Foreign Genetic Material Research Articles

How to survive pig farming: Mechanism of SCCmec element deletion and metabolic stress adaptation in livestock-associated MRSA.

Previous research on methicillin susceptible Staphylococcus aureus (MSSA) belonging to livestock-associated (LA-) sequence type (ST) 398, isolated from pigs and their local surroundings, indicated that differences between these MSSA and their methicillin resistant predecessors (MRSA) are often limited to the absence of the staphylococcal cassette chromosome mec (SCCmec) and few single nucleotide polymorphisms. So far, our understanding on how LA-MRSA endure the environmental conditions associated with pig-farming as well as the putative impact of this particular environment on the mobilisation of SCCmec elements is limited. Thus, we performed in-depth genomic and transcriptomic analyses using the LA-MRSA ST398 strain IMT38951 and its methicillin susceptible descendant. We identified a mosaic-structured SCCmec region including a putative replicative SCCmecVc which is absent from the MSSA chromosome through homologous recombination. Based on our data, such events occur between short repetitive sequences identified within and adjacent to two distinct alleles of the large cassette recombinase genes C (ccrC). We further evaluated the global transcriptomic response of MRSA ST398 to particular pig-farm associated conditions, i.e., contact with host proteins (porcine serum) and a high ammonia concentration. Differential expression of global regulators involved in stress response control were identified, i.e., ammonia-induced alternative sigma factor B-depending activation of genes for the alkaline shock protein 23, the heat shock response and the accessory gene regulator (agr)-controlled transcription of virulence factors. Exposure to serum transiently induced the transcription of distinct virulence factor encoding genes. Transcription of genes reported for mediating the loss of methicillin resistance, especially ccrC, was not significantly different compared to the unchallenged controls. We concluded that, from an evolutionary perspective, bacteria may save energy by incidentally dismissing a fully replicative SCCmec element in contrast to the induction of ccr genes on a population scale. Since the genomic SCCmec integration site is a hot-spot of recombination, occasional losses of elements of 16 kb size may restore capacities for the uptake of foreign genetic material. Subsequent spread of resistance, on the other hand, might depend on the autonomous replication machinery of the deleted SCCmec elements that probably enhance chances for reintegration of SCCmec into susceptible genomes by mere multiplication.

Distribution of CRISPR-Cas systems in the Burkholderiaceae family and its biological implications.

CRISPR-Cas systems are composed of repeated sequences separated by non-repeated sequences that are near genes coding for Cas proteins, which are involved in the function of these systems. Their function has been mostly related to "genetic immunity" against foreign genetic material, among other roles. Interest in them increased after their use in genetic manipulation was uncovered and surveys to find and classify them have been done in several bacterial groups. To determine the presence of these genetic elements in the Burkholderiaceae family members, a bioinformatic approach was followed. Attention in this family comes as it is formed by a great diversity of microorganisms that include opportunistic and true pathogens, and symbiotic and saprophytic organisms, among others. Results show that, in contrast to other bacterial groups, only 8.4% of family members harbor complete CRISPR-Cas systems and the rest either do not have one or have remains or sections of one. Analyses of the spacer sequences indicated that most of them have identity to sections of the same genomes they were found, while a few had identities with either plasmids or phages. The genus with the higher proportion of self-directed spacers is Ralstonia, and their possible roles are discussed. Most of the systems (60%) belong to the class I subtype I-E and a few to subtypes I-C (13.3%), I-F (18.3%), II-C (5%), IV-A (1.7%) and V-C (1.7%). To the best of our knowledge, this is the first study to uncover the CRISPR-Cas system for the whole Burkholderiaceae family.

Prophage Diversity Across Salmonella and Verotoxin-Producing Escherichia coli in Agricultural Niches of British Columbia, Canada.

Prophages have long been regarded as an important contributor to the evolution of Salmonella and Verotoxin-producing E. coli (VTEC), members of the Enterobacteriaceae that cause millions of cases of foodborne illness in North America. In S. Typhimurium, prophages provide many of the genes required for invasion; similarly, in VTEC, the Verotoxin-encoding genes are located in cryptic prophages. The ability of prophages to quickly acquire and lose genes have driven their rapid evolution, leading to highly diversified populations of phages that can infect distantly-related bacterial hosts. To defend against foreign genetic materials (i.e., phages), bacteria have evolved Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) immunity, consisting of variable spacer regions that match short nucleic acid sequences of invaders previously encountered. The number of spacer regions varies widely amongst Enterobacteriaceae, and there is currently no clear consensus if the accumulation of spacers is linked to genomic prophage abundance. Given the immense prophage diversity and contribution to bacterial host phenotypes, we analyzed the prophage sequences within 118 strains of Salmonella and VTEC, 117 of which are of agricultural origin. Overall, 130 unique prophage sequences were identified and they were found to be remarkably diverse with <50% nucleotide similarity, particularly with the Gifsy-1 group which was identified in several Salmonella serovars and interestingly, a strain of VTEC. Additionally, we identified a novel plasmid-like phage that carried antibiotic resistance and bacteriocin resistance genes. The strains analyzed carried at least six distinct spacers which did not possess homology to prophages identified in the same genome. In fact, only a fraction of all identified spacers (14%) possessed significant homology to known prophages. Regression models did not discern a correlation between spacer and prophage abundance in our strains, although the relatively high number of spacers in our strains (an average of 27 in Salmonella and 19 in VTEC) suggest that high rates of infection may occur in agricultural niches and be a contributing driver in bacterial evolution. Cumulatively, these results shed insight into prophage diversity of Salmonella and VTEC, which will have further implications when informing development of phage therapies against these foodborne pathogens.

Random Mutagenesis as a Promising Tool for Microalgal Strain Improvement towards Industrial Production.

Microalgae have become a promising novel and sustainable feedstock for meeting the rising demand for food and feed. However, microalgae-based products are currently hindered by high production costs. One major reason for this is that commonly cultivated wildtype strains do not possess the robustness and productivity required for successful industrial production. Several strain improvement technologies have been developed towards creating more stress tolerant and productive strains. While classical methods of forward genetics have been extensively used to determine gene function of randomly generated mutants, reverse genetics has been explored to generate specific mutations and target phenotypes. Site-directed mutagenesis can be accomplished by employing different gene editing tools, which enable the generation of tailor-made genotypes. Nevertheless, strategies promoting the selection of randomly generated mutants avoid the introduction of foreign genetic material. In this paper, we review different microalgal strain improvement approaches and their applications, with a primary focus on random mutagenesis. Current challenges hampering strain improvement, selection, and commercialization will be discussed. The combination of these approaches with high-throughput technologies, such as fluorescence-activated cell sorting, as tools to select the most promising mutants, will also be discussed.

Emerging Approaches for Enabling RNAi Therapeutics.

RNA interference (RNAi) is a primitive evolutionary mechanism developed to escape incorporation of foreign genetic material. siRNA has been instrumental in achieving the therapeutic potential of RNAi by theoretically silencing any gene of interest in a reversible and sequence-specific manner. Extrinsically administered siRNA generally needs a delivery vehicle to span across different physiological barriers and load into the RISC complex in the cytoplasm in its functional form to show its efficacy. This review discusses the designing principles and examples of different classes of delivery vehicles that have proved to be efficient in RNAi therapeutics. We also briefly discuss the role of RNAi therapeutics in genetic and rare diseases, epigenetic modifications, immunomodulation and combination modality to inch closer in creating a personalized therapy for metastatic cancer. At the end, we present, strategies and look into the opportunities to develop efficient delivery vehicles for RNAi which can be translated into clinics.

Revealing the CRISPR array in bacteria living in our organism

CRISPR (clustered regularly interspaced short palindromic repeats) is an immune system used by bacteria to defend themselves from different types of pathogens. It was discovered that this immune system can modify itself in specific regions called spacers due to previous interaction with foreign genetic material from phages and plasmids. Through our research, we have identified in different bacterial isolates CRISPR arrays belonging to the subtypes I-E (present in 42 samples) and I-F (present in 9 samples). The number of spacers in CRISPR arrays was also estimated based on the array length as a possible connection with the systems activity. Our results yielded arrays as small as 200 bp and as large as 1400 bp.

A target expression threshold dictates invader defense and prevents autoimmunity by CRISPR-Cas13

CRISPR-Cas systems must enact robust immunity against foreign genetic material without inducing cytotoxic autoimmunity. For type VI systems that use Cas13 nucleases and recognize RNA targets, immune activation requires extensive CRISPR RNA (crRNA) guide-target complementarity and a target-flanking motif. Here, we report a third requirement shaping the immune response: the expression of the target transcript exceeding a threshold. We found that endogenous non-essential transcripts targeted by crRNAs rarely elicited autoimmunity. Instead, autoimmune induction required over-expressing the targeted transcripts above a threshold. A genome-wide screen confirmed target expression levels as a global determinant of cytotoxic autoimmunity and revealed that this threshold shifts with each guide-target pair. This threshold further ensured defense against a lytic bacteriophage yetallowed the tolerance of a targeted beneficial gene expressed from an invading plasmid. These findings establish target expression levels as an additional criterion for immune defense by RNA-targeting CRISPR-Cas systems, preventing autoimmunity and distinguishing pathogenic and benign invaders.

Производство молока и численность молочного поголовья КРС в регионах ЮФО

Aim. Assess the features of milk production and dairy cattle breeding in the southern regions of Russia in the face of new global external challenges. Materials and Methods. The main scientific method of research is the horizontal and vertical analysis of open statistical reporting posted on the open website of the Federal State Statistics Service, analytical materials of the Dairy Union of Russia. Mathematical extrapolation tools of the Excel program were used for calculations and making forecasts. To formulate the conclusions, the methods of logical analysis, comparison and generalization of the data presented in the calculation materials and press releases were applied. Results. An assessment was made of the dynamics of raw milk production by all categories of dairy farms in Russia over the past 10 years. It has been established that milk production volumes have increased due to intensive milking and animal husbandry technologies, which made it possible to achieve self-sufficiency in milk up to 80%. Modernization of the dairy complex with the most advanced robotic equipment has allowed farms not only to increase milk yield per cow, but also to significantly improve the quality of milking and the quality of the resulting product. Examples of best practice in terms of milk productivity in the conditions of the South of Russia were demonstrated by enterprises of the Krasnodar Territory and the Volgograd Region. An alarming factor is the reduction in the dairy population of animals. Nevertheless, milk production is intensive against the backdrop of a reduction in the number of dairy cattle, but this factor must be taken into account in the future and its impact should be neutralized through a positive trend in the growth of the number of livestock. Conclusion. On the basis of the study, conclusions were drawn about the need for further expansion of state support measures for the industry and the search for options for introducing Russian-selected animals into production in order to reduce import dependence on foreign genetic material and technologies. The emerging positive trend in the production of raw milk must be supported in the future not only by modernizing production, but also by strengthening the production and resource base.

Convergence of Biofilm Formation and Antibiotic Resistance in Acinetobacter baumannii Infection

Acinetobacter baumannii (A. baumannii) is a leading cause of nosocomial infections as this pathogen has certain attributes that facilitate the subversion of natural defenses of the human body. A. baumannii acquires antibiotic resistance determinants easily and can thrive on both biotic and abiotic surfaces. Different resistance mechanisms or determinants, both transmissible and non-transmissible, have aided in this victory over antibiotics. In addition, the propensity to form biofilms (communities of organism attached to a surface) allows the organism to persist in hospitals on various medical surfaces (cardiac valves, artificial joints, catheters, endotracheal tubes, and ventilators) and also evade antibiotics simply by shielding the bacteria and increasing its ability to acquire foreign genetic material through lateral gene transfer. The biofilm formation rate in A. baumannii is higher than in other species. Recent research has shown how A. baumannii biofilm-forming capacity exerts its effect on resistance phenotypes, development of resistome, and dissemination of resistance genes within biofilms by conjugation or transformation, thereby making biofilm a hotspot for genetic exchange. Various genes control the formation of A. baumannii biofilms and a beneficial relationship between biofilm formation and “antimicrobial resistance” (AMR) exists in the organism. This review discusses these various attributes of the organism that act independently or synergistically to cause hospital infections. Evolution of AMR in A. baumannii, resistance mechanisms including both transmissible (hydrolyzing enzymes) and non-transmissible (efflux pumps and chromosomal mutations) are presented. Intrinsic factors [biofilm-associated protein, outer membrane protein A, chaperon-usher pilus, iron uptake mechanism, poly-β-(1, 6)-N-acetyl glucosamine, BfmS/BfmR two-component system, PER-1, quorum sensing] involved in biofilm production, extrinsic factors (surface property, growth temperature, growth medium) associated with the process, the impact of biofilms on high antimicrobial tolerance and regulation of the process, gene transfer within the biofilm, are elaborated. The infections associated with colonization of A. baumannii on medical devices are discussed. Each important device-related infection is dealt with and both adult and pediatric studies are separately mentioned. Furthermore, the strategies of preventing A. baumannii biofilms with antibiotic combinations, quorum sensing quenchers, natural products, efflux pump inhibitors, antimicrobial peptides, nanoparticles, and phage therapy are enumerated.

Hemp Genome Editing-Challenges and Opportunities.

Hemp (Cannabis sativa L.) is a multipurpose crop with many important uses including medicine, fibre, food and biocomposites. This plant is currently gaining prominence and acceptance for its valuable applications. Hemp is grown as a cash crop for its novel cannabinoids which are estimated to be a multibillion-dollar downstream market. Hemp cultivation can play a major role in carbon sequestration with good CO2 to biomass conversion in low input systems and can also improve soil health and promote phytoremediation. The recent advent of genome editing tools to produce non-transgenic genome-edited crops with no trace of foreign genetic material has the potential to overcome regulatory hurdles faced by genetically modified crops. The use of Artificial Intelligence - mediated trait discovery platforms are revolutionizing the agricultural industry to produce desirable crops with unprecedented accuracy and speed. However, genome editing tools to improve the beneficial properties of hemp have not yet been deployed. Recent availability of high-quality Cannabis genome sequences from several strains (cannabidiol and tetrahydrocannabinol balanced and CBD/THC rich strains) have paved the way for improving the production of valuable bioactive molecules for the welfare of humankind and the environment. In this context, the article focuses on exploiting advanced genome editing tools to produce non-transgenic hemp to improve the most industrially desirable traits. The challenges, opportunities and interdisciplinary approaches that can be adopted from existing technologies in other plant species are highlighted.

CORRELATIONS AND VARIATION STRUCTURE OF GRAIN QUALITY TRAITS AND YIELD OF WHEAT LINES WITH FOREIGN GENETIC MATERIAL

We studied variation structure and correlations between the traits of grain quality and yield in the sample consisting of T. aestivum cultivars, related common wheat species and introgression lines obtained on their basis. In the grain of wheat relatives, two-year average grain macronutrients, micronutrients and protein content were higher than in T. aestivum (1.02 to 2.13 times, depending on the element; 1.4 — for protein). The genetic diversity of mineral composition traits was wider for the lines containing foreign genetic material than for their parental forms, and as a rule, the introgression lines exceeded the original common wheat varieties in the microelements accumulation. The greatest possible number of statistically significant correlations (at least 7) was found for N, P, Zn and Fe content. Protein content was associated with grain's Mg, P, Fe, Zn, Cu. Thousand-kernel weight (TKW) is was negatively correlated with some traits of mineral composition and seed protein content. But the only significant (p = 0.05) association was&#x0D; between TKW and seed nitrogen. We identified lines with optimal mineral composition and high grain productivity, which are of interest for wheat breeding programs.

Multi-species host range of staphylococcal phages isolated from wastewater

The host range of bacteriophages defines their impact on bacterial communities and genome diversity. Here, we characterize 94 novel staphylococcal phages from wastewater and establish their host range on a diversified panel of 117 staphylococci from 29 species. Using this high-resolution phage-bacteria interaction matrix, we unveil a multi-species host range as a dominant trait of the isolated staphylococcal phages. Phage genome sequencing shows this pattern to prevail irrespective of taxonomy. Network analysis between phage-infected bacteria reveals that hosts from multiple species, ecosystems, and drug-resistance phenotypes share numerous phages. Lastly, we show that phages throughout this network can package foreign genetic material enclosing an antibiotic resistance marker at various frequencies. Our findings indicate a weak host specialism of the tested phages, and therefore their potential to promote horizontal gene transfer in this environment.

CRISPR-Cas is associated with fewer antibiotic resistance genes in bacterial pathogens.

The acquisition of antibiotic resistance (ABR) genes via horizontal gene transfer (HGT) is a key driver of the rise in multidrug resistance amongst bacterial pathogens. Bacterial defence systems per definition restrict the influx of foreign genetic material, and may therefore limit the acquisition of ABR. CRISPR-Cas adaptive immune systems are one of the most prevalent defences in bacteria, found in roughly half of bacterial genomes, but it has remained unclear if and how much they contribute to restricting the spread of ABR. We analysed approximately 40 000 whole genomes comprising the full RefSeq dataset for 11 species of clinically important genera of human pathogens, including Enterococcus, Staphylococcus, Acinetobacter and Pseudomonas. We modelled the association between CRISPR-Cas and indicators of HGT, and found that pathogens with a CRISPR-Cas system were less likely to carry ABR genes than those lacking this defence system. Analysis of the mobile genetic elements (MGEs) targeted by CRISPR-Cas supports a model where this host defence system blocks important vectors of ABR. These results suggest a potential 'immunocompromised' state for multidrug-resistant strains that may be exploited in tailored interventions that rely on MGEs, such as phages or phagemids, to treat infections caused by bacterial pathogens. This article is part of the theme issue 'The secret lives of microbial mobile genetic elements'.

Dysregulation of miRNAs in DLBCL: Causative Factor for Pathogenesis, Diagnosis and Prognosis.

MicroRNA is a small non-coding RNA (sncRNA) involved in gene silencing and regulating post-transcriptional gene expression. miRNAs play an essential role in the pathogenesis of numerous diseases, including diabetes, cardiovascular diseases, viral diseases and cancer. Diffuse large B-cell lymphoma (DLBCL) is an aggressive non-Hodgkin’s lymphoma (NHL), arising from different stages of B-cell differentiation whose pathogenesis involves miRNAs. Various viral and non-viral vectors are used as a delivery vehicle for introducing specific miRNA inside the cell. Adenoviruses are linear, double-stranded DNA viruses with 35 kb genome size and are extensively used in gene therapy. Meanwhile, Adeno-associated viruses accommodate up to 4.8 kb foreign genetic material and are favorable for transferring miRNA due to small size of miRNA. The genetic material is integrated into the DNA of the host cell by retroviruses so that only dividing cells are infected and stable expression of miRNA is achieved. Over the years, remarkable progress was made to understand DLBCL biology using advanced genomics and epigenomics technologies enabling oncologists to uncover multiple genetic mutations in DLBCL patients. These genetic mutations are involved in epigenetic modification, ability to escape immunosurveillance, impaired BCL6 and NF-κβ signaling pathways and blocking terminal differentiation. These pathways have since been identified and used as therapeutic targets for the treatment of DLBCL. Recently miRNAs were also identified to act either as oncogenes or tumor suppressors in DLBCL pathology by altering the expression levels of some of the known DLBCL related oncogenes. i.e., miR-155, miR-17-92 and miR-21 act as oncogenes by altering the expression levels of MYC, SHIP and FOXO1, respectively, conversely; miR-34a, mir-144 and miR-181a act as tumor suppressors by altering the expression levels of SIRT1, BCL6 and CARD11, respectively. Hundreds of miRNAs have already been identified as biomarkers in the prognosis and diagnosis of DLBCL because of their significant roles in DLBCL pathogenesis. In conclusion, miRNAs in addition to their role as biomarkers of prognosis and diagnosis could also serve as potential therapeutic targets for treating DLBCL.

Consumer acceptance of gene-edited food products in China

Biotechnology use in food production has been a polarizing topic that has encountered resistance from some consumers. The discovery of genome editing biotechnology, in which no foreign genetic material is introduced into the host organism while making accurate and efficient changes in genomes, has the potential to revolutionize food biotechnology in a more socially acceptable and less polarizing fashion. The success and adoption of gene-edited foods, however, ultimately depends on consumer acceptance. This study reports the results of a geographically disperse Chinese consumer acceptance study (n = 835) in which individuals evaluated rice and pork products that were bio-engineered to address two significant hazards that have recently garnered international attention: cadmium contamination in rice and African swine fever. We explore the role that food technology neophobia has on consumer acceptance and assess how information on the differences between transgenic and gene editing technologies affects consumer preferences. While averse to the use of biotechnology in food products, consumers were considerably more accepting of products that have undergone genome editing rather than transgenic modification. We find differential impacts of information provision on preferences between pork and rice products and on preferences for product provenance. Our analysis indicates that a reduction in consumers’ fear of novel food technologies can substantially increase consumer valuation and market acceptance of bioengineered food products and reinforces the need to consider attitudes in measuring acceptance of novel food products.

The tracrRNA in CRISPR Biology and Technologies.

CRISPR-Cas adaptive immune systems in bacteria and archaea utilize short CRISPR RNAs (crRNAs) to guide sequence-specific recognition and clearance of foreign genetic material. Multiple crRNAs are stored together in a compact format called a CRISPR array that is transcribed and processed into the individual crRNAs. While the exact processing mechanisms vary widely, some CRISPR-Cas systems, including those encoding the Cas9 nuclease, rely on a trans-activating crRNA (tracrRNA). The tracrRNA was discovered in 2011 and was quickly co-opted to create single-guide RNAs as core components of CRISPR-Cas9 technologies. Since then, further studies have uncovered processes extending beyond the traditional role of tracrRNA in crRNA biogenesis, revealed Cas nucleases besides Cas9 that are dependent on tracrRNAs, and established new applications based on tracrRNA engineering. In this review, we describe the biology of the tracrRNA and how its ongoing characterization has garnered new insights into prokaryotic immune defense and enabled key technological advances.

Pathogenicity and Molecular Characterization of Staphylococcus aureus Strains Isolated from the Hospital Environment of CHU-Z Abomey-Calavi/Sô-Ava (Benin).

Staphylococcus aureus is a major human pathogen present on a third of the healthy population. The bacterium possesses an extensive arsenal of virulence factors. The pathogenicity is linked with S. aureus high plasticity and its exceptional ability to incorporate foreign genetic material. The aim of the present study was to perform molecular characterization of Staphylococcus aureus strains isolated from the clinical environment of the CHU-Z Abomey-Calavi/Sô-Ava. Isolation of Staphylococcus aureus bacterium was performed on Chapman agar. Toxin production by isolated S. aureus strains was investigated using the radial immunoprecipitation technique. A colorimetric assay was used to evaluate Staphylococcus aureus lipase (SA-Lipase) production. Finally, the expression of antibiotic resistance genes and genes encoding toxins production was investigated. Our data suggest that none of the isolated Staphylococcus aureus strains expressed the investigated toxin genes. Interestingly, SA-Lipase was produced by 14.28% of our isolated S. aureus strains. The mecA gene was present in 57.14% of the isolated strains, while PVL and TSST-1 genes were identified in 2.85 and 7.14% of S. aureus, respectively. Significant genetic diversity was observed along the hospital environment S. aureus strains. The present study reveals the level of virulence of S. aureus strains isolated in the different units of CHU-Z Abomey Calavi/Sô-Ava through the production of lipase, PVL, and epidermolysins. The molecular study has favored a genetic characterization within the isolated strains.

Epidemiology of Pestivirus H in Brazil and Its Control Implications.

Along with viruses in the Pestivirus A (Bovine Viral Diarrhea Virus 1, BVDV1) and B species (Bovine Viral Diarrhea Virus 2, BVDV2), members of the Pestivirus H are mainly cattle pathogens. Viruses belonging to the Pestivirus H group are known as HoBi-like pestiviruses (HoBiPev). Genetic and antigenic characterization suggest that HoBiPev are the most divergent pestiviruses identified in cattle to date. The phylogenetic analysis of HoBiPev results in at least five subgroups (a–e). Under natural or experimental conditions, calves infected with HoBiPev strains typically display mild upper respiratory signs, including nasal discharge and cough. Although BVDV1 and BVDV2 are widely distributed and reported in many South American countries, reports of HoBiPev in South America are mostly restricted to Brazil. Despite the endemicity and high prevalence of HoBiPev in Brazil, only HoBiPev-a was identified to date in Brazil. Unquestionably, HoBiPev strains in BVDV vaccine formulations are required to help curb HoBiPev spread in endemic regions. The current situation in Brazil, where at this point only HoBiPev-a seems present, provides a more significant opportunity to control these viruses with the use of a vaccine with a single HoBiPev subtype. Despite the lack of differentiation among bovine pestiviruses by current BVDV tests, the reduced genetic variability of HoBiPev in Brazil may allow reliable identification of cases within the region. On the other hand, introducing foreign ruminants, biologicals, and genetic material to South America, especially if it originated from other HoBiPev-endemic countries, should consider the risk of introducing divergent HoBiPev subtypes.

Intracellular DNA transfer events restricted to the genus Convallaria within the Asparagaceae family: Possible mechanisms and potential as genetic markers for biographical studies

Intracellular gene transfer among plant genomes is a common phenomenon. Due to their high conservation and high plastid membrane integrity, chloroplast (cp) genomes incorporate foreign genetic material very rarely. Convallaria is a small monocotyledonous genus consisting of C. keiskei, C. majalis and C. montana. Here, we characterized, analyzed and identified 3.3 and 3.7 kb of mitochondrial DNA sequences in the plastome (MCP) of C. majalis and C. montana, respectively. We identified 6 bp and 23 bp direct repeats and mitochondrial pseudogenes, with rps3, rps19 and rpl10 identified in the MCP region. Additionally, we developed novel plastid molecular genetic markers to differentiate Convallaria spp. based on 21 populations. BEAST and biogeographical analyses suggested that Convallaria separated into Eurasian and North American lineages during the middle Pliocene and originated in East Asia. Vicariance in the genus was followed by dispersal into Europe and southeastern North America. These analyses indicate that the MCP event was restricted to the genus Convallaria of Asparagaceae, in contrast to similar events that occurred in its common ancestors with other families of land plants. However, further mitochondrial and population studies are necessary to understand the integration of the MCP region and gene flow in the genus Convallaria.

RNA Viral Vectors for Accelerating Plant Synthetic Biology.

The tools of synthetic biology have enormous potential to help us uncover the fundamental mechanisms controlling development and metabolism in plants. However, their effective utilization typically requires transgenesis, which is plagued by long timescales and high costs. In this review we explore how transgenesis can be minimized by delivering foreign genetic material to plants with systemically mobile and persistent vectors based on RNA viruses. We examine the progress that has been made thus far and highlight the hurdles that need to be overcome and some potential strategies to do so. We conclude with a discussion of biocontainment mechanisms to ensure these vectors can be used safely as well as how these vectors might expand the accessibility of plant synthetic biology techniques. RNA vectors stand poised to revolutionize plant synthetic biology by making genetic manipulation of plants cheaper and easier to deploy, as well as by accelerating experimental timescales from years to weeks.