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The opportunistic pathogen Acinetobacter baumannii, carries variants of A. baumannii resistance islands (AbaR)-type genomic islands conferring multidrug resistance. Their pervasiveness in the species has remained enigmatic. The dissemination of AbaRs is intricately linked to their horizontal transfer via natural transformation, a process through which bacteria can import and recombine exogenous DNA, effecting allelic recombination, genetic acquisition, and deletion. In experimental populations of the closely related pathogenic Acinetobacter nosocomialis, we quantified the rates at which these natural transformation events occur between individuals. When integrated into a model of population dynamics, they lead to the swift removal of AbaRs from the population, contrasting with the high prevalence of AbaRs in genomes. Yet, genomic analyses show that nearly all AbaRs specifically disrupt comM, a gene encoding a helicase critical for natural transformation. We found that such disruption impedes gene acquisition, and deletion, while moderately impacting acquisition of single nucleotide polymorphism. A mathematical evolutionary model demonstrates that AbaRs inserted into comM gain a selective advantage over AbaRs inserted in sites that do not inhibit or completely inhibit transformation, in line with the genomic observations. The persistence of AbaRs can be ascribed to their integration into a specific gene, diminishing the likelihood of their removal from the bacterial genome. This integration preserves the acquisition and elimination of alleles, enabling the host bacterium--and thus its AbaR--to adapt to unpredictable environments and persist over the long term. This work underscores how manipulation of natural transformation by mobile genetic elements can drive the prevalence of multidrug resistance. [ABSTRACT FROM AUTHOR]

The development of the human central nervous system initiates in the early embryonic period until long after delivery. It has been shown that several neurological and neuropsychiatric diseases originate from prenatal incidents. Mathematical models offer a direct way to understand neurodevelopmental processes better. Mathematical modelling of neurodevelopment during the embryonic period is challenging in terms of how to 'Approach', how to initiate modelling and how to propose the appropriate equations that fit the underlying dynamics of neurodevelopment during the embryonic period while including the variety of elements that are built-in naturally during the process of neurodevelopment. It is imperative to answer where and how to start modelling; in other words, what is the appropriate 'Approach'? Therefore, one objective of this study was to tackle the mathematical issue broadly from different aspects and approaches. The approaches were divided into three embryonic categories: cell division, neural tube growth and neural plate growth. We concluded that the neural plate growth approach provides a suitable platform for simulation of brain formation/neurodevelopment compared to cell division and neural tube growth. We devised a novel equation and designed algorithms that include geometrical and topological algorithms that could fit most of the necessary elements of the neurodevelopmental process during the embryonic period. Hence, the proposed equations and defined mathematical structure would be a platform to generate an artificial neural network that autonomously grows and develops. [ABSTRACT FROM AUTHOR]

The present paper considers the results of long-term (up to 17 years) in situ and laboratory research carried out on oiled French, Spanish, and Russian seacoasts. The objective of this research is to quantify the influence of geographical factors on the rates of natural transformation of the heavy fuel oil stranded ashore and to develop an empirical statistical model in order to evaluate the self-cleansing capacity of the coastal environment. In a number of field campaigns, 363 samples of weathered oil slicks and tar balls have been collected and analysed with the use of thin-layer chromatography combined with optical and gravimetric methods. The results obtained have been subjected to multiple nonlinear regression analyses. It has been shown that heavy fuel oil natural attenuation is more active in continental or estuarine environments influenced by nutrient-rich freshwater runoff and characterised by a higher number of sunny days, solar irradiation, and large temperature fluctuations. On the oceanic coasts, especially in sectors with low hydrodynamic energy, these processes take more time. The resulting model allows for the identification and mapping of the most vulnerable seacoasts, characterised by a low potential to degrade oil pollution. This information may be used in the contingency plans in order to optimise clean-up techniques and associated costs. [ABSTRACT FROM AUTHOR]

ABSTRACT Streptococcus dysgalactiae is an emerging human and animal pathogen. Functional studies of genes involved in virulence of S. dysgalactiae and other pyogenic group streptococci are often hampered by limited genetic tractability. It is known that pyogenic streptococci carry genes required for competence for natural transformation; however, in contrast to other streptococcal subgroups, there is limited evidence for gene transfer by natural transformation in these bacteria. In this study, we systematically assessed the genomes of 179 S. dysgalactiae strains of both human and animal origins (subsp. equisimilis and dysgalactiae, respectively) for the presence of genes required for natural transformation. While a considerable fraction of the strains contained inactive genes, the majority (64.2%) of the strains had an intact gene set. In selected strains, we examined the dynamics of competence activation after addition of competence-inducing pheromones using transcriptional reporter assays and exploratory RNA-seq. Based on these findings, we were able to establish a protocol allowing us to utilize natural transformation to construct deletion mutants by allelic exchange in several S. dysgalactiae strains of both subspecies. As part of the work, we deleted putative lactose utilization genes to study their role in growth on lactose. The data presented here provide new knowledge on the potential of horizonal gene transfer by natural transformation in S. dysgalactiae and, importantly, demonstrates the possibility to exploit natural transformation for genetic engineering in these bacteria.IMPORTANCENumerous Streptococcus spp. exchange genes horizontally through natural transformation, which also facilitates efficient genetic engineering in these organisms. However, for the pyogenic group of streptococci, including the emerging pathogen Streptococcus dysgalactiae, there is limited experimental evidence for natural transformation. In this study, we demonstrate that natural transformation in vitro indeed is possible in S. dysgalactiae strains under optimal conditions. We utilized this method to perform gene deletion through allelic exchange in several strains, thereby paving the way for more efficient gene engineering methods in pyogenic streptococci.

Methicillin-resistant Staphylococcus aureus (MRSA) is an important human pathogen that has emerged through the horizontal acquisition of the staphylococcal cassette chromosome mec (SCCmec). Previously, we showed that SCCmec from heat-killed donors can be transferred via natural transformation in biofilms at frequencies of 10−8-10−7. Here, we show an improved transformation assay of SCCmec with frequencies up to 10−2 using co-cultured biofilms with living donor cells. The Ccr-attB system played an important role in SCCmec transfer, and the deletion of ccrAB recombinase genes reduced the frequency ∼30-fold. SCCmec could be transferred from either MRSA or methicillin-resistant coagulase-negative staphylococci to some methicillin-sensitive S. aureus recipients. In addition, the transformation of other plasmid or chromosomal genes is enhanced by using living donor cells. This study emphasizes the role of natural transformation as an evolutionary ability of S. aureus and in MRSA emergence.

Background: Horizontal gene transfer (HGT) is considered an important mechanism to contribute to the evolution of bacteria, plants, and animals by allowing the movement of genetic material between organisms, in difference to vertical inheritance. Thereby it can also play a significant role in spreading traits like antibiotic resistance among bacteria and virulence factors between pathogens. During the HGT, organisms take up free DNA from the environment and incorporate it into their genomes. Although HGT is known to be carried out by many organisms, there is limited information on how organisms select which genetic material for horizontal transfer. Here we have investigated the preference priority of Saccharomyces cerevisiae between different options of gene source presented under certain stress conditions to repair a double-strand break (DSB) in DNA via HR. Results: Each genetic module was designed with appropriate sequences being homologous for two sides of the DSB, which is important for yeast to repair the fracture with HR. S. cerevisiae made a random selection between two heterologous T1 (44%) and T2 (56%) modules to repair DSB. Interestingly, yeast corrected the DNA break only with the T3 module (almost 100%) when the homologous T3 module was an option for the selection. It seems that S. cerevisiae tends to prefer T3 over alternatives to fix DSBs when it exists among the options. Conclusions: It seems that S. cerevisiae have a preference for priority to select a particular one under certain conditions when it has various DNA options to repair a DSB in its genome, further studies are required to support our findings. [ABSTRACT FROM AUTHOR]

ABSTRACT Genetic competence for natural transformation, considered one of the three main mechanisms leading to horizontal gene transfer in bacteria, is able to promote evolution, through genomic plasticity, and foster antibiotic resistance and virulence factors spreading. Conserved machinery and actors required to perform natural transformation have been shown to accumulate at different cellular localizations depending on the model organism considered. In this study, we investigate the transformation apparatus composition, localization, and dynamics in the human pathogen Staphylococcus aureus. We particularly show that most of the natural transformation actors co-localize in clusters. We also reveal that the localization of natural transformation proteins is dynamic, following the cell cycle. Ultimately, the natural transformation apparatus is preferentially established in the vicinity of the division septum. All these results demonstrate that DNA binding, uptake, and recombination are spatially and temporally coordinated to ensure S. aureus natural transformation. Finally, we hypothesize that S. aureus competent cells would initiate and then block cell division to ensure the success of natural transformation before the final constriction of the cytokinetic ring. IMPORTANCE Natural transformation, considered one of the three main mechanisms leading to horizontal gene transfer in bacteria, is able to promote genomic plasticity and foster antibiotic resistance spreading. Conserved machinery and actors required to perform natural transformation have been shown to accumulate at different cellular localizations depending on the model organism considered. Here, we show in the human pathogen Staphylococcus aureus that DNA binding, uptake, and recombination are spatially and temporally coordinated to ensure S. aureus natural transformation. We also reveal that localization of natural transformation proteins occurs in the vicinity of the division septum allowing S. aureus competent cells to block cell division to ensure the success of natural transformation before the final constriction of the cytokinetic ring.

Notions such as śūnyata, catuskoti, and Indra's net, which figure prominently in Buddhist philosophy, are difficult to readily accommodate within our ordinary thinking about everyday objects. Famous Buddhist scholar Nāgārjuna considered two levels of reality: one called conventional reality, and the other ultimate reality. Within this framework, śūnyata refers to the claim that at the ultimate level objects are devoid of essence or 'intrinsic properties', but are interdependent by virtue of their relations to other objects. Catuskoti refers to the claim that four truth values, along with contradiction, are admissible in reasoning. Indra's net refers to the claim that every part of a whole is reflective of the whole. Here we present category theoretic constructions that are reminiscent of these Buddhist concepts. The universal mapping property definition of mathematical objects, wherein objects of a universe of discourse are defined not in terms of their content, but in terms of their relations to all objects of the universe is reminiscent of śūnyata. The objective logic of perception, with perception modeled as [a category of] two sequential processes (sensation followed by interpretation), and with its truth value object of four truth values, is reminiscent of the Buddhist logic of catuskoti. The category of categories, wherein every category has a subcategory of sets with zero structure within which every category can be modeled, is reminiscent of Indra's net. Our thorough elaboration of the parallels between Buddhist philosophy and category theory can facilitate better understanding of Buddhist philosophy, and bring out the broader philosophical import of category theory beyond mathematics. [ABSTRACT FROM AUTHOR]

In this paper, we introduce the notions of natural transformation between morphisms of quasigroupoids and between morphisms of weak Hopf quasigroups. Also, we prove that natural transformations between morphisms of finite quasigroupoids come from natural transformations between morphisms of weak Hopf quasigroups and, on the other hand, we obtain that every natural transformation for morphisms defined between pointed cosemisimple weak Hopf quasigroups comes from a natural transformation between finite quasigroupoids. [ABSTRACT FROM AUTHOR]

Natural transformation, or the uptake of naked DNA from the external milieu by bacteria, holds a unique place in the history of biology. This is both the beginning of the realization of the correct chemical nature of genes and the first technical step to the molecular biology revolution that sees us today able to modify genomes almost at will. Yet the mechanistic understanding of bacterial transformation still presents many blind spots and many bacterial systems lag behind power horse model systems like Escherichia coli in terms of ease of genetic modification. Using Neisseria gonorrhoeae as a model system and using transformation with multiple DNA molecules, we tackle in this paper both some aspects of the mechanistic nature of bacterial transformation and the presentation of new molecular biology techniques for this organism. We show that similarly to what has been demonstrated in other naturally competent bacteria, Neisseria gonorrhoeae can incorporate, at the same time, different DNA molecules modifying DNA at different loci within its genome. In particular, co-transformation of a DNA molecule bearing an antibiotic selection cassette and another non-selected DNA piece can lead to the integration of both molecules in the genome while selecting only through the selective cassette at percentages above 70%. We also show that successive selections with two selection markers at the same genetic locus can drastically reduce the number of genetic markers needed to do multisite genetic modifications in Neisseria gonorrhoeae. Despite public health interest heightened with the recent rise in antibiotic resistance, the causative agent of gonorrhea still does not possess a plethora of molecular techniques. This paper will extend the techniques available to the Neisseria community while providing some insights into the mechanisms behind bacterial transformation in Neisseria gonorrhoeae. We are providing a suite of new techniques to quickly obtain modifications of genes and genomes in the Neisserial naturally competent bacteria. [ABSTRACT FROM AUTHOR]

Plasmid-mediated horizontal gene transfer (HGT) is a major driver of genetic diversity in bacteria. We experimentally validated the function of a putative mercury resistance operon present on an abundant 8-kbp native plasmid found in groundwater samples without detectable levels of mercury. Phylogenetic analyses of the plasmid-encoded mercury reductases from the studied groundwater site show them to be distinct from those reported in proximal metal-contaminated sites. We synthesized the entire native plasmid and demonstrated that the plasmid was sufficient to confer functional mercury resistance in Escherichia coli Given the possibility that natural transformation is a prevalent HGT mechanism in the low-cell-density environments of groundwaters, we also assayed bacterial strains from this environment for competence. We used the native plasmid-encoded metal resistance to design a screen and identified 17 strains positive for natural transformation. We selected 2 of the positive strains along with a model bacterium to fully confirm HGT via natural transformation. From an ecological perspective, the role of the native plasmid population in providing advantageous traits combined with the microbiome's capacity to take up environmental DNA enables rapid adaptation to environmental stresses.IMPORTANCE Horizontal transfer of mobile genetic elements via natural transformation has been poorly understood in environmental microbes. Here, we confirm the functionality of a native plasmid-encoded mercury resistance operon in a model microbe and then query for the dissemination of this resistance trait via natural transformation into environmental bacterial isolates. We identified 17 strains including Gram-positive and Gram-negative bacteria to be naturally competent. These strains were able to successfully take up the plasmid DNA and obtain a clear growth advantage in the presence of mercury. Our study provides important insights into gene dissemination via natural transformation enabling rapid adaptation to dynamic stresses in groundwater environments.

Natural transformation, the process whereby a cell acquires DNA directly from the environment, is an important driver of evolution in microbial populations, yet the mechanism of DNA uptake is only characterized in bacteria. To expand our understanding of natural transformation in archaea, we undertook a genetic approach to identify a catalog of genes necessary for transformation in Methanococcus maripaludis. Using an optimized method to generate random transposon mutants, we screened 6144 mutant strains for defects in natural transformation and identified 25 transformation-associated candidate genes. Among these are genes encoding components of the type IV-like pilus, transcription/translation associated genes, genes encoding putative membrane bound transport proteins, and genes of unknown function. Interestingly, similar genes were identified regardless of whether replicating or integrating plasmids were provided as a substrate for transformation. Using allelic replacement mutagenesis, we confirmed that several genes identified in these screens are essential for transformation. Finally, we identified a homolog of a membrane bound substrate transporter in Methanoculleus thermophilus and verified its importance for transformation using allelic replacement mutagenesis, suggesting a conserved mechanism for DNA transfer in multiple archaea. These data represent an initial characterization of the genes important for transformation which will inform efforts to understand gene flow in natural populations. Additionally, knowledge of the genes necessary for natural transformation may assist in identifying signatures of transformation machinery in archaeal genomes and aid the establishment of new model genetic systems for studying archaea. [ABSTRACT FROM AUTHOR]

C. jejuni is an important food-borne pathogen displaying high genetic diversity, substantially based on natural transformation. The mechanism of DNA uptake from the environment depends on a type II secretion/type IV pilus system, whose components are partially known. Here, we quantified DNA uptake in C. jejuni at the single cell level and observed median transport capacities of approximately 30 kb per uptake location. The process appeared to be limited by the initialization of DNA uptake, was finite, and, finalized within 30 min of contact to DNA. Mutants lacking either the outer membrane pore PilQ or the inner membrane channel ComEC were deficient in natural transformation. The periplasmic DNA binding protein ComE was negligible for DNA uptake, which is in contrast to its proposed function. Intriguingly, a mutant lacking the unique periplasmic protein Cj0683 displayed rare but fully functional DNA uptake events. We conclude that Cj0683 was essential for the efficient initialization of DNA uptake, consistent with the putative function as a competence pilus protein. Unravelling features important in natural transformation might lead to target identification, reducing the adaptive potential of pathogens. [ABSTRACT FROM AUTHOR]

Natural transformation, or the uptake of naked DNA from the external milieu by bacteria, holds a unique place in the history of biology. This is both the beginning of the realization of the correct chemical nature of genes and the first technical step to the molecular biology revolution that sees us today able to modify genomes almost at will. Yet the mechanistic understanding of bacterial transformation still presents many blind spots and many bacterial systems lag behind power horse model systems like Escherichia coli in terms of ease of genetic modification. Using Neisseria gonorrhoeae as a model system and using transformation with multiple DNA molecules, we tackle in this paper both some aspects of the mechanistic nature of bacterial transformation and the presentation of new molecular biology techniques for this organism. We show that similarly to what has been demonstrated in other naturally competent bacteria, Neisseria gonorrhoeae can incorporate, at the same time, different DNA molecules modifying DNA at different loci within its genome. In particular, co-transformation of a DNA molecule bearing an antibiotic selection cassette and another non-selected DNA piece can lead to the integration of both molecules in the genome while selecting only through the selective cassette at percentages above 70%. We also show that successive selections with two selection markers at the same genetic locus can drastically reduce the number of genetic markers needed to do multisite genetic modifications in Neisseria gonorrhoeae. Despite public health interest heightened with the recent rise in antibiotic resistance, the causative agent of gonorrhea still does not possess a plethora of molecular techniques. This paper will extend the techniques available to the Neisseria community while providing some insights into the mechanisms behind bacterial transformation in Neisseria gonorrhoeae. We are providing a suite of new techniques to quickly obtain modifications of genes and genomes in the Neisserial naturally competent bacteria.

ABSTRACT Avibacterium paragallinarum is the pathogen involved in infectious coryza (IC), an acute infectious upper respiratory disease in chickens. The prevalence of IC has increased in China in recent years. There is a lack of reliable and effective procedures for gene manipulation, which has limited the research on the bacterial genetics and pathogenesis of A. paragallinarum. Natural transformation has been developed as a method of gene manipulation in Pasteurellaceae by the introduction of foreign genes or DNA fragments into bacterial cells, but there has been no report on natural transformation in A. paragallinarum. In this study, we analyzed the existence of homologous genetic factors and competence proteins underlying natural transformation in A. paragallinarum and established a method for transformation in it. Through bioinformatics analysis, we identified 16 homologs of Haemophilus influenzae competence proteins in A. paragallinarum. We found that the uptake signal sequence (USS) was overrepresented in the genome of A. paragallinarum (1,537 to 1,641 copies of the core sequence ACCGCACTT). We then constructed a plasmid, pEA-KU, that carries the USS and a plasmid, pEA-K, without the USS. These plasmids can be transferred via natural transformation into naturally competent strains of A. paragallinarum. Significantly, the plasmid that carries USS showed a higher transformation efficiency. In summary, our results demonstrate that A. paragallinarum has the ability to undergo natural transformation. These findings should prove to be a valuable tool for gene manipulation in A. paragallinarum. IMPORTANCE Natural transformation is an important mechanism for bacteria to acquire exogenous DNA molecules during the process of evolution. Additionally, it can also be used as a method to introduce foreign genes into bacteria under laboratory conditions. Natural transformation does not require equipment such as an electroporation apparatus. It is easy to perform and is similar to gene transfer under natural conditions. However, there have been no reports on natural transformation in Avibacterium paragallinarum. In this study, we analyzed the presence of homologous genetic factors and competence proteins underlying natural transformation in A. paragallinarum. Our results indicate that natural competence could be induced in A. paragallinarum serovars A, B, and C. Furthermore, the method that we established to transform plasmids into naturally competent A. paragallinarum strains was stable and efficient.

Given an ∞-bicategory D with underlying ∞-category D_0, we construct a Cartesian fibration Tw(D) --> Tw(D_0) x D_0op, which we call the enhanced twisted arrow ∞-category, classifying the restricted mapping category functor Map_D:D_0op x D_0 --> Dop\times D --> Cat_∞. With the aid of this new construction, we provide a description of the ∞-category of natural transformations Nat(F,G) as an end for any functors F and G from an ∞-category to an ∞-bicategory. As an application of our results, we demonstrate that the definition of weighted colimits studied by Gepner-Haugseng-Nikolaus satisfies the expected 2-dimensional universal property. [ABSTRACT FROM AUTHOR]

Despite a general recognition that there are two styles of think-ing: fast, reflexive and relatively effortless (Type 1) versus slow,reflective and effortful (Type 2), dual-process theories of cogni-tion remain controversial, in particular, for their vagueness. Toaddress this lack of formal precision, we take a mathematicalcategory theory approach towards understanding what under-lies the relationship between dual cognitive processes. Fromour category theory perspective, we show that distinguishingfeatures of Type 1 versus Type 2 processes are exhibited viaadjoint and representable functors. These results suggest thatcategory theory provides a useful formal framework for devel-oping dual-process theories of cognition.

Category theorists and cognitive scientists study the structural (analogical) relations between domains of interest albeit in different contexts, that is, formal and psychological systems, respectively. Despite this basic commonality, very few cognitive scientists take a category theory approach toward understanding the structure of cognition which raises the question, What is category theory to cognitive science? An answer is given as the slogan "Category theory is to cognitive science as functor is to representation; as natural transformation is to comparison" to make category theory more accessible and informative for cognitive scientists. [ABSTRACT FROM AUTHOR]

Ralstonia solanacearum species complex (RSSC) is a group of Gram-negative bacterial pathogen capable of infecting numerous plants and crops, causing severe vascular wilt diseases. Functional analysis of the genes associated with bacterial virulence is critical for elucidating the molecular mechanisms that govern the bacterial pathogenicity. To this end, an efficient gene deletion method would be of great help. In this study, we set to develop an efficient and simple markerless gene deletion method by exploiting its natural transformation competence and the FLP/FRT recombination system. We found that natural transformation using PCR products provided much higher transformation frequency than the plasmid-based triparental mating and electroporation. We thus generated the gene deletion fusion PCR fragments by incorporating the upstream and downstream DNA fragments of the target gene and an antibiotic resistance gene flanked by FRT sites, and delivered the PCR products into R. solanacearum cells through natural transformation. Using this method, we knocked out the epsB and phcA genes, which are associated with exopolysaccharide (EPS) biosynthesis and regulation, respectively, in several R. solanacearum strains isolated from different host plants at a frequency from 5 (1E-08) to 45 (1E-08). To remove the antibiotic marker gene, the plasmid expressing the FLP enzyme was introduced into the above knockout mutants, which enabled removal of the marker gene. The effective combination of natural transformation and the FLP/FRT recombination system thus offers a simple and efficient method for functional study of putative virulence genes and for elucidation of R. solanacearum pathogenic mechanisms. [ABSTRACT FROM AUTHOR]

We show that the \Sigma ^1_1- and \Sigma ^1_2-boundedness theorems extend to the category of continuous dilators. We then apply these results to conclude the corresponding theorems for the category of sharps of real numbers, thus establishing another connection between Proof Theory and Set Theory, and extending work of Girard-Normann [J. Symbolic Logic 57 (1992), pp. 659–676] and Kechris-Woodin [Ann. Pure Appl. Logic 52 (1991), pp. 93–97]. [ABSTRACT FROM AUTHOR]

In the post‐genome era, reverse genetic engineering is an indispensable methodology for experimental molecular biology to provide a deeper understanding of the principal relationship between genomic features and biological phenotypes. Technically, genetic engineering is carried out through allele replacement of a target genomic locus with a designed nucleotide sequence, so called site‐directed mutagenesis. To artificially manipulate allele replacement through homologous recombination, researchers have improved various methodologies that are optimized to the bacterial species of interest. Here, we review widely used genetic engineering technologies, particularly for streptococci and enterococci, and recent advances that enable more effective and flexible manipulation. The development of genetic engineering has been promoted by synthetic biology approaches based on basic biological knowledge of horizontal gene transfer systems, such as natural conjugative transfer, natural transformation, and the CRISPR/Cas system. Therefore, this review also describes basic insights into molecular biology that underlie improvements in genetic engineering technology. [ABSTRACT FROM AUTHOR]

Antibiotic resistance genes (ARGs) as emerging environmental contaminants exacerbate the risk of spreading antibiotic resistance. Natural organic matter (NOM) is ubiquitous in aquatic environments and plays a crucial role in biogeochemical cycles. However, its impact on the dissemination of extracellular antibiotic resistance genes (eARGs) under sunlight exposure remains elusive. This study reveals that environmentally relevant levels of NOM (0.1-20 mg/L) can significantly enhance the natural transformation frequency of the model bacterium Acinetobacter baylyi ADP1 by up to 7.6-fold under simulated sunlight. Similarly, this enhancement was consistently observed in natural water and wastewater systems. Further mechanism analysis revealed that reactive oxygen species (ROS) generated by NOM under sunlight irradiation, primarily singlet oxygen and hydroxyl radicals, play a crucial role in this process. These ROS induce intracellular oxidative stress and elevated cellular membrane permeability, thereby indirectly boosting ATP production and enhancing cell competence of extracellular DNA uptake and integration. Our findings highlight a previously underestimated role of natural factors in the dissemination of eARGs within aquatic ecosystems and deepen our understanding of the complex interplay between NOM, sunlight, and microbes in environmental water bodies. This underscores the importance of developing comprehensive strategies to mitigate the spread of antibiotic resistance in aquatic environments.

Natural transformation refers to the process in which bacteria acquire new traits by uptaking naked DNA from the environment and integrating it into their genome through homologous recombination when they are in the specialized physiological state of competence. The natural transformation was first described in Streptococcus pneumoniae. Since Frederick Griffith first described natural transformations in S. pneumoniae in 1928, this phenomenon has been studied extensively. Induction of competence before natural transformation has been reported to involve about 10% of the pneumococcal genome. In addition to natural transformation, multiple physiological processes are involved, including biofilm formation, bacteriocin production, and fratricide. In this review, we summarized current knowledge about natural transformation in S. pneumoniae and described its competence regulation mechanism. This review also introduces the development of novel drugs and vaccines against S. pneumoniae infection by utilizing the existing knowledge of competence and natural transformation., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

The sexually transmitted pathogen, Neisseria gonorrhoeae, undergoes natural transformation at high frequency. This property has led to the rapid dissemination of antibiotic resistance markers and the panmictic structure of the gonococcal population. However, high-frequency transformation also makes N. gonorrhoeae one of the easiest bacterial species to manipulate genetically in the laboratory. Techniques have been developed that result in transformation frequencies >50%, allowing the identification of mutants by screening and without selection. Constructs have been created to take advantage of this high-frequency transformation, facilitating genetic mutation, complementation, and heterologous gene expression. Similar methods have been developed for N. meningitidis and nonpathogenic Neisseria including N. mucosa and N. musculi. Techniques are described for genetic manipulation of N. gonorrhoeae and commensal Neisseria species, as well as for growth of these fastidious organisms. © 2024 The Author(s). Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Spot transformation of Neisseria gonorrhoeae on agar plates Basic Protocol 2: Spot transformation of commensal Neisseria on agar plates Basic Protocol 3: Transformation of Neisseria gonorrhoeae in liquid culture Basic Protocol 4: Electroporation of Neisseria gonorrhoeae Basic Protocol 5: Creation of unmarked mutations using a positive and negative selection cassette Basic Protocol 6: In vitro mutagenesis of Neisseria gonorrhoeae chromosomal DNA using EZ-Tn5 Basic Protocol 7: Chemical mutagenesis Basic Protocol 8: Complementation on the Neisseria gonorrhoeae chromosome Alternate Protocol 1: Complementation with replicating plasmids Alternate Protocol 2: Complementation on the Neisseria musculi or Neisseria mucosa chromosome Basic Protocol 9: Preparation of chromosomal DNA from Neisseria gonorrhoeae grown on solid medium Alternate Protocol 3: Preparation of chromosomal DNA from Neisseria gonorrhoeae grown in broth Support Protocol: Preparing PCR templates from Neisseria gonorrhoeae colonies., (© 2024 The Author(s). Current Protocols published by Wiley Periodicals LLC.)

Category theorists and cognitive scientists study the structural (analogical) relations between domains of interest albeit in different contexts, that is, formal and psychological systems, respectively. Despite this basic commonality, very few cognitive scientists take a category theory approach toward understanding the structure of cognition which raises the question, What is category theory to cognitive science? An answer is given as the slogan “Category theory is to cognitive science as functor is to representation; as natural transformation is to comparison” to make category theory more accessible and informative for cognitive scientists.

Glaesserella parasuis is an important bacterial pathogen that affects the swine industry worldwide. Research on the pathogenic mechanism and genetically engineered vaccine remains undeveloped because an effective markerless and multiple-gene knockout system is unavailable for G. parasuis yet. To establish a markerless knockout, deleted allelic genes with kanamycin resistance (KanR) cassettes were introduced into the genome of G. parasuis by using natural transformation with suicide plasmids. Then, the KanR cassette was excised with a thermosensitive plasmid pGF conferring a constitutive Flp expression. To realize the markerless and multiple-gene knockout, plasmid pGAF was constructed by placing the Flp gene under the control of an arabinose-inducible promoter. Firstly, pGAF was introduced into G. parasuis by electroporation, and the marked mutants were produced following natural transformation. Finally, the KanR cassette was excised from the genome by the inducible expression of Flp upon arabinose action. Based on the natural transformation and the inducible expression of Flp, the markerless single-gene knockout mutants of ΔhsdR, ΔneuA2, ΔespP2, Δapd, and ΔnanH were constructed. In addition, a five-gene knockout mutant of ΔhsdRΔneuA2ΔespP2ΔapdΔnanH was generated by successive natural transformation with five suicide plasmids. Taken together, a markerless and multiple-gene deletion system was established for G. parasuis in the present study for the first time. This system is simple, efficient, and easy to manipulate for G. parasuis; thus, our technique will substantially aid the understanding of the etiology, pathogenesis, and genetic engineering of G. parasuis and other bacteria that can be naturally transformed in laboratory conditions. Key points: • Flp recombinase excised the KanR gene flanked by FRT sites in Glaesserella parasuis. • The regulatory expression of Flp enabled a multiple-gene knockout forG. parasuis. • The technique will promote the understanding of Glässer's disease pathogens. [ABSTRACT FROM AUTHOR]

Natural transformation is a mechanism by which a particular bacterial species takes up foreign DNA and integrates it into its genome. The swine pathogen Glaesserella parasuis (G. parasuis) is a naturally transformable bacterium. The regulation of competence, however, is not fully understood. In this study, the natural transformability of 99 strains was investigated. Only 44% of the strains were transformable under laboratory conditions. Through a high-resolution melting curve and phylogenetic analysis, we found that genetic differences in the core regulator of natural transformation, the tfoX gene, leads to two distinct natural transformation phenotypes. In the absence of the tfoX gene, the highly transformable strain SC1401 lost its natural transformability. In addition, when the SC1401 tfoX gene was replaced by the tfoX of SH0165, which has no natural transformability, competence was also lost. These results suggest that TfoX is a core regulator of natural transformation in G. parasuis, and that differences in tfoX can be used as a molecular indicator of natural transformability. Transcriptomic and proteomic analyses of the SC1401 wildtype strain, and a tfoX gene deletion strain showed that differential gene expression and protein synthesis is mainly centered on pathways related to glucose metabolism. The results suggest that tfoX may mediate natural transformation by regulating the metabolism of carbon sources. Our study provides evidence that tfoX plays an important role in the natural transformation of G. parasuis. [ABSTRACT FROM AUTHOR]

The biotechnologically important and naturally transformable cyanobacterium, Synechococcus elongatus PCC 7942, possesses multiple genome copies irrespective of its growth rate or condition. Hence, segregating mutations across all genome copies typically takes several weeks. In this study, Synechococcus 7942 cultivation on a solid growth medium was optimised using different concentrations of agar, the addition of antioxidants, and overexpression of the catalase gene to facilitate the rapid acquisition of colonies and fully segregated lines. Synechococcus 7942 was grown at different temperatures and nutritional conditions. The miniploid cells were identified using flow cytometry and fluorimetry. The natural transformation was carried out using miniploid cells and validated with PCR and high performance liquid chromatography (HPLC). We identified that 0.35% agar concentration and 200 IU of catalase could improve the growth of Synechococcus 7942 on a solid growth medium. Furthermore, overexpression of a catalase gene enhanced the growth rate and supported diluted culture to grow on a solid medium. Our results reveal that high temperature and phosphate-depleted cells contain the lowest genome copies (2.4 ± 0.3 and 1.9 ± 0.2) and showed the potential to rapidly produce fully segregated mutants. In addition, higher antibiotic concentrations improve the selection of homozygous transformants while maintaining similar genome copies at a constant temperature. Based on our observation, we have an improved cultivation and natural transformation protocol for Synechococcus 7942 by optimising solid media culturing, generating low-ploidy cells that ultimately reduced the time required for the complete segregation of engineered lines. [ABSTRACT FROM AUTHOR]

Nowadays, the growing human population exacerbates the need for sustainable resources. Inspiration and achievements in nutrient production or human/animal health might emanate from microorganisms and their adaptive strategies. Here, we exemplify the benefits of lactic acid bacteria (LAB) for numerous biotechnological applications and showcase their natural transformability as a fast and robust method to hereditarily influence their phenotype/traits in fundamental and applied research contexts. We described the biogenesis of the transformation machinery and we analyzed the genome of hundreds of LAB strains exploitable for human needs to predict their transformation capabilities. Finally, we provide a stepwise rational path to stimulate and optimize natural transformation with standard and synthetic biology techniques. A comprehensive understanding of the molecular mechanisms driving natural transformation will facilitate and accelerate the improvement of bacteria with properties that serve broad societal interests. [ABSTRACT FROM AUTHOR]

Motivated by the category of probabilistic convergence spaces — a supercategory of the category of topological spaces; recently, we brought to light the categories of probabilistic convergence groups, probabilistic metric probabilistic convergence groups, probabilistic convergence transformation groups, along with their underpinning natural examples. The purpose of this paper is, first, to establish a result on the isomorphism between the categories of probabilistic metric groups, and probabilistic metric probabilistic convergence groups. Second, among others, we explore a monad in relation with probabilistic convergence groups, and probabilistic convergence spaces, and their related algebras. In so doing, we consider a product of the categories of probabilistic convergence groups and probabilistic convergence spaces in an attempt to construct a monad on it such that the corresponding category of algebras, the so-called Eilenberg–Moore category, is isomorphic to the category of probabilistic convergence transformation groups. Finally, invoking so-called Beck's theorem on characterization of algebras, and starting with a particular adjunction, we achieve a monad. Conversely, given a monad, we obtain an adjunction which coincides with the original monad. [ABSTRACT FROM AUTHOR]

ABSTRACT Lateral gene transfer plays an important role in the evolution of genetic diversity in prokaryotes. DNA transfer via natural transformation depends on the ability of recipient cells to actively transport DNA from the environment into the cytoplasm, termed natural competence, which relies on the presence of type IV pili and other competence proteins. Natural competence has been described in cyanobacteria for several organisms, including unicellular and filamentous species. However, natural competence in cyanobacteria that differentiate specialized cells for N2-fixation (heterocysts) and form branching or multiseriate cell filaments (termed subsection V) remains unknown. Here, we show that genes essential for natural competence are conserved in subsection V cyanobacteria. Furthermore, using the replicating plasmid pRL25C, we experimentally demonstrate natural competence in a subsection V organism: Chlorogloeopsis fritschii PCC 6912. Our results suggest that natural competence is a common trait in cyanobacteria forming complex cell filament morphologies. IMPORTANCE Cyanobacteria are crucial players in the global biogeochemical cycles, where they contribute to CO2- and N2-fixation. Their main ecological significance is the primary biomass production owing to oxygenic photosynthesis. Cyanobacteria are a diverse phylum, in which the most complex species differentiate specialized cell types and form true-branching or multiseriate cell filament structures (termed subsection V cyanobacteria). These bacteria are considered a peak in the evolution of prokaryotic multicellularity. Among others, species in that group inhabit fresh and marine water habitats, soil, and extreme habitats such as thermal springs. Here, we show that the core genes required for natural competence are frequent in subsection V cyanobacteria and demonstrate for the first time natural transformation in a member of subsection V. The prevalence of natural competence has implications for the role of DNA acquisition in the genome evolution of cyanobacteria. Furthermore, the presence of mechanisms for natural transformation opens up new possibilities for the genetic modification of subsection V cyanobacteria.

The biotechnologically important and naturally transformable cyanobacterium, Synechococcus elongatus PCC 7942, possesses multiple genome copies irrespective of its growth rate or condition. Hence, segregating mutations across all genome copies typically takes several weeks. In this study, Synechococcus 7942 cultivation on a solid growth medium was optimised using different concentrations of agar, the addition of antioxidants, and overexpression of the catalase gene to facilitate the rapid acquisition of colonies and fully segregated lines. Synechococcus 7942 was grown at different temperatures and nutritional conditions. The miniploid cells were identified using flow cytometry and fluorimetry. The natural transformation was carried out using miniploid cells and validated with PCR and high performance liquid chromatography (HPLC). We identified that 0.35% agar concentration and 200 IU of catalase could improve the growth of Synechococcus 7942 on a solid growth medium. Furthermore, overexpression of a catalase gene enhanced the growth rate and supported diluted culture to grow on a solid medium. Our results reveal that high temperature and phosphate-depleted cells contain the lowest genome copies (2.4 ± 0.3 and 1.9 ± 0.2) and showed the potential to rapidly produce fully segregated mutants. In addition, higher antibiotic concentrations improve the selection of homozygous transformants while maintaining similar genome copies at a constant temperature. Based on our observation, we have an improved cultivation and natural transformation protocol for Synechococcus 7942 by optimising solid media culturing, generating low-ploidy cells that ultimately reduced the time required for the complete segregation of engineered lines.

C. jejuni is an important food-borne pathogen displaying high genetic diversity, substantially based on natural transformation. The mechanism of DNA uptake from the environment depends on a type II secretion/type IV pilus system, whose components are partially known. Here, we quantified DNA uptake in C. jejuni at the single cell level and observed median transport capacities of approximately 30 kb per uptake location. The process appeared to be limited by the initialization of DNA uptake, was finite, and, finalized within 30 min of contact to DNA. Mutants lacking either the outer membrane pore PilQ or the inner membrane channel ComEC were deficient in natural transformation. The periplasmic DNA binding protein ComE was negligible for DNA uptake, which is in contrast to its proposed function. Intriguingly, a mutant lacking the unique periplasmic protein Cj0683 displayed rare but fully functional DNA uptake events. We conclude that Cj0683 was essential for the efficient initialization of DNA uptake, consistent with the putative function as a competence pilus protein. Unravelling features important in natural transformation might lead to target identification, reducing the adaptive potential of pathogens.

Antibiotic-resistant bacteria and the spread of antibiotic resistance genes (ARGs) pose a serious risk to human and veterinary health. While many studies focus on the movement of live antibiotic-resistant bacteria to the environment, it is unclear whether extracellular ARGs (eARGs) from dead cells can transfer to live bacteria to facilitate the evolution of antibiotic resistance in nature. Here, we use eARGs from dead, antibiotic-resistant Pseudomonas stutzeri cells to track the movement of eARGs to live P. stutzeri cells via natural transformation, a mechanism of horizontal gene transfer involving the genomic integration of eARGs. In sterile, antibiotic-free agricultural soil, we manipulated the eARG concentration, soil moisture, and proximity to eARGs. We found that transformation occurred in soils inoculated with just 0.25 mg of eDNA g-1 soil, indicating that even low concentrations of soil eDNA can facilitate transformation (previous estimates suggested ~2 to 40 µg eDNA g-1 soil). When eDNA was increased to 5 mg g-1 soil, there was a 5-fold increase in the number of antibiotic-resistant P. stutzeri cells. We found that eARGs were transformed under soil moistures typical of terrestrial systems (5 to 30% gravimetric water content) but inhibited at very high soil moistures (.30%). Overall, this work demonstrates that dead bacteria and their eARGs are an overlooked path to antibiotic resistance. More generally, the spread of eARGs in antibiotic-free soil suggests that transformation allows genetic variants to establish in the absence of antibiotic selection and that the soil environment plays a critical role in regulating transformation. [ABSTRACT FROM AUTHOR]

A Multiplicative-Exponential Linear Logic (MELL) proof-structure can be expanded into a set of resource proof-structures: its Taylor expansion. We introduce a new criterion characterizing (and deciding in the finite case) those sets of resource proof-structures that are part of the Taylor expansion of some MELL proof-structure, through a rewriting system acting both on resource and MELL proof-structures. We also prove semi-decidability of the type inhabitation problem for cut-free MELL proof-structures. [ABSTRACT FROM AUTHOR]

Vibrio natriegens is a gram-negative bacterium, which has received increasing attention due to its very fast growth with a doubling time of under 10 min under optimal conditions. To enable a wide range of projects spanning from basic research to biotechnological applications, we developed NT-CRISPR as a new method for genome engineering. This book chapter provides a step-by-step protocol for the use of this previously published tool. NT-CRISPR combines natural transformation with counterselection through CRISPR-Cas9. Thereby, genomic regions can be deleted, foreign sequences can be integrated, and point mutations can be introduced. Furthermore, up to three simultaneous modifications are possible., (© 2025. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

In this article, exact solutions of some Laplace-type fractional boundary value problems (FBVPs) are investigated via natural decomposition method. The fractional derivatives are described within Caputo operator. The natural decomposition technique is applied for the first time to boundary value problems (BVPs) and found to be an excellent tool to solve the suggested problems. The graphical representation of the exact and derived results is presented to show the reliability of the suggested technique. The present study is mainly concerned with the approximate analytical solutions of some FBVPs. Moreover, the solution graphs have shown that the actual and approximate solutions are very closed to each other. The comparison of the proposed and variational iteration methods is done for integer-order problems. The comparison, support strong relationship between the results of the suggested techniques. The overall analysis and the results obtained have confirmed the effectiveness and the simple procedure of natural decomposition technique for obtaining the solution of BVPs.

ABSTRACT Helicobacter pylori is a human pathogen competent for natural transformation. Intrinsic and acquired antibiotic resistance contribute to the survival and multiplication of H. pylori under antibiotics. While drug-resistance dissemination by natural transformation (NT)-mediated horizontal gene transfer remains poorly understood in H. pylori. The purpose of the study was to investigate the role of H. pylori porins (HopA, HopB, HopC, HopD, and HopE) in the intrinsic antibiotic resistance and to preliminarily reveal the potential effect of HopE and HopD porins in streptomycin resistance acquisition after NT in the presence of antibiotics. Using traditional antibiotic susceptibility tests and growth curve analysis, we found the MIC values of metronidazole, clarithromycin, levofloxacin, tetracycline, rifampin, and streptomycin in mutants lacking HopE and/or HopD were significantly elevated compare to those in wild-type strain. The quantitative analysis of the tetramethyl rhodamine isothiocyanate (TRITC)-labeled streptomycin accumulation at the single-cell level showed reduced streptomycin intracellular fluorescence in ΔhopE and ΔhopD mutant cells. Furthermore, in the presence of translation-inhibiting antibiotic streptomycin, the resistance acquisition frequency was decreased in the wild-type strain, which could be reversed by mutants lacking HopE and HopD that restored relatively high resistance acquisition frequencies. By transforming a pUC19-rpsLmut-sfgfp linear plasmid carrying a streptomycin conferring mutation, we observed that the impaired ability of rpsLmut synthesis in the wild-type strain was restored in the ΔhopE and ΔhopD mutant transformants. Our study revealed that in the presence of streptomycin, resistance acquisition at least partially relied on the deletion of the hopE and hopD genes, because their loss reduced streptomycin concentration in the cell and thus restored the expression of the resistance-conferring gene, which was inhibited by streptomycin in wild-type strain. The loss of HopE and HopD influx activity may also preserve resistance acquisition by transformation in the presence of antibiotics with other modes of action. IMPORTANCE Helicobacter pylori is constitutively competent for natural transformation (NT) and possesses an efficient system for homologous recombination, which could be utilized to study the NT-mediated horizontal gene transfer induced antibiotic resistance acquisition. Bacterial porins have drawn renewed attention because of their crucial role in antibiotic susceptibility. From the perspective of porin-mediated influx in H. pylori, our study preliminarily revealed the important role of HopE and HopD porins not only in preserving the intrinsic susceptibility to specific antibiotic but also in evading acquired antibiotic resistance by NT in the presence of translation-inhibiting antimicrobial. Therefore, the loss of HopE or HopD porin in H. pylori genomes, combined with the large number of secreted or cell-free genetic elements carrying mutations conferring antibiotic resistance, may raise the possibility that this mechanism plays a potential role in the propagation of antibiotic resistance within H. pylori communities.

This study characterizes a total of 21 wastewater samples collected from Al Amal hospital, and aims to determine if untreated hospital wastewater may impose a potentially detrimental impact on the downstream municipal biological wastewater treatment process. By means of solid phase extraction and liquid chromatography with tandem mass spectrometry (LC-MS/MS), chemical contaminants in these wastewater samples were determined in a non-targeted manner. In-silico characterization for the mutagenicity and reactive oxygen species (ROS) producing capabilities was performed by checking against database and literature. However, majority of the chemical contaminants have no prior information available and remain uncharacterized for both traits. Instead, in-vitro mutagenicity tests by means of Ames test showed that majority of the samples were non-mutagenic except for 5 samples that imposed mutagenic effect at high concentrations of >×10. In-vitro tests to determine for intracellular ROS production further showed that one of the mutagenic samples collected on Jun-22 positively induce ROS production and subsequently increased horizontal gene transfer via natural transformation. The findings in this study suggest that a specialty hospital like Al Amal does not frequently contribute mutagenic compounds and ROS to the wastewater streams, and in instances where it contributed positively, would require a high concentration to do so. Hence in general, wastewater streams from a specialty hospital like Al Amal may be unlikely to significantly perturb the downstream environment.

Acinetobacter baumannii is a serious threat to public health, and there is increasing attention to the development of antibiotic resistance in this bacterium. Natural transformation is a major horizontal gene transfer mechanism that can lead to antibiotic resistance. To better understand the mechanism of natural transformation in A. baumannii , we selected a clinical isolate that was transformable but had no visible extracellular type IV pili (T4P) filaments and then examined the effects of multiple single-gene knockouts on natural plasmid transformation. Among 33 candidate genes, 28 knockout mutants had severely or completely impaired transformability. Some of these genes had established roles in T4P biogenesis; DNA transfer across the outer membrane, periplasm, or inner membrane; and protection of intracellular single-stranded DNA (ssDNA). Other genes had no previously reported roles in natural transformation of A. baumannii , including competence activator cAMP receptor protein (CRP), a periplasmic protein that may function in T4P assembly (TonB2), a T4P secretin-associated protein (TsaP), and two type II secretion system (T2SS) minor pseudopilus assembly prime complex competent proteins (XcpV and XcpW). The deletion of the T2SS assembly platform protein X had no effect on transformation, and the minor pseudopilins were capable of initiating major pilin assembly. Thus, we speculate that XcpV and XcpW may function in DNA uptake with major pilin assembly, a non-T2SS-dependent mechanism and that a competence pseudopilus similar to T4P constituted the central part of the DNA uptake complex. These results may help guide future research on the alarming increase of antibiotic resistance in this pathogen. [ABSTRACT FROM AUTHOR]

Acinetobacter baumannii is a serious threat to public health, and there is increasing attention to the development of antibiotic resistance in this bacterium. Natural transformation is a major horizontal gene transfer mechanism that can lead to antibiotic resistance. To better understand the mechanism of natural transformation in A. baumannii , we selected a clinical isolate that was transformable but had no visible extracellular type IV pili (T4P) filaments and then examined the effects of multiple single-gene knockouts on natural plasmid transformation. Among 33 candidate genes, 28 knockout mutants had severely or completely impaired transformability. Some of these genes had established roles in T4P biogenesis; DNA transfer across the outer membrane, periplasm, or inner membrane; and protection of intracellular single-stranded DNA (ssDNA). Other genes had no previously reported roles in natural transformation of A. baumannii , including competence activator cAMP receptor protein (CRP), a periplasmic protein that may function in T4P assembly (TonB2), a T4P secretin-associated protein (TsaP), and two type II secretion system (T2SS) minor pseudopilus assembly prime complex competent proteins (XcpV and XcpW). The deletion of the T2SS assembly platform protein X had no effect on transformation, and the minor pseudopilins were capable of initiating major pilin assembly. Thus, we speculate that XcpV and XcpW may function in DNA uptake with major pilin assembly, a non-T2SS-dependent mechanism and that a competence pseudopilus similar to T4P constituted the central part of the DNA uptake complex. These results may help guide future research on the alarming increase of antibiotic resistance in this pathogen. [ABSTRACT FROM AUTHOR]

The remarkable genomic plasticity of Streptococcus pneumoniae largely depends on its ability to undergo natural genetic transformation. To take up extracellular DNA, S. pneumoniae assembles competence pili composed of the major pilin ComGC. In addition to ComGC, four minor pilins ComGD, E, F, and G are expressed during bacterial competence, but their role in pilus biogenesis and transformation is unknown. Here, using a combination of protein-protein interaction assays we show that all four proteins can directly interact with each other. Pneumococcal ComGG stabilizes the minor pilin ComGD and ComGF and can interact with and stabilize the major pilin ComGC, thus, deletion of ComGG abolishes competence pilus assembly. We further demonstrate that minor pilins are present in sheared pili fractions and find ComGF to be incorporated along the competence pilus by immunofluorescence and electron microscopy. Finally, mutants of the invariant Glu5 residue (E5), ComGDE5A or ComGEE5A, but not ComGFE5A, were severely impaired in pilus formation and function. Together, our results suggest that ComGG, lacking E5, is essential for competence pilus assembly and function, and plays a central role in connecting the pneumococcal minor pilins to ComGC. [ABSTRACT FROM AUTHOR]

We describe completely the so called jet like functors of a vector bundle E over an m-dimensional manifold M, i.e. bundles FE over M canonically depending on E such that F(E1 ×M E2) = FE1 ×M FE2 for any vector bundles E1 and E2 over M. Then we study how a linear vector field on E can induce canonically a vector field on FE. [ABSTRACT FROM AUTHOR]

In the present research article, we implemented two well-known analytical techniques to solve fractional-order multi-dimensional Navier–Stokes equation. The proposed methods are the modification of Adomian decomposition method and variational iteration method by using natural transformation. Furthermore, some illustrative examples are presented to confirm the validity of the suggested methods. The solutions graphs and tables are constructed for both fractional and integer-order problems. It is investigated that the suggested techniques have the identical solutions of the problems. The solution comparison via graphs and tables have also supported the greater accuracy and higher rate of convergence of the present methods.

Acinetobacter baumannii is a serious threat to public health, and there is increasing attention to the development of antibiotic resistance in this bacterium. Natural transformation is a major horizontal gene transfer mechanism that can lead to antibiotic resistance. To better understand the mechanism of natural transformation in A. baumannii, we selected a clinical isolate that was transformable but had no visible extracellular type IV pili (T4P) filaments and then examined the effects of multiple single-gene knockouts on natural plasmid transformation. Among 33 candidate genes, 28 knockout mutants had severely or completely impaired transformability. Some of these genes had established roles in T4P biogenesis; DNA transfer across the outer membrane, periplasm, or inner membrane; and protection of intracellular single-stranded DNA (ssDNA). Other genes had no previously reported roles in natural transformation of A. baumannii, including competence activator cAMP receptor protein (CRP), a periplasmic protein that may function in T4P assembly (TonB2), a T4P secretin-associated protein (TsaP), and two type II secretion system (T2SS) minor pseudopilus assembly prime complex competent proteins (XcpV and XcpW). The deletion of the T2SS assembly platform protein X had no effect on transformation, and the minor pseudopilins were capable of initiating major pilin assembly. Thus, we speculate that XcpV and XcpW may function in DNA uptake with major pilin assembly, a non-T2SS-dependent mechanism and that a competence pseudopilus similar to T4P constituted the central part of the DNA uptake complex. These results may help guide future research on the alarming increase of antibiotic resistance in this pathogen.

The remarkable genomic plasticity of Streptococcus pneumoniae largely depends on its ability to undergo natural genetic transformation. To take up extracellular DNA, S. pneumoniae assembles competence pili composed of the major pilin ComGC. In addition to ComGC, four minor pilins ComGD, E, F, and G are expressed during bacterial competence, but their role in pilus biogenesis and transformation is unknown. Here, using a combination of protein-protein interaction assays we show that all four proteins can directly interact with each other. Pneumococcal ComGG stabilizes the minor pilin ComGD and ComGF and can interact with and stabilize the major pilin ComGC, thus, deletion of ComGG abolishes competence pilus assembly. We further demonstrate that minor pilins are present in sheared pili fractions and find ComGF to be incorporated along the competence pilus by immunofluorescence and electron microscopy. Finally, mutants of the invariant Glu5 residue (E5), ComGDE5A or ComGEE5A, but not ComGFE5A, were severely impaired in pilus formation and function. Together, our results suggest that ComGG, lacking E5, is essential for competence pilus assembly and function, and plays a central role in connecting the pneumococcal minor pilins to ComGC.

Streptococcus dysgalactiae is an emerging human and animal pathogen. Functional studies of genes involved in virulence of S. dysgalactiae and other pyogenic group streptococci are often hampered by limited genetic tractability. It is known that pyogenic streptococci carry genes required for competence for natural transformation; however, in contrast to other streptococcal subgroups, there is limited evidence for gene transfer by natural transformation in these bacteria. In this study, we systematically assessed the genomes of 179 S . dysgalactiae strains of both human and animal origins (subsp. equisimilis and dysgalactiae, respectively) for the presence of genes required for natural transformation. While a considerable fraction of the strains contained inactive genes, the majority (64.2%) of the strains had an intact gene set. In selected strains, we examined the dynamics of competence activation after addition of competence-inducing pheromones using transcriptional reporter assays and exploratory RNA-seq. Based on these findings, we were able to establish a protocol allowing us to utilize natural transformation to construct deletion mutants by allelic exchange in several S. dysgalactiae strains of both subspecies. As part of the work, we deleted putative lactose utilization genes to study their role in growth on lactose. The data presented here provide new knowledge on the potential of horizonal gene transfer by natural transformation in S. dysgalactiae and, importantly, demonstrates the possibility to exploit natural transformation for genetic engineering in these bacteria., Importance: Numerous Streptococcus spp. exchange genes horizontally through natural transformation, which also facilitates efficient genetic engineering in these organisms. However, for the pyogenic group of streptococci, including the emerging pathogen Streptococcus dysgalactiae , there is limited experimental evidence for natural transformation. In this study, we demonstrate that natural transformation in vitro indeed is possible in S. dysgalactiae strains under optimal conditions. We utilized this method to perform gene deletion through allelic exchange in several strains, thereby paving the way for more efficient gene engineering methods in pyogenic streptococci., Competing Interests: The authors declare no conflict of interest.