Studying the Humoral Response against SARS-CoV-2 in Cuban COVID-19 Recovered Patients.

Understanding the immune response generated by SARS-CoV-2 is critical for assessing efficient therapeutic protocols and gaining insights into the durability of protective immunity. The current work was aimed at studying the specific humoral responses against SARS-CoV-2 in Cuban COVID-19 convalescents. We developed suitable tools and methods based on ELISA methodology, for supporting this evaluation. Here, we describe the development of an ELISA for the quantification of anti-RBD IgG titers in a large number of samples and a similar test in the presence of NH4SCN as chaotropic agent for estimating the RBD specific antibody avidity. Additionally, a simple and rapid ELISA based on antibody-mediated blockage of the binding RBD-ACE2 was implemented for detecting, as a surrogate of conventional test, the levels of anti-RBD inhibitory antibodies in convalescent sera. In a cohort of 273 unvaccinated convalescents, we identified higher anti-RBD IgG titer (1 : 1,330, p < 0.0001) and higher levels of inhibitory antibodies blocking RBD-ACE2 binding (1 : 216, p < 0.05) among those who had recovered from severe illness. Our results suggest that disease severity, and not demographic features such as age, sex, and skin color, is the main determinant of the magnitude and neutralizing ability of the anti-RBD antibody response. An additional paired longitudinal assessment in 14 symptomatic convalescents revealed a decline in the antiviral antibody response and the persistence of neutralizing antibodies for at least 4 months after the onset of symptoms. Overall, SARS-CoV-2 infection elicits different levels of antibody response according to disease severity that declines over time and can be monitored using our homemade serological assays.

Safety and immunogenicity of the FINLAY-FR-1A vaccine in COVID-19 convalescent participants: an open-label phase 2a and double-blind, randomised, placebo-controlled, phase 2b, seamless, clinical trial.

BACKGROUND: A phase 1, clinical trial to evaluate FINLAY-FR-1A vaccine in COVID-19 convalescent individuals was completed. Here, we report results of the phase 2, clinical trial. METHODS: We studied 450 convalescent participants with a history of asymptomatic, mild, or moderate COVID-19 at the National Institute of Hematology and Immunology and the National Centre for Sexual Education in Havana, Cuba. The study included adults aged 19-78 years who had recovered from COVID-19 and had had a negative PCR test at least 2 months before the initiation of the study. Phase 2 was done sequentially in two stages. The first stage to assess safety comprised an open, non-controlled phase 2a study in participants aged 60-78 years who received a single dose of the FINLAY-FR-1A vaccine (50 µg of recombinant dimeric receptor binding domain [RBD]). The second stage comprised the placebo-controlled, double-blind, phase 2b trial in participants aged 19-78 years, where participants were randomly assigned (4:1) into two groups: an experimental group vaccinated with a single dose of the FINLAY-FR-1A vaccine, and a control (placebo) group injected with vaccine excipient. The primary outcomes were safety, evaluated 28 days after vaccination by the occurrence of serious adverse events in all participants, and successful immune response, assessed by neutralising antibody ELISA, and defined as half-maximal surrogate virus neutralisation titres of 250 or more. Secondary endpoints included vaccine immunogenicity assessed by ELISA anti-RBD and live-virus neutralisation test. All randomly assigned participants were included in the safety analysis (safety population), and immunogenicity was evaluated in participants without study interruptions (per-protocol population). The trial is registered with the Cuban Public Registry of Clinical Trials, RPCEC00000366-En and WHO-ICTRP and is complete. FINDINGS: From April 9, 2021, to April 17, 2021, 663 COVID-19 convalescent participants were enrolled in the study; 213 participants did not meet the selection criteria and 450 volunteers were recruited. 20 participants aged 60-78 years were included in the open, single-group, phase 2a study and 430 participants were randomly assigned to the experimental (n=344) or control groups (n=86) in the phase 2b study of participants aged 19-78 years. 19 (95%) of 20 phase 2a volunteers achieved a successful immune response after vaccination. No vaccine-associated serious adverse events were reported in the whole study population. Minor adverse events were found, the most common being pain at the injection site (105 [29%] of 364 in the intervention group; 13 [15%] of 86 in the placebo group). A successful immune response was found in 289 (81%) of 358 participants 28 days after vaccination. The vaccine elicited a greater than 31-times increase in anti-RBD-IgG antibodies compared with prevaccination rates, and the seroconversion rate was 302 (84%) of 358 on day 28 after vaccination; the geometric mean titres of live-virus neutralisation test increased from 15·4 (95% CI 10·3-23·2) to 400·3 (272·4-588·1) and high response was found against alpha, beta, and delta variants of concern. INTERPRETATION: A single dose of the FINLAY-FR-1A vaccine against SARS-CoV-2 strengthened the pre-existing natural immunity, with excellent safety profile. FUNDING: Cuba's Ministry of Science, Technology, and Environment.

An enzyme immunoassay for determining albumin in human urine samples using an ultra-microanalytical system.

Microalbuminuria is a term to describe a moderate increase in the level of albumin in urine. It is an important prognostic marker for kidney damage in diseases such as diabetes mellitus and hypertension. A simple sandwich-type ultramicroELISA assay (UMELISA) has been developed for the measurement of albumin in human urine samples. Strips coated with a high affinity monoclonal antibody directed against albumin are used as solid phase, to ensure the specificity of the assay. The albumin assay was completed in 1 hr and 30 min, with a measuring range of 1.44-200 ng/mL. The intra- and inter-assay coefficients of variation were 3.98-4.35% and 7.59-8.92%, respectively, depending on the albumin concentrations evaluated. Percentage recovery ranged from 94.26 to 98.50%. Regression analysis showed a good correlation with the commercial quantitative turbidimetric test Microalbumin-turbilatex (n = 240, r = 0.994, p < .01). The analytical performance characteristics of our UMELISA MICROALBUMINA endorse its use for the quantification of albumin in human urine samples. This test will make a cost-effective diagnostic kit accessible to low-income countries such as Latin American countries and is now available in the Cuban Public Health System.

Performance characteristics of a fast real-time PCR assay for hepatitis B virus DNA quantification.

Detection of hepatitis B virus (HBV) DNA is particularly important for detection of early acute and of occult HBV infection. On the other hand, HBV DNA detection and quantification are essential to diagnose and treat chronic HBV infection. In this study, we evaluated the performance of the real-time PCR SUMASIGNAL VHB (un paso) (Immunoassay Center, Cuba). The clinical and analytical specificity of the assay was 100%. Intra-assay and inter-assay coefficients of variation ranged from 0.50 to 2.53% and from 1.23 to 3.03%, respectively. A strong correlation (r=0.926; P<0.001) with the COBAS® AmpliPrep/COBAS® TaqMan® HBV Test, v2.0 (Roche Molecular Systems, Inc.) was obtained. The limit of detection using the third WHO international standard for HBV DNA was 377.47 IU/mL. The test was able to detect the most prevalent HBV genotypes (A-G) equally well. In conclusion, the SUMASIGNAL VHB (un paso) is a sensitive, specific, precise and accurate assay for the quantification of serum and plasma HBV DNA. Thus, this simple and fast real-time PCR test can be used as an aid in diagnosing an HBV infection and monitoring drug efficacy.

Identificación en especies de Aeromonas aisladas en Cuba / Identification of Aeromonas species isolated in Cuba

Durante los meses de enero y julio de 1990, se estudiaron 100 cepas de Aeromonas, aisladas de niños menores de 5 años con enfermedad diarreica aguda, procedentes de diferentes centros de salud del país. Utilizando el esquema de Popoff y Veron amplaido por Janda, se logró identificar en especies el 63 por ciento de las cepas mediante el uso de la pruebas primarias, y el 37 por ciento restante complementado con la pruebas suplementarias, fueron identificadas el 100 por ciento de las cepas en Aeromonas sobria, Aeromonas hydrophila y Aeromonas caviae