Meat Exudate for Detection of African Swine Fever Virus Genomic Material and Anti-ASFV Antibodies

African swine fever (ASF) is one of the most important viral diseases of pigs caused by the ASF virus (ASFV). The virus is highly stable over a wide range of temperatures and pH and can survive in meat and meat products for several months, leading to long-distance transmission of ASF. Whole blood, serum, and organs from infected pigs are used routinely as approved sample types in the laboratory diagnosis of ASF. However, these sample types may not always be available. Here, we investigated meat exudate as an alternative sample type for the detection of ASFV-specific nucleic acids and antibodies. Pigs were infected with various ASFV strains: the highly virulent ASFV Malawi LIL 18/2 strain, the moderately-virulent ASFV Estonia 2014 strain, or the low-virulent ASFV OURT/88/3 strain. The animals were euthanized on different days post-infection (dpi), and meat exudates were collected and tested for the presence of ASFV-specific nucleic acids and antibodies. Animals infected with the ASFV Malawi LIL 18/2 developed severe clinical signs and succumbed to the infection within seven dpi, while pigs infected with ASFV Estonia 2014 also developed clinical signs but survived longer, with a few animals seroconverting before succumbing to the ASFV infection or being euthanized as they reached humane endpoints. Pigs infected with ASFV OURT/88/3 developed transient fever and seroconverted without mortality. ASFV genomic material was detected in meat exudate from pigs infected with ASFV Malawi LIL 18/2 and ASFV Estonia 2014 at the onset of viremia but at a lower amount when compared to the corresponding whole blood samples. Low levels of ASFV genomic material were detected in the whole blood of ASFV OURT/88/3-infected pigs, and no ASFV genomic material was detected in the meat exudate of these animals. Anti-ASFV antibodies were detected in the serum and meat exudate derived from ASFV OURT/88/3-infected pigs and in some of the samples derived from the ASFV Estonia 2014-infected pigs. These results indicate that ASFV genomic material and anti-ASFV antibodies can be detected in meat exudate, indicating that this sample can be used as an alternative sample type for ASF surveillance when routine sample types are unavailable or are not easily accessible.

Spatial separation of ribosomes and DNA in Asgard archaeal cells

The origin of the eukaryotic cell is a major open question in biology. Asgard archaea are the closest known prokaryotic relatives of eukaryotes, and their genomes encode various eukaryotic signature proteins, indicating some elements of cellular complexity prior to the emergence of the first eukaryotic cell. Yet, microscopic evidence to demonstrate the cellular structure of uncultivated Asgard archaea in the environment is thus far lacking. We used primer-free sequencing to retrieve 715 almost full-length Loki- and Heimdallarchaeota 16S rRNA sequences and designed novel oligonucleotide probes to visualize their cells in marine sediments (Aarhus Bay, Denmark) using catalyzed reporter deposition-fluorescence in situ hybridization (CARD-FISH). Super-resolution microscopy revealed 1–2 µm large, coccoid cells, sometimes occurring as aggregates. Remarkably, the DNA staining was spatially separated from ribosome-originated FISH signals by 50–280 nm. This suggests that the genomic material is condensed and spatially distinct in a particular location and could indicate compartmentalization or membrane invagination in Asgard archaeal cells.

Expanding the classical paradigm: what we have learnt from vertebrates about sex chromosome evolution

Until recently, the field of sex chromosome evolution has been dominated by the canonical unidirectional scenario, first developed by Muller in 1918. This model postulates that sex chromosomes emerge from autosomes by acquiring a sex-determining locus. Recombination reduction then expands outwards from this locus, to maintain its linkage with sexually antagonistic/advantageous alleles, resulting in Y or W degeneration and potentially culminating in their disappearance. Based mostly on empirical vertebrate research, we challenge and expand each conceptual step of this canonical model and present observations by numerous experts in two parts of a theme issue of Phil. Trans. R. Soc. B. We suggest that greater theoretical and empirical insights into the events at the origins of sex-determining genes (rewiring of the gonadal differentiation networks), and a better understanding of the evolutionary forces responsible for recombination suppression are required. Among others, crucial questions are: Why do sex chromosome differentiation rates and the evolution of gene dose regulatory mechanisms between male versus female heterogametic systems not follow earlier theory? Why do several lineages not have sex chromosomes? And: What are the consequences of the presence of (differentiated) sex chromosomes for individual fitness, evolvability, hybridization and diversification? We conclude that the classical scenario appears too reductionistic. Instead of being unidirectional, we show that sex chromosome evolution is more complex than previously anticipated and principally forms networks, interconnected to potentially endless outcomes with restarts, deletions and additions of new genomic material. This article is part of the theme issue ‘Challenging the paradigm in sex chromosome evolution: empirical and theoretical insights with a focus on vertebrates (Part II)’.

Genome sequencing and de novo assembly of the giant unicellular alga Acetabularia acetabulum using droplet MDA

The macroscopic single-celled green alga Acetabularia acetabulum has been a model system in cell biology for more than a century. However, no genomic information is available from this species. Since the alga has a long life cycle, is difficult to grow in dense cultures, and has an estimated diploid genome size of almost 2 Gb, obtaining sufficient genomic material for genome sequencing is challenging. Here, we have attempted to overcome these challenges by amplifying genomic DNA using multiple displacement amplification (MDA) combined with microfluidics technology to distribute the amplification reactions across thousands of microscopic droplets. By amplifying and sequencing DNA from five single cells we were able to recover an estimated ~ 7–11% of the total genome, providing the first draft of the A. acetabulum genome. We highlight challenges associated with genome recovery and assembly of MDA data due to biases arising during genome amplification, and hope that our study can serve as a reference for future attempts on sequencing the genome from non-model eukaryotes.

Mechanisms for Chromosome Segregation in Bacteria

The process of DNA segregation, the redistribution of newly replicated genomic material to daughter cells, is a crucial step in the life cycle of all living systems. Here, we review DNA segregation in bacteria which evolved a variety of mechanisms for partitioning newly replicated DNA. Bacterial species such as Caulobacter crescentus and Bacillus subtilis contain pushing and pulling mechanisms that exert forces and directionality to mediate the moving of newly synthesized chromosomes to the bacterial poles. Other bacteria such as Escherichia coli lack such active segregation systems, yet exhibit a spontaneous de-mixing of chromosomes due to entropic forces as DNA is being replicated under the confinement of the cell wall. Furthermore, we present a synopsis of the main players that contribute to prokaryotic genome segregation. We finish with emphasizing the importance of bottom-up approaches for the investigation of the various factors that contribute to genome segregation.

Pitfalls in Tick and Tick-Borne Pathogens Research, Some Recommendations and a Call for Data Sharing

An understanding of the relationships of ticks and tick-borne pathogens can only be achieved by integrating data from multiple studies. The publication of raw material is a necessary step for wide-area meta-analyses and study design, data collection and reporting require harmonization. This is an opinion paper, not a consensus position, and is open to debate. This work reflects our view about how data should be communicated in mainstream journals. We indicate rules that should be observed in recording weather data, to avoid serendipitous correlations between the density of ticks and climate variables and recommend the inclusion of raw data in reports. We stress the need for standardized methods to collect ticks that cannot be obtained by standard flagging. The reporting of infection rates of pathogens in ticks should avoid conclusions based on pure molecular findings in feeding ticks. Studies demonstrating the vectorial capacity of ticks should not be supported only by molecular surveys of feeding ticks. Vacuous conclusions about vectorial or reservoir status based solely on the finding of genomic material of a pathogen should be discouraged. We stress that phylogenetic studies based on random selection of sequences from GenBank are unsuitable. We firmly support the development of a dedicated server of curated sequences of ticks and pathogens as a standard for future studies.

VACCINE RACE TO BECOME A REALITY AGAINST THE HIGHLY INFECTIOUS CORONAVIRUS (COVID19)

COVID-19, is a global pandemic started in November 2019, extending beyond its biological scale to infect lives globally by inserting their genomic material inside the host cells. This global threat has till now claimed numerous lives with no possible full proof cure till date. There is no unanimity amongst the countries in the design and development of specific vaccines to cure COVID-19 and its distribution among the global population. India Government acquired “cold chain management” for safe and even distribution of present vaccines (Covaxin, Covisheild). Here, authors intend to put some light on the brief introduction, stages and perspective of cold chain logistics of vaccine distribution in India with 150crores of population.

HIV-1 Capsid Core: A Bullet to the Heart of the Target Cell

The first step of the intracellular phase of retroviral infection is the release of the viral capsid core in the cytoplasm. This structure contains the viral genetic material that will be reverse transcribed and integrated into the genome of infected cells. Up to recent times, the role of the capsid core was considered essentially to protect this genetic material during the earlier phases of this process. However, increasing evidence demonstrates that the permanence inside the cell of the capsid as an intact, or almost intact, structure is longer than thought. This suggests its involvement in more aspects of the infectious cycle than previously foreseen, particularly in the steps of viral genomic material translocation into the nucleus and in the phases preceding integration. During the trip across the infected cell, many host factors are brought to interact with the capsid, some possessing antiviral properties, others, serving as viral cofactors. All these interactions rely on the properties of the unique component of the capsid core, the capsid protein CA. Likely, the drawback of ensuring these multiple functions is the extreme genetic fragility that has been shown to characterize this protein. Here, we recapitulate the busy agenda of an HIV-1 capsid in the infectious process, in particular in the light of the most recent findings.

Environmental (e)RNA advances the reliability of eDNA by predicting its age

Environmental DNA (eDNA) analysis has advanced conservation biology and biodiversity management. However, accurate estimation of age and origin of eDNA is complicated by particle transport and the presence of legacy genetic material, which can obscure accurate interpretation of eDNA detection and quantification. To understand the state of genomic material within the environment, we investigated the degradation relationships between (a) size of fragments (long vs short), (b) genomic origins (mitochondrial vs nuclear), (c) nucleic acids (eDNA vs eRNA), and (d) RNA types (messenger (m)RNA vs ribosomal (r)RNA) from non-indigenous Dreissena mussels. Initial concentrations of eRNA followed expected transcriptional trends, with rRNAs found at > 1000 × that of eDNA, and a mitosis-associated mRNA falling below detection limits within 24 h. Furthermore, the ratio of eRNA:eDNA significantly decreased throughout degradation, potentially providing an estimate for the age of genomic material. Thus, eRNA quantification can increase detection due to the high concentrations of rRNAs. Furthermore, it may improve interpretation of positive detections through the eRNA:eDNA ratio and/or by detecting low abundant mitosis-associated mRNAs that degrade within ~ 24 h.

Development of approaches to the diagnosis of cattle leukemia in the system of antiepizootic measures in the Belgorod Region

The aim of the research is the use of serological and molecular genetic methods for detecting virus-infected cattle leukemia, as well as determining the significance of PCR in identifying BLV infected calves in the system of antiepizootic health measures. The developed technique for early diagnosis of leukemia in cattle made it possible to accelerate the process of recovery of disadvantaged farms in the Belgorod region by increasing the frequency of studies from 6 months to 2-3 months and an increase in the sensitivity of the agar-gel immunodiffusion test. This, in turn, leads to an increase in the sensitivity of the agar-gel immunodiffusion test and makes it possible to detect, on average, from 8.8% to 20.25% more animals infected with the leukemia virus compared to the standard reaction of the agar-gel immunodiffusion test. The additional use of molecular genetic tests for the detection of proviral DNA of the leukemia virus makes it possible to identify at the early stages of the development of the leukemia process, in calves from 15 days of age, the genomic material of bovine leukemia virus, which will also allow in a shorter time to carry out a qualitative improvement of young cattle in dysfunctional farms.