral flow immunoassay (LFIA) test was developed for quantitative measurement of the total binding antibody units (BAUs) (BAU/mL) against SARS-CoV-2 spike protein receptor-binding domain (S-RBD). AIM: This study aimed to evaluate the performance of the fluorescence LFIA FinecareTM 2019-nCoV S-RBD test along with its reader (Model No.: FS-113). METHODS: Plasma from 150 reverse trancriptase-PCR (RT-PCR)-confirmed positive individuals and 100 prepandemic samples were tested by FincareTM to access sensitivity and specificity. For qualitative and quantitative validation of the FinCareTM measurements, BAU/mL results of FinCareTM were compared with results of 2 reference assays: the surrogate virus-neutralizing test (sVNT, GenScript Biotech, USA) and the VIDAS®3 automated assay (BioMérieux, France). RESULTS: FinecareTM showed 92% sensitivity and 100% specificity compared with PCR. Cohen's Kappa statistic denoted moderate and excellent agreement with sVNT and VIDAS®3, with values being 0.557 (95% CI: 0.32-0.78) and 0.731 (95% CI: 0.51-0.95), respectively. A strong correlation was observed between FinecareTM/sVNT (r = 0.7, p < 0.0001) and FinecareTM/VIDAS®3 (r = 0.8, p < 0.0001). CONCLUSION: FinecareTM is a reliable assay and can be used as a surrogate to assess binding and neutralizing antibody response after infection or vaccination, particularly in none or small laboratory settings.

Human herpes simplex virus-6 (HHV-6) detection and seroprevalence among Qatari nationals and immigrants residing in Qatar.

Background: Human herpes simplex virus-6 (HHV-6) is the causative agent of exanthema subitum. Transmission mainly occurs through salivary secretions, yet blood transfusions and organ transplantations have also been reported as routes of transmission. Studies of seroprevalence of HHV-6 in the Middle East and North Africa (MENA) region and other parts of Asia are scarce. As such, this study aimed to estimate the seroprevalence of HHV-6 among healthy blood donors in Qatar. Methods: In total, 620 healthy blood donors from different nationalities residing in Qatar, mainly from the MENA region and Southeast Asia, were tested using a commercial anti-HHV-6 immunoglobulin G (IgG) enzyme-linked immunosorbent assay kit. In addition, HHV-6 DNA from randomly selected samples was tested and quantified using quantitative reverse transcriptase polymerase chain reaction. Results: Anti-HHV-6 IgG was detected in 71.7% (445/620) [95% confidence interval (CI) 68.2-75.3%] of the tested samples, while 24.3% (61/251) (95% CI 20.0-29.6%) had detectable HHV-6 viraemia. Only 22.5% of individuals with positive IgG status had detectable HHV-6 DNA in their blood, indicating a weak association between viraemia and IgG positivity (P=0.08). Furthermore, no significant difference was associated between HHV-6 viraemia and demographic characteristics, except for nationality. Conclusion: The seroprevalence of HHV-6 in Qatar was found to be similar to rates reported in other parts of the world.

Evaluation of commercially available fully automated and ELISA-based assays for detecting anti-SARS-CoV-2 neutralizing antibodies.

Rapid and accurate measurement of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV2)-specific neutralizing antibodies (nAbs) is paramount for monitoring immunity in infected and vaccinated subjects. The current gold standard relies on pseudovirus neutralization tests which require sophisticated skills and facilities. Alternatively, recent competitive immunoassays measuring anti-SARS-CoV-2 nAbs are proposed as a quick and commercially available surrogate virus neutralization test (sVNT). Here, we report the performance evaluation of three sVNTs, including two ELISA-based assays and an automated bead-based immunoassay for detecting nAbs against SARS-CoV-2. The performance of three sVNTs, including GenScript cPass, Dynamiker, and Mindray NTAb was assessed in samples collected from SARS-CoV-2 infected patients (n = 160), COVID-19 vaccinated individuals (n = 163), and pre-pandemic controls (n = 70). Samples were collected from infected patients and vaccinated individuals 2-24 weeks after symptoms onset or second dose administration. Correlation analysis with pseudovirus neutralization test (pVNT) and immunoassays detecting anti-SARS-CoV-2 binding antibodies was performed. Receiver operating characteristic (ROC) curve analysis was generated to assess the optimal threshold for detecting nAbs by each assay. All three sVNTs showed an excellent performance in terms of specificity (100%) and sensitivity (100%, 97.0%, and 97.1% for GenScript, Dynamiker, and Mindray, respectively) in samples collected from vaccinated subjects. GenScript demonstrated the strongest correlation with pVNT (r = 0.743, R2 = 0.552), followed by Mindray (r = 0.718, R2 = 0.515) and Dynamiker (r = 0.608, R2 = 0.369). Correlation with anti-SARS-CoV-2 binding antibodies was variable, but the strongest correlations were observed between anti-RBD IgG antibodies and Mindray (r = 0.952, R2 = 0.907). ROC curve analyses demonstrated excellent performance for all three sVNT assays in both groups, with an AUC ranging between 0.99 and 1.0 (p < 0.0001). Also, it was shown that the manufacturer's recommended cutoff values could be modified based on the tested cohort without significantly affecting the sVNT performance. The sVNT provides a rapid, low-cost, and scalable alternative to conventional neutralization assays for measuring and expanding nAbs testing across various research and clinical settings. Also, it could aid in evaluating actual protective immunity at the population level and assessing vaccine effectiveness to lay a foundation for boosters' requirements.

Neutralizing antibodies against SARS-CoV-2 are higher but decline faster in mRNA vaccinees compared to individuals with natural infection.

BACKGROUND: Waning protection against emerging SARS-CoV-2 variants by pre-existing antibodies elicited because of current vaccination or natural infection is a global concern. Whether this is due to the waning of immunity to SARS-COV-2 remains unclear. AIM: We aimed to investigate the dynamics of antibody isotype responses amongst vaccinated naïve (VN) and naturally infected (NI) individuals. METHODS: We followed up antibody levels in COVID-19 messenger RNA (mRNA)-vaccinated subjects without prior infection (VN, n = 100) in two phases: phase-I (P-I) at ~ 1.4 and phase-II (P-II) at ~ 5.3 months. Antibody levels were compared with those of unvaccinated and naturally infected subjects (NI, n = 40) at ~ 1.7 (P-1) and 5.2 (P-II) months post-infection. Neutralizing antibodies (NTAb), anti-S-RBD-IgG, -IgM and anti-S-IgA isotypes were measured. RESULTS: The VN group elicited significantly greater antibody responses (P < 0.001) than the NI group at P-I, except for IgM. In the VN group, a significant waning in antibody response was observed in all isotypes. There was about an ~ 4-fold decline in NTAb levels (P < 0.001), anti-S-RBD-IgG (~5-fold, P < 0.001), anti-S-RBD-IgM (~6-fold, P < 0.001) and anti-S1-IgA (2-fold, P < 0.001). In the NI group, a significant but less steady decline was notable in S-RBD-IgM (~2-fold, P < 0.001), and a much smaller but significant difference in NTAb (<2-fold, P < 0.001) anti-S-RBD IgG (<2-fold, P = 0.005). Unlike the VN group, the NI group mounted a lasting anti-S1-IgA response with no significant decline. Anti-S1-IgA, which were ~ 3-fold higher in VN subjects compared with NI in P-1 (P < 0.001), dropped to almost the same levels, with no significant difference observed between the two groups in P-II. CONCLUSION: Whereas double-dose mRNA vaccination boosted antibody levels, vaccinated individuals' 'boost' was relatively short-lived.

Assessment of the Neutralizing Antibody Response of BNT162b2 and mRNA-1273 SARS-CoV-2 Vaccines in Naïve and Previously Infected Individuals: A Comparative Study.

The currently authorized mRNA COVID-19 vaccines, Pfizer-BNT162b2 and Moderna-mRNA-1273, offer great promise for reducing the spread of the COVID-19 by generating protective immunity against SARS-CoV-2. Recently, it was shown that the magnitude of the neutralizing antibody (NAbs) response correlates with the degree of protection. However, the difference between the immune response in naïve mRNA-vaccinated and previously infected (PI) individuals is not well studied. We investigated the level of NAbs in naïve and PI individuals after 1 to 26 (median = 6) weeks of the second dose of BNT162b2 or mRNA-1273 vaccination. The naïve mRNA-1273 vaccinated group (n = 68) generated significantly higher (~2-fold, p ≤ 0.001) NAbs than the naïve BNT162b2 (n = 358) group. The P -vaccinated group (n = 42) generated significantly higher (~3-fold; p ≤ 0.001) NAbs levels than the naïve-BNT162b2 (n = 426). Additionally, the older age groups produced a significantly higher levels of antibodies than the young age group (<30) (p = 0.0007). Our results showed that mRNA-1273 generated a higher NAbs response than the BNT162b2 vaccine, and the PI group generated the highest level of NAbs response regardless of the type of vaccine.

Detection of Antinuclear Antibodies Targeting Intracellular Signal Transduction, Metabolism, Apoptotic Processes and Cell Death in Critical COVID-19 Patients.

Background and Objectives: The heterogeneity of the coronavirus disease of 2019 (COVID-19) lies within its diverse symptoms and severity, ranging from mild to lethal. Acute respiratory distress syndrome (ARDS) is a leading cause of mortality in COVID-19 patients, characterized by a hyper cytokine storm. Autoimmunity is proposed to occur as a result of COVID-19, given the high similarity of the immune responses observed in COVID-19 and autoimmune diseases. Here, we investigate the level of autoimmune antibodies in COVID-19 patients with different severities. Results: Initial screening for antinuclear antibodies (ANA) IgG using ELISA revealed that 1.58% (2/126) and 4% (5/126) of intensive care unit (ICU) COVID-19 cases expressed strong and moderate ANA levels, respectively. An additional sample was positive with immunofluorescence assays (IFA) screening. However, all the non-ICU cases (n=273) were ANA negative using both assays. Samples positive for ANA were further confirmed with large-scale autoantibody screening by phage immunoprecipitation-sequencing (PhIP-Seq). The majority of the ANA-positive samples showed "speckled" ANA pattern by microscopy and revealed autoantibody specificities that targeted proteins involved in intracellular signal transduction, metabolism, apoptotic processes, and cell death by PhIP-Seq; further denoting reactivity to nuclear and cytoplasmic antigens. Conclusion: Our results further support the notion of routine screening for autoimmune responses in COVID-19 patients, which might help improve disease prognosis and patient management. Further, results provide compelling evidence that ANA-positive individuals should be excluded from being donors for convalescent plasma therapy in the context of COVID-19.