Vehicles, Replicators, and Intercellular Movement of Genetic Information: Evolutionary Dissection of a Bacterial Cell

Prokaryotic biosphere is vastly diverse in many respects. Any given bacterial cell may harbor in different combinations viruses, plasmids, transposons, and other genetic elements along with their chromosome(s). These agents interact in complex environments in various ways causing multitude of phenotypic effects on their hosting cells. In this discussion I perform a dissection for a bacterial cell in order to simplify the diversity into components that may help approach the ocean of details in evolving microbial worlds. The cell itself is separated from all the genetic replicators that use the cell vehicle for preservation and propagation. I introduce a classification that groups different replicators according to their horizontal movement potential between cells and according to their effects on the fitness of their present host cells. The classification is used to discuss and improve the means by which we approach general evolutionary tendencies in microbial communities. Moreover, the classification is utilized as a tool to help formulating evolutionary hypotheses and to discuss emerging bacterial pathogens as well as to promote understanding on the average phenotypes of different replicators in general. It is also discussed that any given biosphere comprising prokaryotic cell vehicles and genetic replicators may naturally evolve to have horizontally moving replicators of various types

Population-level consequences of risky dispersal

Achieving sufficient connectivity between populations is essential for persistence, but costs of dispersal may select against individual traits or behaviours that, if present, would improve connectivity. Existing dispersal models tend to ignore the multitude of risks to individuals: while many assess the effect of mortality costs, there is also a risk of failing to find new habitat, especially when the entire inhabitable area remains both small and fragmented. There are few known rules governing whether individuals evolve to disperse more, or less, than what is ideal for population connectivity and persistence. Here we aim to fill this gap, while also noting that evolution might not only produce suboptimal dispersal behaviour: it also influences individual heterogeneity in dispersal. Intuitively, we might expect heterogeneity to improve connectivity, as some individuals will travel far. However, we show that this is only true if dispersal distances on average are quite short; heterogeneity can also lead to reduced connectivity because it can reduce the proportion of the most profitable (‘safest’) intermediate dispersal distances. In general, our results show that conditions typically associated with conservation concerns (small and fragmented habitats inhabited by a species with a low birth rate) are also ones that are most likely to lead to suboptimal dispersal traits. This prompts the question of assisted dispersal in cases of urgent conservation concern

Population-level consequences of risky dispersal

Achieving sufficient connectivity between populations is essential for persistence, but costs of dispersal may select against individual traits or behaviours that, if present, would improve connectivity. Existing dispersal models tend to ignore the multitude of risks to individuals: while many assess the effect of mortality costs, there is also a risk of failing to find new habitat, especially when the entire inhabitable area remains both small and fragmented. There are few known rules governing whether individuals evolve to disperse more, or less, than what is ideal for population connectivity and persistence. Here we aim to fill this gap, while also noting that evolution might not only produce suboptimal dispersal behaviour: it also influences individual heterogeneity in dispersal. Intuitively, we might expect heterogeneity to improve connectivity, as some individuals will travel far. However, we show that this is only true if dispersal distances on average are quite short; heterogeneity can also lead to reduced connectivity because it can reduce the proportion of the most profitable (‘safest’) intermediate dispersal distances. In general, our results show that conditions typically associated with conservation concerns (small and fragmented habitats inhabited by a species with a low birth rate) are also ones that are most likely to lead to suboptimal dispersal traits. This prompts the question of assisted dispersal in cases of urgent conservation concern

Caracterización de los fagos de Klebsiella pneumoniae con potencial biotecnológico

The extensive use and misuse of antibiotics has led to an increased emergence of multidrug-resistant Klebsiella pneumoniae strains. They are a serious concern worldwide due to their propensity to spread and the scarce effective treatments left. Consequently, phage therapy is garnering renewed interest as an alternative method to defeat antibiotic resistant bacteria. Phages – natural pathogens of bacteria – have several properties: high capacity to replicate and host specificity that turns them into a great advantage over antibiotics. Eight bacteriophages infecting Klebsiella pneumoniae were characterized according to their genetic material and morphology by performing endonuclease digestions and transmission electron microscopy imaging with 1% phosphotungstic acid or 2% uranyl acetate as staining dyes. Then, they were classified in agreement with their morphological characterization. Seven phages (EKP3P1, EKP3P2, EKP3P4, EKP3P5, EKP8P2, EKP8P3 and EKP8P4) were classified into Siphoviridae family showing hexagonal heads with long non- contractile, sometimes flexible tails and closely related restriction patterns. EKP8P1 phage was classified into Podoviridae family showing an icosahedral head with a short non-contractile tail and a different restriction pattern. They all belong to Caudovirales order. Moreover, a prophage was found in EKP8P1 sample, and classified into Siphoviridae family according to its morphology. The genome of EKP3P5 phage, a double stranded DNA of 47,622 bp long, was sequenced and annotated manually. EKP3P5 phage is a temperate phage encoding integrase, holin and endolysin proteins, among others. Therefore, EKP3P5 could not be used in phage therapy due to the risk of transferring virulence and resistance genes to the host bacteria. For all the above reasons, this thesis provides detailed knowledge of the physical structure along with genomic qualities of eight bacteriophages infecting multidrug- resistant Klebsiella pneumoniae strains. This is important for determining the potential of phages as therapeutic agents and the first step to improve phage therapy

Midbiotics : conjugative plasmids for genetic engineering of natural gut flora

ABSTRACTThe possibility to modify gut bacterial flora has become an important goal, and various approaches are used to achieve desirable communities. However, the genetic engineering of existing microbes in the gut, which are already compatible with the rest of the community and host immune system, has not received much attention. Here, we discuss and experimentally evaluate the possibility to use modified and mobilizable CRISPR-Cas9-endocing plasmid as a tool to induce changes in bacterial communities. This plasmid system (briefly midbiotic) is delivered from bacterial vector into target bacteria via conjugation. Compared to, for example, bacteriophage-based applications, the benefits of conjugative plasmids include their independence of any particular receptor(s) on host bacteria and their relative immunity to bacterial defense mechanisms (such as restriction-modification systems) due to the synthesis of the complementary strand with host-specific epigenetic modifications. We show that conjugative plasmid in association with a mobilizable antibiotic resistance gene targeting CRISPR-plasmid efficiently causes ESBL-positive transconjugants to lose their resistance, and multiple gene types can be targeted simultaneously by introducing several CRISPR RNA encoding segments into the transferred plasmids. In the rare cases where the midbiotic plasmids failed to resensitize bacteria to antibiotics, the CRISPR spacer(s) and their adjacent repeats or larger regions were found to be lost. Results also revealed potential caveats in the design of conjugative engineering systems as well as workarounds to minimize these risks.Peer reviewe

Indirect Selection against Antibiotic Resistance via Specialized Plasmid-Dependent Bacteriophages

Antibiotic resistance genes of important Gram-negative bacterial pathogens are residing in mobile genetic elements such as conjugative plasmids. These elements rapidly disperse between cells when antibiotics are present and hence our continuous use of antimicrobials selects for elements that often harbor multiple resistance genes. Plasmid-dependent (or male-specific or, in some cases, pilus-dependent) bacteriophages are bacterial viruses that infect specifically bacteria that carry certain plasmids. The introduction of these specialized phages into a plasmid-abundant bacterial community has many beneficial effects from an anthropocentric viewpoint: the majority of the plasmids are lost while the remaining plasmids acquire mutations that make them untransferable between pathogens. Recently, bacteriophage-based therapies have become a more acceptable choice to treat multi-resistant bacterial infections. Accordingly, there is a possibility to utilize these specialized phages, which are not dependent on any particular pathogenic species or strain but rather on the resistance-providing elements, in order to improve or enlengthen the lifespan of conventional antibiotic approaches. Here, we take a snapshot of the current knowledge of plasmid-dependent bacteriophages

Systematic Comparison of Epidemic and Non-Epidemic Carbapenem Resistant Klebsiella pneumoniae Strains

Over the past few decades, extensively drug resistant (XDR) resistant Klebsiella pneumoniae has become a notable burden to healthcare all over the world. Especially carbapenemase-producing strains are problematic due to their capability to withstand even last resort antibiotics. Some sequence types (STs) of K. pneumoniae are significantly more prevalent in hospital settings in comparison to other equally resistant strains. This provokes the question whether or not there are phenotypic characteristics that may render certain K. pneumoniae more suitable for epidemic dispersal between patients, hospitals, and different environments. In this study, we selected seven epidemic and non-epidemic carbapenem resistant K. pneumoniae isolates for extensive systematic characterization for phenotypic and genotypic qualities in order to identify potential factors that precede or emerge from epidemic successfulness. Studied characteristics include growth rates and densities in different conditions (media, temperature, pH, resource levels), tolerance to alcohol and drought, inhibition between strains, ability to compensate pH, as well as various genomic features. Overall, there are clear differences between isolates, yet, only drought tolerance was found to notably associate with non-epidemic K. pneumoniae strains. We further report a preliminary study on the potential to control K. pneumoniae ST11 with an antimicrobial component produced by a non-epidemic K. pneumoniae. This component initially restricts bacterial growth, but stable resistance develops rapidly in vitro

New insights and trends in undergraduate research: evidence from Jordan

Quality-filtered VCF file for treatment-replicate 2BV-II

Extracellular vesicles provide a capsid-free vector for oncolytic adenoviral DNA delivery

Extracellular vesicles (EVs) have been showcased as auspicious candidates for delivering therapeutic cargo, including oncolytic viruses for cancer treatment. Delivery of oncolytic viruses in EVs could provide considerable advantages, hiding the viruses from the immune system and providing alternative entry pathways into cancer cells. Here we describe the formation and viral cargo of EVs secreted by cancer cells infected with an oncolytic adenovirus (IEVs, infected cell-derived EVs) as a function of time after infection. IEVs were secreted already before the lytic release of virions and their structure resembled normally secreted EVs, suggesting that they were not just apoptotic fragments of infected cells. IEVs were able to carry the viral genome and induce infection in other cancer cells. As such, the role of EVs in the life cycle of adenoviruses may be an important part of a successful infection and may also be harnessed for cancer- and gene therapy.Peer reviewe

Dual-stressor selection alters eco-evolutionary dynamics in experimental communities

Recognizing when and how rapid evolution drives ecological change is fundamental for our understanding of almost all ecological and evolutionary processes such as community assembly, genetic diversification and the stability of communities and ecosystems. Generally, rapid evolutionary change is driven through selection on genetic variation and is affected by evolutionary constraints, such as tradeoffs and pleiotropic effects, all contributing to the overall rate of evolutionary change. Each of these processes can be influenced by the presence of multiple environmental stressors reducing a population's reproductive output. Potential consequences of multistressor selection for the occurrence and strength of the link from rapid evolution to ecological change are unclear. However, understanding these is necessary for predicting when rapid evolution might drive ecological change. Here we investigate how the presence of two stressors affects this link using experimental evolution with the bacterium Pseudomonas fluorescens and its predator Tetrahymena thermophila. We show that the combination of predation and sublethal antibiotic concentrations delays the evolution of anti-predator defence and antibiotic resistance compared with the presence of only one of the two stressors. Rapid defence evolution drives stabilization of the predator-prey dynamics but this link between evolution and ecology is weaker in the two-stressor environment, where defence evolution is slower, leading to less stable population dynamics. Tracking the molecular evolution of whole populations over time shows further that mutations in different genes are favoured under multistressor selection. Overall, we show that selection by multiple stressors can significantly alter eco-evolutionary dynamics and their predictability.Peer reviewe