In vivo secondary structural analysis of Influenza A virus genomic RNA.

Influenza A virus (IAV) is a respiratory virus that causes epidemics and pandemics. Knowledge of IAV RNA secondary structure in vivo is crucial for a better understanding of virus biology. Moreover, it is a fundament for the development of new RNA-targeting antivirals. Chemical RNA mapping using selective 2'-hydroxyl acylation analyzed by primer extension (SHAPE) coupled with Mutational Profiling (MaP) allows for the thorough examination of secondary structures in low-abundance RNAs in their biological context. So far, the method has been used for analyzing the RNA secondary structures of several viruses including SARS-CoV-2 in virio and in cellulo. Here, we used SHAPE-MaP and dimethyl sulfate mutational profiling with sequencing (DMS-MaPseq) for genome-wide secondary structure analysis of viral RNA (vRNA) of the pandemic influenza A/California/04/2009 (H1N1) strain in both in virio and in cellulo environments. Experimental data allowed the prediction of the secondary structures of all eight vRNA segments in virio and, for the first time, the structures of vRNA5, 7, and 8 in cellulo. We conducted a comprehensive structural analysis of the proposed vRNA structures to reveal the motifs predicted with the highest accuracy. We also performed a base-pairs conservation analysis of the predicted vRNA structures and revealed many highly conserved vRNA motifs among the IAVs. The structural motifs presented herein are potential candidates for new IAV antiviral strategies.

Differences in BNT126b2 and ChAdOx1 Homologous Vaccination Antibody Response among Teachers in Poznan, Poland.

Children are among the best vectors to spread respiratory viruses, including emerging variants of SARS-CoV-2 due to the asymptomatic or relatively mild course of infection and simultaneously high titres of pathogens in the respiratory tract. Therefore, individuals who have constant contact with children, e.g., teachers should be vaccinated against COVID-19 as essential workers within the first phases of a vaccination campaign. In Poland, primary and secondary school teachers were vaccinated with ChAdOx1 from February 2021 with a three month interval between the two doses, while lecturers at medical universities, who are simultaneously healthcare workers, received the BNT126b2 vaccine from December 2020 with three weeks between the first and second doses. The aim of this study was to compare the antibody responses at two weeks and three months after vaccination and to estimate the vaccine effectiveness against COVID-19 among infection-naïve teachers vaccinated with mRNA and a vector vaccine. We found that the anti-SARS-CoV-2 spike protein antibodies were significantly higher among the lecturers but antibody waning was slower among the schoolteachers. However, those vaccinated with ChAdOx1 complained significantly more often of vaccine side effects. In addition, during the three months after the second vaccine dose no study participants were infected with SARS-CoV-2. The BNT126b2 vaccine gave higher antibody titres in comparison with ChAdOx1 but protection against COVID-19 in both cases was similar. Moreover, we did not find any anti-SARS-CoV-2 nucleoprotein antibodies at two weeks as well as at three months after vaccination among the study participants, which shows a very high vaccine effectiveness in the occupational group with a high SARS-CoV-2-infection risk.

The Longitudinal Analysis on the Anti-SARS-CoV-2 Antibodies among Healthcare Workers in Poland-Before and after BNT126b2 mRNA COVID-19 Vaccination.

One of the groups most vulnerable to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is healthcare workers (HCWs) who have direct contact with suspected and confirmed coronavirus diseases 2019 (COVID-19) patients. Therefore, this study aimed to (i) conduct a longitudinal analysis of the seroprevalence of SARS-CoV-2 infection among HCWs working in two healthcare units (HCUs) in Poland and (ii) identify anti-SARS-CoV-2 IgG antibody (Ab) response factors following infection and anti-COVID-19 vaccination. The overall seroprevalence increased from 0% at baseline in September 2020 to 37.8% in December 2020. It reached 100% in February 2021 after BNT126b2 (Pfizer New York, NY, USA/BioNTech Mainz, Germany) full vaccination and declined to 94.3% in September 2021. We observed significant differences in seroprevalence between the tested high- and low-risk infection HCUs, with the highest seropositivity among the midwives and nurses at the Gynecology and Obstetrics Ward, who usually have contact with non-infectious patients and may not have the proper training, practice and personal protective equipment to deal with pandemic infections, such as SARS-CoV-2. We also found that anti-SARS-CoV-2 Ab levels after coronavirus infection were correlated with disease outcomes. The lowest Ab levels were found among HCWs with asymptomatic coronavirus infections, and the highest were found among HCWs with severe COVID-19. Similarly, antibody response after vaccination depended on previous SARS-CoV-2 infection and its course: the highest anti-SARS-CoV-2 Ab levels were found in vaccinated HCWs after severe COVID-19. Finally, we observed an approximately 90-95% decrease in anti-SARS-CoV-2 Ab levels within seven months after vaccination. Our findings show that HCWs have the highest risk of SARS-CoV-2 infection, and due to antibody depletion, extra protective measures should be undertaken. In addition, in the context of the emergence of new pathogens with pandemic potential, our results highlight the necessity for better infectious disease training and regular updates for the low infection risk HCUs, where the HCWs have only occasional contact with infectious patients.

Prevalence of Anti-SARS-CoV-2 Antibodies in Poznan, Poland, after the First Wave of the COVID-19 Pandemic.

In comparison to other European countries, during the first months of the COVID-19 pandemic, Poland reported a relatively low number of confirmed cases of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections. To estimate the scale of the pandemic in Poland, a serosurvey of antibodies against SARS-CoV-2 was performed after the first wave of COVID-19 in Europe (March-May 2020). Within this study, we collected samples from 28 July to 24 September 2020 and, based on the ELISA results, we found that 1.67% (25/1500, 95% CI 1.13-2.45) of the Poznan (Poland) metropolitan area's population had antibodies against SARS-CoV-2 after the first wave of COVID-19. However, the presence of anti-SARS-CoV-2 IgG antibodies was confirmed with immunoblotting in 56% (14/25) samples, which finally resulted in a decrease in seroprevalence, i.e., 0.93% (14/1500, 95% CI 0.56-1.56). The positive anti-SARS-CoV-2 IgG results were associated with age, occupation involving constant contact with people, travelling abroad, non-compliance with epidemiological recommendations and direct contact with the novel coronavirus. Our findings confirm the low SARS-CoV-2 incidence in Poland and imply that the population had little herd immunity heading into the second and third wave of the pandemic, and therefore, that herd immunity contributed little to preventing the high numbers of SARS-CoV-2 infections and COVID-19-related deaths in Poland during these subsequent waves.

A novel Bacillus thuringiensis Cry9Ea-like protein with high insecticidal activity towards Cydia pomonella larvae.

BACKGROUND: The host specificity of known Bacillus thuringiensis Cry and Vip pesticidal proteins still needs extensive investigation and the proteins currently used in crop protection are not effective against many pest species. Cydia pomonella (L.) is a widespread and economically important pest of apples, very difficult to control, since the larvae spend most of their life inside a fruit. Currently, large amounts of broad-spectrum, detrimental synthetic agents are used to combat this herbivore and therefore biopesticides with high activity against C. pomonella are very much needed. RESULTS: The toxicity of B. thuringiensis Cry9Ea along with five distinct pesticidal proteins (Cry1Aa, Cry1Ca, Cry1Ia, Cry2Ab and Vip3Aa) has been determined towards the first-instar larvae of C. pomonella. Cry9Ea has much higher activity than the remaining tested proteins (30-1200-fold lower LC50 ) and possibly is the most potent B. thuringiensis pesticidal protein bioassayed against C. pomonella so far. In contrast, Cry9Ea is not toxic towards Spodoptera exigua (Hübn.), indicating a potentially narrow spectrum of activity. Both insect species show high variability in susceptibility to the remaining Cry/Vip proteins. CONCLUSIONS: The obtained results extend the existing knowledge regarding B. thuringiensis pesticidal protein host range and indicate Cry9Ea as a promising candidate for successful biological control of C. pomonella. © 2020 Society of Chemical Industry.

RNA Secondary Structure as a First Step for Rational Design of the Oligonucleotides towards Inhibition of Influenza A Virus Replication.

Influenza is an important research subject around the world because of its threat to humanity. Influenza A virus (IAV) causes seasonal epidemics and sporadic, but dangerous pandemics. A rapid antigen changes and recombination of the viral RNA genome contribute to the reduced effectiveness of vaccination and anti-influenza drugs. Hence, there is a necessity to develop new antiviral drugs and strategies to limit the influenza spread. IAV is a single-stranded negative sense RNA virus with a genome (viral RNA-vRNA) consisting of eight segments. Segments within influenza virion are assembled into viral ribonucleoprotein (vRNP) complexes that are independent transcription-replication units. Each step in the influenza life cycle is regulated by the RNA and is dependent on its interplay and dynamics. Therefore, viral RNA can be a proper target to design novel therapeutics. Here, we briefly described examples of anti-influenza strategies based on the antisense oligonucleotide (ASO), small interfering RNA (siRNA), microRNA (miRNA) and catalytic nucleic acids. In particular we focused on the vRNA structure-function relationship as well as presented the advantages of using secondary structure information in predicting therapeutic targets and the potential future of this field.