A Non-phylogeny-dependent Reassortment Detection Method for Influenza A Viruses

Influenza A virus is a segmented RNA virus whose genome consists of 8 single-stranded negative-sense RNA segments. This unique genetic structure allows viruses to exchange their segments through reassortment when they infect the same host cell. Studying the determination and nature of influenza A virus reassortment is critical to understanding the generation of pandemic strains and the spread of viruses across species. Reassortment detection is the first step in influenza A virus reassortment research. Several methods for automatic detection of reassortment have been proposed, which can be roughly divided into two categories: phylogenetic methods and distance methods. In this article, we proposed a reassortment detection method that does not require multiple sequence alignment and phylogenetic analysis. We extracted the codon features from the segment sequence and expressed the sequence as a feature vector, and then used the clustering method of self-organizing map to cluster the sequence for each segment. Based on the clustering results and the epidemiological information of the virus, the reassortment detection was implemented. We used this method to perform reassortment detection on the collected 7,075 strains from Asia and identified 516 reassortment events. We also conducted a statistical analysis of the identified reassortment events and found conclusions consistent with previous studies. Our method will provide new insights for automating reassortment detection tasks and understanding the reassortment patterns of influenza A viruses.

Perspectives on Assembling Coronavirus Spikes on Fiber Optics to Reveal Broadly Recognizing Antibodies and Generate a Universal Coronavirus Detector

In time of COVID-19 biological detection technologies are of crucial relevance. We propose here the use of state of the art optical fiber biosensors to address two aspects of the fight against SARS-CoV-2 and other pandemic human coronaviruses (HCoVs). Fiber optic biosensors functionalized with HCoV spikes could be used to discover broadly neutralizing antibodies (bnAbs) effective against known HCoVs (SARS-CoV, MERS-CoV and SARS-CoV-2) and likely future ones. In turn, identified bnAbs, once immobilized onto fiber optic biosensors, should be capable to detect HCoVs as diagnostic and environmental sensing devices. The therapeutic and preventative value of bnAbs is immense as they can be used for passive immunization and for the educated development of a universal vaccine (active immunization). Hence, HCoV bnAbs represent an extremely important resource for future preparedness against coronavirus-borne pandemics. Furthermore, the assembly of bnAb-based biosensors constitutes an innovative approach to counteract public health threats, as it bears diagnostic competence additional to environmental detection of a range of pandemic strains. This concept can be extended to different pandemic viruses, as well as bio-warfare threats that entail existing, emerging and extinct viruses (e.g., the smallpox-causing Variola virus). We report here the forefront fiber optic biosensor technology that could be implemented to achieve these aims.

Enhanced Immunogenicity of a Whole-Inactivated Influenza A Virus Vaccine Using Optimised Irradiation Conditions

Influenza A virus presents a constant pandemic threat due to the mutagenic nature of the virus and the inadequacy of current vaccines to protect against emerging strains. We have developed a whole-inactivated influenza vaccine using γ-irradiation (γ-Flu) that can protect against both vaccine-included strains as well as emerging pandemic strains. γ-irradiation is a widely used inactivation method and several γ-irradiated vaccines are currently in clinical or pre-clinical testing. To enhance vaccine efficacy, irradiation conditions should be carefully considered, particularly irradiation temperature. Specifically, while more damage to virus structure is expected when using higher irradiation temperatures, reduced radiation doses will be required to achieve sterility. In this study, we compared immunogenicity of γ-Flu irradiated at room temperature, chilled on ice or frozen on dry ice using different doses of γ-irradiation to meet internationally accepted sterility assurance levels. We found that, when irradiating at sterilising doses, the structural integrity and vaccine efficacy were well maintained in all preparations regardless of irradiation temperature. In fact, using a higher temperature and lower radiation dose appeared to induce higher neutralising antibody responses and more effective cytotoxic T cell responses. This outcome is expected to simplify irradiation protocols for manufacturing of highly effective irradiated vaccines.

Vibrio cholerae El Tor strains linked to global cholera are homogeneous by pulsed-field gel electrophoresis

Vibrio cholerae O1 El Tor, causative agent of the ongoing seventh cholera pandemic, is native to the aquatic environment of the Ganges Delta, Bay of Bengal (GDBB). Recent studies traced pandemic strains to the GDBB and proposed global spread of cholera had occurred via intercontinental transmission. In the research presented here, Not I-digested genomic DNA extracted from V. cholerae O1 clinical and environmental strains isolated in Bangladesh during 2004 – 2014 was analyzed by pulsed-field gel electrophoresis (PFGE). Results of cluster analysis showed 94.67% of the V. cholerae isolates belonged to clade A and included the majority of clinical isolates of spatio-temporal origin and representing different cholera endemic foci. The rest of the strains were estuarine, all environmental isolates from Mathbaria, Bangladesh, and occurred as singletons, clustered in clades B and C, or in the small clades D and E. Cluster analysis of the Bangladeshi strains and including 157 El Tor strains from thirteen countries in Asia, Africa, and the Americas revealed 85% of the total set of isolates belonged to clade A, indicating all were related, yet did not form an homogeneous cluster. Overall, 15% of the global strains comprised multiple small clades or segregated as singletons. Three sub-clades could be discerned within the major clade A, reflecting distinct lineages of V. cholerae El Tor associated with cholera in Asia, Africa, and the Americas. The presence in Asia and the Americas of non-pandemic V. cholerae El Tor populations differing by PFGE and from strains associated with cholera globally suggests different ecotypes are resident in distant geographies.

Type VI secretion system mutations reduced competitive fitness of classical Vibrio cholerae biotype

The gram-negative bacterium Vibrio cholerae is the causative agent of the diarrhoeal disease cholera and is responsible for seven recorded pandemics. Several factors are postulated to have led to the decline of 6th pandemic classical strains and the rise of El Tor biotype V. cholerae, establishing the current 7th pandemic. We investigated the ability of classical V. cholerae of the 2nd and 6th pandemics to engage their type six secretion system (T6SS) in microbial competition against non-pandemic and 7th pandemic strains. We report that classical V. cholerae underwent sequential mutations in T6SS genetic determinants that initially exposed 2nd pandemic strains to microbial attack by non-pandemic strains and subsequently caused 6th pandemic strains to become vulnerable to El Tor biotype V. cholerae intraspecific competition. The chronology of these T6SS-debilitating mutations agrees with the decline of 6th pandemic classical strains and the emergence of 7th pandemic El Tor V. cholerae.

Coding-Complete Genome Sequence of Swine Influenza Virus Isolate A/Swine/Karaganda/04/2020 (H1N1) from Kazakhstan

Here, we report the coding-complete genome sequence of a clinical sample of influenza virus obtained from a pig at a livestock farm in Karaganda, Central Kazakhstan, during a pig study in 2020. Isolate A/Swine/Karaganda/04/2020 (H1N1) belongs to clade 1A.3.2.2 lineage 1A, which includes the 2009 H1N1 pandemic strains.

Genomic stability among O3:K6 V. parahaemolyticus pandemic strains isolated between 1996 to 2012 in American countries

Background The V. parahaemolyticus pandemic clone, results in the development of gastrointestinal illness in humans. Toxigenic strains of this species are frequently isolated from aquatic habitats and organisms such as mollusks and crustaceans. Reports on the isolation of the pandemic clone started in 1996, when a new O3:K6 clone was identified in Asia, that rapidly spread worldwide, becoming the predominant clone isolated from clinical cases. In this study whole genome sequencing was accomplished with an Illumina MiniSeq platform, upon six novel V. parahaemolyticus strains, that have been isolated in Mexico since 1998 and three representative genomes of strains that were isolated from reported outbreaks in other American countries, and were deposited in the GenBank. These nine genomes were compared against the reference sequence of the O3:K6 pandemic strain (RIMD 2210633), which was isolated in 1996, to determine sequence differences within American isolates and between years of isolation. Results The results indicated that strains that were isolated at different times and from different countries, were highly genetically similar, among them as well as to the reference strain RIMD 2210633, indicating a high level of genetic stability among the strains from American countries between 1996 to 2012, without significant genetic changes relative to the reference strain RIMD 2210633, which was isolated in 1996 and was considered to be representative of a novel O3:K6 pandemic strain. Conclusions The genomes of V. parahaemolyticus strains isolated from clinical and environmental sources in Mexico and other American countries, presented common characteristics that have been reported for RIMD 2210633 O3:K6 pandemic strain. The major variations that were registered in this study corresponded to genes non associated to virulence factors, which could be the result of adaptations to different environmental conditions. Nevertheless, results do not show a clear pattern with the year or locality where the strains were isolated, which is an indication of a genomic stability of the studied strains.

VicPred: A Vibrio cholerae Genotype Prediction Tool

Genomic information can be used to predict major pathogenic traits of pathogens without the need for laboratory experimentation. However, no Vibrio cholerae genome-based trait identification tools currently exist. The aim of this study was to develop a web-based prediction tool to identify Vibrio pathogenic traits using publicly available 796 whole-genome sequences of V. cholerae. Using this application, 68 structural O-antigen gene clusters belonging to 49 serogroups of V. cholerae were classified, and the composition of the genes within the O-antigen cluster of each serogroup was identified. The arrangement and location of the CTX prophage and related elements of the seventh cholera pandemic strains were also revealed. With the versatile tool, named VicPred, we analyzed the assemblage of various SXTs (sulfamethoxazole/trimethoprim resistance element) and major genomic islands (GIs) of V. cholerae, and the increasing trend in drug-resistance revealing high resistance of the V. cholerae strains to certain antibiotics. The pathogenic traits of newly sequenced V. cholerae strains could be analyzed based on these characteristics. The accumulation of further genome data will expedite the establishment of a more precise genome-based pathogenic traits analysis tool.

A gap-free tomato genome built from complementary PacBio and Nanopore long DNA sequences reveals extensive linkage drag during breeding

The assembly and scaffolding of plant crop genomes facilitates the characterization of genetically diverse cultivated and wild germplasm. The cultivated tomato has been improved through the introgression of genetic material from related wild species, including resistance to pandemic strains of Tobacco Mosaic virus (TMV) from Solanum peruvianum. Here we applied PacBio HiFi and ONT nanopore sequencing to develop independent, highly contiguous and complementary assemblies of an inbred TMV-resistant tomato variety. We merged the HiFi and ONT assemblies to generate a long-read-only assembly where all twelve chromosomes were represented as twelve contiguous sequences (N50=68.5 Mbp). The merged assembly was validated by chromosome conformation capture data and is highly consistent with previous tomato assemblies that made use of genetic maps and HiC for scaffolding. Our long-read-only assembly reveals that a complex series of structural variants linked to the TMV resistance gene likely contributed to linkage drag of a 64.1 Mbp region of the S. peruvianum genome during tomato breeding. We show that this minimal introgression region is present in six cultivated tomato hybrid varieties developed in three commercial breeding programs. Our results suggest that complementary long read technologies can facilitate the rapid generation of near complete genome sequences.

Inhibition of Phosphatidylinositol 3-Kinase by Pictilisib Blocks Influenza Virus Propagation in Cells and in Lungs of Infected Mice

Influenza virus (IV) infections are considered to cause severe diseases of the respiratory tract. Beyond mild symptoms, the infection can lead to respiratory distress syndrome and multiple organ failure. Occurrence of resistant seasonal and pandemic strains against the currently licensed antiviral medications points to the urgent need for new and amply available anti-influenza drugs. Interestingly, the virus-supportive function of the cellular phosphatidylinositol 3-kinase (PI3K) suggests that this signaling module may be a potential target for antiviral intervention. In the sense of repurposing existing drugs for new indications, we used Pictilisib, a known PI3K inhibitor to investigate its effect on IV infection, in mono-cell-culture studies as well as in a human chip model. Our results indicate that Pictilisib is a potent inhibitor of IV propagation already at early stages of infection. In a murine model of IV pneumonia, the in vitro key findings were verified, showing reduced viral titers as well as inflammatory response in the lung after delivery of Pictilisib. Our data identified Pictilisib as a promising drug candidate for anti-IV therapies that warrant further studying. These results further led to the conclusion that the repurposing of previously approved substances represents a cost-effective and efficient way for development of novel antiviral strategies.