Social and Behavioral Science: The Missing Ingredient to Successful Clinical Trials

This talk considers the role of social and behavioral science at every stage of the clinical trial process from design to enrolment, participation, retention and outcomes. Based on a review of the literature and three decades' experience as a social scientist conducting leading HIV and COVID studies, it argues that understanding human behavior and decision-making alongside the context in which these decisions are made are key to effective, efficient and quality clinical trials.

SEXUAL BEHAVIOR AND STI INCIDENCE DURING THE FIRST 4 YEARS OF PrEP USE AMONG MSM

Background: A prospective demonstration project in Amsterdam (AMPrEP) provided pre-exposure prophylaxis (PrEP) to people vulnerable to HIV in 2015- 2020. Data on long-term trends in sexual behavior and incidence of STIs during PrEP use are needed to inform future PrEP programs. Therefore, we assessed sexual behavior and incidence rates of STIs among MSM and transgender women on PrEP over four years. Method(s): AMPrEP participants chose between oral PrEP daily (dPrEP) or event-driven (edPrEP) at baseline and could switch regimens at each 3-monthly study visit. They were tested for STIs at these visits and if necessary in between. Follow-up began at PrEP initiation and continued until 48 months of follow-up or was censored at March 15, 2020 (start COVID-19), whichever occurred first. We assessed changes over time in incidence rates (IR) of chlamydia, gonorrhea, and infectious syphilis using Poisson regression. We estimated the IR of Hepatitis C (HCV) diagnoses per consecutive year. We described the number of HIV diagnoses, and sexual behavior (i.e. number of sex partners, condomless anal sex acts with casual partners [CAS]). Result(s): A total of 367 (365 MSM) started PrEP and contributed 1249 person-years of observation. IRs of any STI was 87[95%CI 82-93]/100PY. There was no change in the IR of any STI and infectious syphilis over time on PrEP. We observed a slight decrease in incident chlamydia and gonorrhea in daily PrEP users (Table). Two incident HIV cases were diagnosed in the first year of follow-up. IRs for HCV were 1.5[0.6-3.6], 2.5[1.3-5.0], 0.7[0.2-2.7], and 0.4[0.1- 2.8]/100PY, per consecutive year on PrEP. Median number of sex partners per 3-month period decreased from 16[IQR 8-34] and 12[6-25] (dPrEP and edPrEP, respectively) at baseline, 15[7-30] and 8[3-16] at 24 months, and 12[6-26] and 5[2-12] at 48 months. Median number of CAS acts with casual partners were respectively 7[3-15] and 4[1-9] at baseline, 14[5-25] and 4[1-12] at 24 months, and 12[4-25] and 4[1-9] at 48 months. Conclusion(s): Over the first 4 years of PrEP use overall STI incidence was high and stable. Chlamydia and gonorrhea incidence declined slightly in daily users. Numbers of sex partners seemed to decrease in both dPrEP and edPrEP users. Number of CAS acts with casual partners appeared to increase first, and then stabilized. Notably, this did not result in increased incidence of STIs. Regular testing and treatment of STIs remain a priority among PrEP users. Biomedical prevention of STIs can be examined in this context.

Circulation of genetically diverse non-polio enteroviruses in respiratory samples during COVID-19 pandemic period (2021-22)

Enteroviruses, beyond poliovirus, are important pathogens. Several non-polio enteroviruses (NPEVs) are causing epidemics all around the world. Limited data is available on the prevalence and diversity of these viruses from India. Objective(s): Detection and characterization of NPEVs in respiratory samples during the COVID-19 pandemic period. Material(s) and Method(s): COVID-19 negative samples from acute respiratory infections (ARI) [n = 105] and severe-acute respiratory infections (SARI) [n = 148] during the period 2021-22 were screened for NPEVs. Detection was carried out using the one step RT-PCR method targeting the 5'UTR region followed by molecular analysis. Results and Conclusion(s): Total positivity of NPEVs was noted in 35.23% and 31.08% of the ARI and SARI cases, respectively. Comparison within the two groups studied, showed significant difference in the age-wise distribution for cases>18 years of age. Year round seasonality for ARI cases while autumn seasonality for SARI cases was observed. Sequencing of representative samples of ARI cases showed prevalence of Rhinovirus A (RVA), Rhinovirus B (RVB), Rhinovirus C (RVC) and Echovirus, while predominance of RVC followed by RVA was observed for the SARI cases. Phylogenetic analysis of all the strains showed clustering of RVC strains in different clusters. Divergence was also noted in RVA and RVB strains studied. Circulation of a rare Echovirus-29 strain was noted in the ARI cases. The study highlighted significant divergence in the Rhinovirus strains studied. It warrants the need for surveillance of NPEVs, whole-genome sequencing of the circulating strains for better understanding of biodiversity among the NPEVs and the potential health burden.

Mental disorders during the pandemic of a new coronavirus infection: causes and predisposing factors.

Objective: to study the causes and predictors of mental disorders during the COVID-19 epidemic in those who turned to psychiatrist for the first time, as well as in patients with already diagnosed mental illness. Patients and methods. We examined 100 patients who turned to a psychiatrist due to a deterioration in their mental state during the pandemic, 50 patients were newly diagnosed (Group 1) and 50 with previously diagnosed mental disorders (Group 2). The study was carried out by a clinical method using a specially designed map, followed by statistical processing of the obtained data. Results and discussion. Mental disorders caused by the COVID-19 pandemic more often occurred at a young age, in patients with higher and secondary specialized education, and in single patients. In the 1st group, as a result of exposure to psychogenic factors (the influence of the media, quarantine, economic changes), anxiety (36.8%) and depressive (21.1%) disorders occurred more often, and after the coronavirus infection, depressive disorders were in the first place (54.2%). The 2nd group mostly included patients with endogenous disorders (bipolar affective disorder - 24%, recurrent depressive disorder - 20%, schizophrenia - 20%), which were exacerbated more often as a result of COVID-19, to a lesser extent - psychogenic (experiences associated with a change in material status and illness of relatives). Obsessive-compulsive disorder, generalized anxiety disorder, somatoform disorders have been associated with epidemic factors. Conclusion. The results obtained indicate that there are differences between the mental disorders that first appeared during the pandemic and the exacerbations of the condition in mentally ill patients, which relate to the predictors, causes and clinical manifestations of these disorders.Copyright © 2022 Ima-Press Publishing House. All rights reserved.

Online survey results among social network users in terms of the COVID-19 problem.

Aim. An online survey among social network users was conducted to assess the frequency of COVID-19 cases, the spectrum of medications used for treatment, and the subjective assessment of clinical manifestations of the disease. Material and methods. An anonymous online survey was conducted among users of various social networks using a questionnaire created on the SurveyMonkey survey and research platform. During the first month of December 2021, the survey included 23 questions regarding the clinical and demographic characteristics of respondents, the number of COVID-19 cases, clinical manifestations, and severity, as well as the need for medical help and medication. Results. 752 respondents took part in the online survey, more than 70% of them are under 50 years old. Among the respondents 59.73% had a new coronavirus infection (COVID-19). More than 40% of the participants had COVID-19 in the period from September 2020 to April 2021 (2nd wave in Russia). In 79.2% of people, the presence of a new coronavirus infection was confirmed by one of the diagnostic methods: polymerase chain reaction (PCR test), radiography, the presence of antibodies to Ig G/M, and took into account the presence of contact with infected SARS-CoV-2. 411 participants observed any clinical manifestations of the disease. Most often respondents who had COVID-19 indicated weakness, cough, dyspnea, disappearance or decrease in the acuity of smell and taste. The volume of lung tissue damage in 36.5% of cases was less than 25%. The disappearance of any clinical manifestations of the disease immediately after recovery was noted by 32.0% of respondents. Most of the patients (59.2%) sought medical help at the polyclinic, 38.9% had to self-medicate. 71.9% respondents indicated they had been vaccinated against COVID-19, but without specifying the timing and completeness of the course. Side effects after immunization (fever, weakness, soreness, and redness at the injection site) were subjectively assessed by 41.9% of respondents. Conclusion. Among the surveyed respondents, 62.7% of the disease symptoms were mild. The highest number of cases occurred in the 2nd and 4th waves of COVID-19 morbidity in Russia. Most often respondents indicated symptoms of acute respiratory infection. The complete disappearance of clinical manifestations of the disease immediately after recovery was noted by 32.0 % of respondents, and the persistence of symptoms for up to a year - 7.5. More than 70% of the participants in the online survey reported vaccination against COVID-19, but the questionnaire did not include questions about the timing of vaccinations (before or after COVID-19) and the completeness of the course.Copyright © Eco-Vector, 2023. All rights reserved.

Tocilizumab, a cutting-edge therapeutic resource against SARS-CoV-2 infection: A scoping review.

OBJECTIVE: To explore the medical evidence published until April 20, 2022, about the efficacy and safety of tocilizumab in COVID-19 patients. METHODOLOGY: Scoping review that included PubMed and Scopus, searching for clinical trials and observational studies in English and Spanish. Additionally, records of clinical trials from the International Clinical Trials Registry Platform were analyzed. RESULT(S): Fifty-four documents were included: retrospective cohort studies (n = 20), randomized clinical trials (n = 16), case control studies (n = 7), non-randomized clinical trials (n = 5) and prospective cohort studies (n = 6), with a total study population of 20,007 patients. There were 15 records of clinical trials of which 10 were registered in the US National Library of Medicine. CONCLUSION(S): Tocilizumab could be effective and safe to treat patients with moderate to critical COVID-19, in conjunction with additional immunomodulators and antivirals. A greater number of randomized clinical trials, however, are needed to explore the efficacy and safety of tocilizumab.Copyright © 2023 Comunicaciones Cientificas Mexicanas S.A. de C.V.. All rights reserved.

Rising level of pulmonary dioxin might trigger CMV and HSV-1 reactivation via bHLH/PAS/AhR:ARNT transcription pathway in lungs of immunocompetent COVID-19 patients with ARDS

Study objective. It has been shown that human common viruses are new target genes for host cell dioxin receptor transcriptional (AhR-ARNT) complex initially proven to up-regulate mammalian genes containing dioxin-response elements (DRE) in the promoters [doi:10.1016/j.ijid.2012.05.265]. Initially, transactivation of HIV-1 and HBV by 2,3,7,8-tetrachlodibenzop- dioxin (TCDD) at low nanomolar range was demonstrated [doi:10.3109/00498259309057034]. Noteworthy, transactivation of human cytomegalovirus (CMV) was shown with 0.3 ppt dioxin, i.e. lower than its current background level in the general population (~3.0 ppt). Recently, reactivation of CMV infection was found to influence worse clinical outcome following SARS-CoV-2 infection (doi: 10.1186/s12979-020- 00185-x). Other findings showed that CMV and herpes simplex virus 1 (HSV-1) reactivation were observed in immunocompetent patients with COVID-19 acute respiratory distress syndrome (ARDS) (doi.org/10.1186/s13054-020-03252-3). Addressing occurrence of Herpesviridae reactivation in severe COVID-19 patients, and still unspecified real triggers of CMV and HSV-1 reactivations, we tested TCDD, which current body burden (DBB) ranges from 20 pg/g (TEQ in fat) in general population to 100 pg/g in older people. Methods. In Silico quantitation of active DRE in promoters of viral genes. Virus DNA hybridization assay. Clinical and epidemiological analyses. Results and Discussion. In this study, a computational search for DRE in CMV and HSV-1 genes was performed by SITECON, a tool recognizing potentially active transcriptional factor binding sites. In silico analysis revealed in regulatory region of CMV IE genes from 5 to 10 DRE, and from 6 to 8 DRE in regulatory region of HSV-1 IE genes.We established that a low picomolar TCDD can trigger up-regulation of CMV and HSV-1 genes via AhR:Arnt transcription factor in macrophage(doi.org/10.1016/ j.ijid.2012.05.265) and glial human cell lines (doi.org/10.1016/j. jalz.2016.06.1268), respectively. In fact, viral reactivation may be triggered in COVID-19 ARDS patients by higher pulmonary TCDD concentrations, because "lipid storm" within lungs of severe COVID-19 patients has been recently reported (doi.org/ 10.1101/2020.12.04.20242115). TCDD is known as the most potent xenobiotic, which bioaccumulates and has estimation half-life in humans of up to 10 yr. Due to hydrophobic character (Log P octanol/water: 7.05), TCDD partitions into inflammatory lipids in lung tissue thus augmenting its local concentration. Population-based epidemiological data on SARS-CoV-2 first wave of pandemic revealed high level of CMV seropositivity and cumulative mortality rate 4.5 times in Lombardi region of Italy, where after Seveso industrial accident TCDD plasma level in pre-exposed subjects is 15 times the level in rest of Italy (doi. org/10.3389/fpubh.2020.620416). Also, Arctic Native (AN) peoples consume dioxin-contaminated fat in seafood and have TCDD DBB, i.e. 7 times that in general population. To the point of this paper, their COVID-19 mortality is 2.2 times of that among non-AN Alaskans (doi: 10.15585/mmwr.mm6949a3). Conclusion(s): TCDD in the picomolar range may trigger CMV expression in lung cells and commit virus to the lytic cycle, which can be applied to reactivation of Herpesviridae infection in immunocompetent patients with COVID-19 ARDS syndrome.Copyright © 2023 The American Society for Biochemistry and Molecular Biology, Inc.

VSV-BASED COVID VACCINE FOR THE PREVENTION OF BREAKTHROUGH SARS-CoV-2 INFECTIONS

Background: More than 12 billion doses of COVID-19 vaccine administrations and over 630 million natural infections should have developed adequate levels of herd immunity over the last three years. However, there have been many new waves of coronavirus infections. The development of safe and effective vaccines to control breakthrough SARS-CoV-2 infections remain an urgent priority. We have developed a recombinant VSV vector-based vaccine to fulfill this worldwide need. Method(s): We have used a recombinant vesicular stomatitis virus (rVSV)-based prime-boost immunization strategy to develop an effective COVID-19 recall vaccine candidate. We have constructed an attenuated recombinant VSV genome carrying the full-length Spike protein gene of SARS-CoV-2. Adding the honeybee melittin signal peptide (msp) at the N-terminus enhanced the protein expression and adding the VSV G protein transmembrane domain and the cytoplasmic tail (Gtc) at the C-terminus of the Spike protein allowed efficient incorporation of the Spike protein into pseudotype VSV. Result(s): In immunized mice, rVSV with chimeric rVSV-msp-S-Gtc induced high levels of potent neutralizing antibodies (nAbs) and CD8+ T cell responses, while the full-length Spike with Gtc proved to be the superior immunogen. More importantly, rVSV-msp-S-Gtc-vaccinated animals were completely protected from subsequent SARS-CoV-2 challenges. Furthermore, rVSV-Wuhan and rVSV-Delta vaccines, and an rVSV-Trivalent (mixed rVSV-Wuhan, -Beta and -Delta) vaccine elicited potent nAbs against live SARS-CoV-2 Wuhan (USAWA1), Beta (B.1.351), Delta (B.1.617.2) and Omicron (B.1.1.529) viruses. Heterologous boosting of rVSV-Wuhan with rVSV-Delta induced strong nAb responses against Delta and Omicron viruses, with the rVSV-Trivalent vaccine consistently inducing effective nAbs against all the SARS-CoV-2 variants tested. All rVSV-msp-S-Gtc vaccines also elicited an immunodominant Spike-specific CD8+ T cell response. Conclusion(s): rVSV vaccines targeting SARS-CoV-2 variants of concern can be considered as an effective booster vaccine in the global fight against COVID-19.

SARS-CoV-2 INFECTION AMONG PERSONS WHO INJECT DRUGS AND THEIR PARTNERS IN KENYA

Background: Despite increased social vulnerability and barriers to care, there has been a paucity of data on SARS-CoV-2 incidence among key populations in sub-Saharan Africa. We seek to characterize active infections and define transmission dynamics of SARS-CoV-2 among people who inject drugs (PWID) and their sexual and injecting partners from Nairobi and the coastal region in Kenya. Method(s): This was a nested cross-sectional study of SARS-CoV-2 infection from April to July 2021 within a cohort study of assisted partner services for PWID in Kenya. A total of 1000 PWID and their partners (500 living with and 500 living without HIV) were recruited for SARS-CoV-2 antibody testing, of whom 440 were randomly selected to provide self-collected nasal swabs for real-time PCR testing. Whole genome sequencing (WGS) was completed on a limited subset of samples (N=23) with cycle threshold values 32.0. Phylogenetic tree construction and analysis was performed using the Nextstrain pipeline and compared with publicly available SARS-CoV-2 sequences from GenBank. Result(s): A total of 438 (99.5%) participants provided samples for SARS-CoV-2 PCR testing. Median age was 37 (IQR 32-42);128 (29.2%) were female;and 222 (50.7%) were living with HIV. The overall prevalence of SARS-CoV-2 infection identified by RT-PCR was 86 (19.6%). In univariate analyses, there was no increased relative risk of SARSCoV- 2 infection related to positive HIV status, frequenting an injection den, methadone treatment, unstable housing, report of any high-risk exposure, or having a sexual or injecting partner diagnosed with COVID-19 or who died from COVID-19 or flu-like illness. Eight samples were successfully sequenced via WGS and classified as WHO variants of concern: 3 Delta, 3 Alpha, and 2 Beta. Seven were classified into clades predominantly circulating in Kenya during 2021. Notably, two sequences were identical and matched identically to another Kenyan sequence, which is consistent with, though not indictive of, a transmission linkage. Conclusion(s): Overall, the risk of SARS-CoV-2 infection in this population of PWID and their partners was not significantly associated with risk factors related to injection drug use. At a genomic level, the SARS-CoV-2 strains in this study were consistent with contemporary Kenyan lineages circulating during the time and not unique to PWID. Prevention efforts, therefore, must also focus on marginalized groups for control given the substantial amount of mixing that likely occurs between populations.

MAINTAINING IMMUNOLOGICAL MEMORY TO THE SARS-CoV-2 VIRUS DURING COVID-19 PANDEMIC.

The question on the duration and effectiveness of post-infection vs post-vaccination SARS-CoV-2 immunity remains in the focus of numerous studies. The aim of the work was to examine the duration of maintained post-infection and post-vaccination SARS-CoV-2 immunity as well as formation of hybrid (vaccination after infection) and breakthrough (repeated disease or disease after vaccination) immunity in the context of an ongoing COVID-19 pandemic. 107 adults with mild or moderate COVID-19 3-18 months after the disease and 30 subjects vaccinated twice with the Sputnik V vaccine were examined 1-6 times. Antibodies against SARS-CoV-2 virus were determined by ELISA on the "SARSCoV-2-IgG quantitative-ELISA-BEST" test systems. The antibody avidity was measured by additional incubation with and without denaturing solution. Mononuclear cells were isolated from blood by gradient centrifugation, incubated with and without coronavirus S-protein for 20 hours, stained with fluorescently labeled antibodies, and the percentage of CD8highCD107a+ was counted using FACSCanto II cytometer. It was shown that in the group of convalescent and vaccinated subjects, the level of virus-specific antibodies decreased more deeply in individuals with initially high humoral response, but 9 months later the decrease slowed down and reached a plateau. The antibody avidity rose up to 50% and persisted for 18 months. Cellular immunity in recovered patients did not change for 1.5 years, while in vaccinated patients it gradually decreased 6 months later, but remained at detectable level. After revaccination, a significant increase in the level of antibodies, avidity up to 67.6% and cellular immunity returned to the initial level were noted. Hybrid immunity turned out to be significantly higher than post-infection and post-vaccination immunity. The level of antibodies increased to 1218.2 BAU/ml, avidity - to 69.85%, and cellular immunity - to 9.94%. Breakthrough immunity was significantly higher than that after the first disease. The level of antibodies rose to 1601 BAU/ml, avidity - up to 81.6%, cellular immunity - up to 13.71%. Using dynamic observation of four COVID-19 convalescents, it has been shown that in the context of the ongoing pandemic and active coronavirus mutation, natural boosting occurs both asymptomatically and as a result of a mild re-infection, which prevents disappearance of SARS-CoV-2 humoral and cellular immunity.Copyright © 2023 Saint Petersburg Pasteur Institute. All rights reserved.

MECHANISM AND RESISTANCE STUDIES OF SARS-CoV-2 MPRO INHIBITOR POMOTRELVIR (PBI-0451)

Background: The unprecedented scale of the COVID-19 pandemic and rapid evolution of SARS-CoV-2 variants underscores the need for broadly active inhibitors with a high barrier to resistance. The coronavirus main protease (Mpro) is an essential viral enzyme required for viral polyprotein processing and is highly conserved across human coronaviruses. Pomotrelvir (PBI-0451) is a novel Mpro inhibitor currently completing phase 2 clinical trial. Here we describe the mechanism of action, broad activity against SARS-CoV-2 clinical isolates, combination studies with other SARS-CoV-2 inhibitors and favorable resistance profile of pomotrelvir. Method(s): The kinetic parameters of pomotrelvir Mpro inhibition and its interaction with nirmaltrevir were determined in a kinetic protease assay. The IC50s of pomotrelvir on mutant Mpro proteins were measured in an endpoint Mpro assay. Combination studies of pomotrelvir with remdesivir and molnupiravir were carried out in A549-hACE2 cells infected with SARS-CoV-2 NLuc virus. Activity against SARS-CoV-2 clinical variants was assessed by infection of A549-ACE2-TMPRSS2 cells followed by immunostaining of the viral nucleocapsid protein. Result(s): Pomotrelvir is a potent competitive inhibitor of SARS-CoV-2 Mpro (Ki =2.7 nM). Binding of pomotrelvir and the Mpro inhibitor nirmatrelvir to the active site is mutually exclusive. In the SARS-CoV-2 NLuc assay, pomotrelvir is additive when combined with remdesivir or molnupiravir, two nucleoside analogs targeting viral RNA synthesis. When the effect of Mpro substitutions previously selected in a resistance study of pomotrelvir were analyzed in an enzyme assay, only Mpro-N133H showed a significant increase in IC50 (45-fold). The catalytic efficiency of Mpro-N133H is reduced by 10-fold and the recombinant virus SARSCoV-2 (WA1) -N133H is not viable, suggesting that N133H has lower replicative fitness. Lastly, pomotrelvir exhibits broad activity against all SARS-CoV-2 clinical isolates tested to date, including five omicron variants. Conclusion(s): PBI-0451 is a potent competitive inhibitor of SARS-CoV-2 Mpro and is broadly active against SARS-CoV-2 clinical isolates including omicron variants. Results from inhibitor interaction studies support the potential combination of pomotrelvir with remdesivir and molnupiravir but not nirmatrelvir. Enzymatic characterization of in vitro selected pomotrelvir resistant variants indicates they either confer no resistance or have reduced fitness.

Use of daily home spirometry to predict clinical outcomes in persons with cystic fibrosis

Background: Use of home spirometry to monitor lung function has been increasing in popularity in persons with cystic fibrosis (PwCF) since the start of the COVID-19 pandemic. Although clinic spirometry is interpreted from validated standards, expected test-to-test variation of home spirometry and how variation during baseline health may relate to clinical changes are unknown. The aim of this study was to determine variation in baseline lung function during daily home spirometry and identify associations with clinical outcomes. Method(s): Subjectswere selected based on available spirometry data from a cohort of PwCF enrolled in a long-term airway microbiome study. Subjects were provided with a PiKo-6 hand-held spirometer (nSpire Health, Inc., Longmont, CO) and asked to perform spirometry maneuvers three times per. Validity of home spirometry (percentage predicted forced expiratory volume in 11 second (FEV1pp)) was compared with that clinic spirometry using Bland-Altman plots. Spirometry acceptability across multiple maneuvers in the same day was assessed using the American Thoracic Society (ATS) guidelines, with grade A or B (two or more maneuvers within 150 mL) considered acceptable. Variation in FEV1pp was assessed by calculating a mean FEV1pp and coefficient of variation (CoV). The association between CoV and pulmonary exacerbations (PEx) was tested using Cox proportional hazards regression models. Result(s): Thirteen subjects (62% female) with a mean age of 28.7 +/- 8.3 and mean FEV1pp of 59.9 +/- 8.2%were included. Median study durationwas 377 days (range, 33-730 days). Subjects used the home spirometer on average 51.2% of the study days (range 15-97%). On average, 58.9% of subjects (range 12-100%) used the home spirometer at least twice aweek, and 76.8% (range 65-100%) at least once aweek. To focus on periods of baseline health, days associated with PEx (spirometry performed 2 weeks before and during times of antibiotic therapy) were excluded. A median of 204 days (range 11-728 days) of baseline spirometry readings was available for further analysis. Comparing validity of home spirometry with that of clinic spirometry, Bland-Altman plots demonstrated overall good agreement with a slight bias (+0.042 L) toward higher readings for clinic FEV1pp (95% limits of agreement, -0.11-0.19 L). Spirometry quality was graded as acceptable on most study days (mean 90.6 +/- 4.6%) in which two or more maneuvers were recorded. Intra-individual variation in baseline FEV1pp was high, with a mean variation of 17.6 +/- 5.9% day to day and 15.2 +/- 6.2% week to week. Neither rates of acceptable spirometry grades nor CoV was associated with lung disease severity. Of the 13 subjects, 10 experienced one or more PEx, for a total of 32 PEx during the study. CoV was not associated with time to first PEx (hazard ratio [HR], 0.78;95% confidence interval [CI], 0.51-1.21;p = 0.24) or time to subsequent PEx (HR, 0.91;95% CI, 0.73-1.12;p = 0.28) during the study. Conclusion(s): Although home spirometry has generally good validity and acceptability, variation in lung function during baseline health is present and often exceeds expected variation in clinic spirometry per ATS standards. Variability may represent normal physiological variation or be related to the home spirometer itself or other factors but did not portend upcoming PEx. Recognition of variation during baseline health provides important context for interpretation of home spirometry.Copyright © 2022, European Cystic Fibrosis Society. All rights reserved

Risk factors for rejection following COVID-19 in lung transplant recipients

Introduction/Aim: The prevalence of and risk factors for acute cellular (ACR) and antibody mediated rejection (AMR) in lung transplant (LTx) recipients is unclear. Method(s): We performed a retrospective cohort study of all living LTx recipients between January 2020 and September 2022. Recipients with COVID-19 infection and those diagnosed with and/or treated for ACR or AMR were identified. Baseline demographics are described. A logistic regression univariate analysis was used to identify risk factors for rejection. Result(s): 128/387 (33%) LTx recipients tested positive to SARS-CoV-2 during the study period. 44 (32.3%) patients were investigated for graft dysfunction, with persistent loss of >=10% of FEV 1 at >=90-days in 37 (31.4%), median was 54.5 years (23-76). There was no significant difference between gender, disease severity or presence of chronic lung allograft dysfunction (CLAD) at time of COVID-19 infection. 9(20.5%) recipients experienced rejection, 3 (6.8%) with AMR, 5 (11.4%) ACR, and 1 (2.3%) both. Median time to onset of rejection was 59 days (16-239). Change in FEV 1 post COVID-19 was not significantly different between recipients with and without rejection, with mean volume loss in rejection group 559 mL (SD 678 mL, 22.9%), and 842 mL (SD 824 mL, 42.9%) in non-rejecters. Univariate logistic regression of risk factors demonstrated younger patients were at higher risk of rejection (OR 0.95 [95% CI 0.90-1.00] p = 0.05). Female gender was weakly associated with rejection (OR 0.21 [95% CI 0.04-1.18] p = 0.08). Time post-transplant, severe COVID illness, early COVID-19 treatment did not show association. Conclusion(s): Acute rejection occurs frequently following COVID infection and should be considered a differential in persistent allograft dysfunction. Younger age and female gender were associated with increased risk of rejection. The volume of lung function lost did not differentiate between those who did and did not suffer rejection;we hypothesise due to non-alloimmune inflammatory processes.

Hospitalisation and death in lung transplant recipients following COVID-19 infection

Introduction/Aim: Rates of hospitalisation and death from COVID-19 in lung transplant (LTx) recipients vary internationally. We aimed to assess risk factors for this in an Australian cohort. Method(s): We performed a retrospective cohort study of all LTx recipients between January 2020 and September 2022. LTx recipients with COVID-19 were included. Baseline characteristics and treatments were recorded. Multivariate logistic regression was performed to identify risk factors associated with hospitalisation and death. Result(s): 128/387 (33%) recipients tested positive to SARS-CoV-2 during the study period, 97.6% during the Omicron waves with 40(31.3%) requiring hospitalisation and 10 (7.8%) died. The median (IQR) recipient age was 50.6 (22-77). The cohort was of Caucasian ethnicity 105 (82%), 48% were female with high vaccination rates (98.4%). Chronic lung allograft dysfunction (CLAD) was present in 48 (37.5%). 103 (80.5%) of patients received early SARS-CoV-2 treatment with either Sotrovimab 84(65%), Molnupirivir 50(39%) or combination 31(24%). 25 patients (19.5%) received no early treatment. All hospitalised patients received Remdesivir and Dexamethasone as per local treatment protocols. Regarding risk of hospitalisation, multivariate analysis showed that recipient age (1-unit change OR 1.04 95% CI 1.01-1.07 p = 0.019) was associated with an increased risk, whereas Molnupiravir was protective (OR 0.32 95% CI 0.13-0.80 p = 0.02). In univariable analysis, increasing age (1-unit change, OR 1.07 95% CI 1.02-1.129 p = 0.01) and severe disease (OR 9.95 95% CI 2.58-38.32 p =< 0.001) were associated with an increased risk of death. Male gender, non-Caucasian ethnicity, CLAD, CKD stage 3-5 were correlated with death with weak association. Conclusion(s): Recipient age is a significant risk factor for both hospitalisation and death, and older patients with COVID-19 should be monitored closely during COVID-19 illness. Molnupirivir is protective against hospitalisation, with Sotrovimab having a weak association. Further analysis of the protective effect of pre-exposure prophylaxis with emerging therapies such as Evusheld would be helpful to fully evaluate the currently available early disease therapies in Australia.

COVID pneumonitis, a risk factor for persistent lung allograft dysfunction

Introduction/Aim: There is a paucity of data regarding the impact of COVID-19 on allograft function in lung transplant (LTx) recipients. Method(s): We performed a retrospective cohort study of all living LTx recipients in our service between January 2020 and September 2022. Patients with COVID-19 were identified and baseline characteristics recorded. Pre- and post-COVID-19 spirometry was used to identify persistent decline in allograft function (>=10% of FEV 1 decline at 90 days after infection and failure to return to baseline during the study period). Multivariable logistic regression was performed to identify risk factors associated with persistent allograft decline. Result(s): 128/387 (33%) LTx recipients tested positive to SARS-CoV-2 during the study period. The majority, 125 (97%) during the Omicron waves. In those with COVID-19, the median (IQR) recipient age was 50.6 (22-77) with median time post-transplant of 1522 (17-9842) days. The cohort was of Caucasian ethnicity, 105 (82%), with vaccination rates (98.4%) and 48% female. Chronic lung allograft dysfunction (CLAD) was present at time of infection in 48 (37.5%). Severe disease (oxygen requirement) was present in 40 (31%) patients and 10 (7.8%) died. Recipients were followed for median of 172 days (range 90-339) post infection. Persistent FEV 1 decline occurred in 37 (31.4%). Multivariate analysis showed severe disease was independently associated with an increased risk of persistent FEV 1 decline (OR 5.55 [95% CI 2.28-13.48] p =< 0.001). Non-Caucasian ethnicity (OR 2.83, [95% CI 0.92-8.65], p = 0.07) and the presence of CLAD (OR 2.39, [95% CI 0.94-6.08], p = 0.06), were positively correlated, with weak association. No significant association between recipient age, gender, time post-transplant, early COVID therapy, SARS-CoV-2 variant with persistent FEV 1 decline was seen. Conclusion(s): Persistent decline in lung allograft function is common post COVID-19 infection. Severe disease is strongly associated with this outcome and these patients should be monitored for poor long-term allograft recovery. Further investigation into pathological mechanisms responsible for persistent allograft decline is required.

SARS-CoV-2 VARIANTS-DEPENDENT INFLAMMASOME ACTIVATION

Background: Innate immunity is the first line of defense in response to pathogens, which acts locally and also leads the stimulation of adaptive immunity through at least with IL-1beta secretion. It has been shown that SARSCoV- 2 infection triggered the NLRP-3 inflammasome activation and the IL-1beta secretion. The aim of this study was to analyze and compare the level of IL-1beta secretion that is one of the most important innate immunity cytokines, in monocyte-like cells infected with 6 different variants of the SARS-CoV-2. Method(s): Six SARS-CoV-2 variants (historical (B.1, D614G), Alpha, Beta, Gamma, Delta and Omicron BA.1) were isolated from COVID-19 hospitalized patients. Viral stocks were obtained by inoculation in Vero and Vero-TRMPSS2 cells. THP-1 monocyte-like cells were cultured with RPMI-hepes 10% FBS-0.05 mM 2-mercaptoethanol. A total of 5 x 104 of THP-1 cells was plated per well in 96-wells plate and differentiated with 10nM of PMA for 24h. Differenciated- THP-1 were first primed with LPS 1mug/ml for 2h and infected with different SARS-CoV-2 variants with a MOI 0.1. IL-1beta was measured by luminescence in the supernatant after 24 h of infection. Result(s): We analyzed and compared IL-1beta secretion between SARS-CoV-2 virus 6 sublineages after infection of monocytes like THP-1. We observed that THP-1 cells infected with SARS-CoV-2 variants presented a significantly higher IL-1beta secretion than non-infected cells. Moreover, some SARS-CoV-2 variants led to a stronger IL-1beta secretion, and particularly we observed a significantly higher level of IL-1beta cells infected with Omicron BA.1 sublineage compared to other variants. Indeed, Omicron BA.1 infected cells presented the higher IL-1beta secretion (median 385.7 pg/ml IQR[302.6-426.3]) follows by the Delta variants and the historical variants (median 303.6 [266.3-391.9] and 281.9 [207.2-410], respectively). Alpha, Beta and Gamma variants presented the lowest IL-1beta secretion (median 228.1 [192.5-276.4], 219.1 [185.1-354.2] and 211 [149.8- 228.8]). Conclusion(s): We observed the inflammasome activation for the 6 SARS-CoV-2 sublineages with a variation in level of IL-1beta secretion. Indeed, our results suggested that Omicron BA.1 was more recognized by the innate immune cells than other SARS-CoV-2, which could in part, with its upper respiratory tract tropism, possibly explain its less clinical virulence. Taking together, these results suggest that the innate immunity response and precisely, IL-1beta secretion pathways were activated in a SARS-CoV-2 variants-dependent manner.

Hiv Detection in Wastewater as a New Epidemiological Tool

Background: Wastewater represents a broad, immediate, and unbiased accounting of the pathgens in the population. We aimed to develop methods to track HIV in wastewater utilizing a viral detection pipeline adapted from platforms developed to track SARS-COV-2. Method(s): We used samples from 6 wastewater treatment plants in the Houston area. We focused on regions of higher prevalence and lower prevalence. First, employing wastewater processing and nucleic acid extraction methods described by our group to detect SARS-COV-2, we tested a single high and low prevalence site in triplicate with all 3 primer sets. nucleic acid extracts from HIV and SIV cell culture supernatants were used as controls. Next, in subsequent samples, RT-PCR reactions with detections were subjected to gel electrophoresis to determine the amplified product sizes. To further confirm HIV detection, we sequenced the RT-PCR products and compared the proportion of reads which mapped to the expected amplified product. In a later set of studies, we fractionated samples into supernatant and pellet. We further tested HIV presence by performing whole virome sequencing on the extracts from some samples that produced detections and mapped reads to published genomes. A crAssphage genome was used as a negative control. Result(s): Samples from all sites resulted in signal detection at least once. Only reactions with gag and pol primers appeared to amplify the expected product. Products from the HIV positive control mapped almost exclusively to the HIV genome (97-100% of reads), with a fraction of reads from the SIV negative control doing the same (16-18% of reads). The ltr and pol products did not map the HIV genome while gag products did (34-44% of reads). Among the fractionated sample, in total, 6 supernatant fractions produced no detection compared to 7 of 8 pellet fractions. The whole virome sequencing produced reads that mapped to the HIV genome with at least 8X depth coverage. The sample with the lowest Ct detection (26) yielded HIV coverage several logs greater than those samples with higher Ct detection (37). Reads from all samples mapped to at least 20% of the HIV genome. Conclusion(s): This work provides the first evidence that HIV can be detected in municipal wastewater systems and has the potential to be developed into a new public health tool.

Placental Thrombosis Resulting in Hypoxic Ischemic Encephalopathy in an Infant of a Mother with Covid-19

Case Report: Since the beginning of the Coronavirus Disease 2019 (COVID-19) pandemic, there has been much work to understand the negative effects of SARS-CoV-2 on tissues expressing the Angiotensin Converting Enzyme-2 (ACE2) receptor, including the placenta. However, there is limited information regarding placental pathology findings in mothers with COVID-19 and the effects of SARS-CoV-2 on the placenta. The available research reports effects on the fetus ranging from minimal to intrauterine fetal demise. Case Description: A 4680g baby boy was born at 38+1 weeks of gestation to 36y old G4P1021 female via repeat cesarian section. The pregnancy was complicated by advanced maternal age, chronic hypertension with superimposed pre-eclampsia with severe features, BMI of 80, and SARS-CoV-2 infection. The mother had mild COVID-19 symptoms and did not require hospitalization or oxygen support. Prenatal ultrasounds were limited due to body habitus. At the time of delivery, there was clear amniotic fluid. Upon delivery the infant was cyanotic and limp and was brought to the warmer immediately. Non-invasive positive pressure ventilation was initiated at 5 minutes of life with improvement in infant color and oxygen saturation. He was then admitted to the Neonatal Intensive Care Unit (NICU). APGARs were 2, 3, 5, and 7 at 1, 5, 10, and 15 minutes respectively. Cord gases showed severe metabolic acidosis. The patient was diagnosed with hypoxic-ischemic encephalopathy (HIE) and therapeutic hypothermia was initiated. Both the NICU and obstetric teams were unable to identify a clear perinatal cause of HIE in this patient. Later, the placenta pathology report revealed a large placenta for estimated gestational age corresponding to the 75th percentile, villous parenchyma with focal chorangiosis and thrombi, with unremarkable fetal membrane and three vessel umbilical cord. The cause of HIE was then thought to be due to the placental thrombi likely caused by SARS-CoV-2 infection. Discussion(s): Fetal vascular malperfusion and fetal vascular thrombus have been noted as a common finding in the placentas of pregnant women who test positive for SARS-CoV-2. There are various causes of HIE, from maternal, placental and fetal factors. This patient had no clinically evident hypoxic event, but information was limited due to the lack of monitoring of the fetus in utero. Given the mother's SARS-CoV-2 infection and the placental pathology findings, it is likely that the cause of this patient's HIE was related to the effects on the placenta from SARS-CoV-2. Conclusion(s): As more information comes to light about the effects of SARS-CoV-2 on the placenta, it is important to consider a maternal SARS-CoV-2 infection during pregnancy as a cause of HIE in a newborn.