Pathogenomics analysis of high-risk clone ST147 multidrug-resistant Klebsiella pneumoniae isolated from a patient in Egypt.

BACKGROUND: The emergence of multi-drug-resistant Klebsiella pneumoniae (MDR-KP) represents a serious clinical health concern. Antibiotic resistance and virulence interactions play a significant role in the pathogenesis of K. pneumoniae infections. Therefore, tracking the clinical resistome and virulome through monitoring antibiotic resistance genes (ARG) and virulence factors in the bacterial genome using computational analysis tools is critical for predicting the next epidemic. METHODS: In the current study, one hundred extended spectrum ß-lactamase (ESBL)-producing clinical isolates were collected from Mansoura University Hospital, Egypt, in a six-month period from January to June 2022. One isolate was selected due to the high resistance phenotype, and the genetic features of MDR-KP recovered from hospitalized patient were investigated. Otherwise, the susceptibility to 25 antimicrobials was determined using the DL Antimicrobial Susceptibility Testing (AST) system. Whole genome sequencing (WGS) using Illumina NovaSeq 6000 was employed to provide genomic insights into K. pneumoniae WSF99 clinical isolate. RESULTS: The isolate K. pneumoniae WSF99 was phenotypically resistant to the antibiotics under investigation via antibiotic susceptibility testing. WGS analysis revealed that WSF99 total genome length was 5.7 Mb with an estimated 5,718 protein-coding genes and a G + C content of 56.98 mol%. Additionally, the allelic profile of the WSF99 isolate was allocated to the high-risk clone ST147. Furthermore, diverse antibiotic resistance genes were determined in the genome that explain the high-level resistance phenotypes. Several ß-lactamase genes, including blaCTX-M-15, blaTEM-1, blaTEM-12, blaSHV-11, blaSHV-67, and blaOXA-9, were detected in the WSF99 isolate. Moreover, a single carbapenemase gene, blaNDM-5, was predicted in the genome, positioned within a mobile cassette. In addition, other resistance genes were predicted in the genome including, aac(6')-Ib, aph(3')-VI, sul1, sul2, fosA, aadA, arr-2, qnrS1, tetA and tetC. Four plasmid replicons CoIRNAI, IncFIB(K), IncFIB(pQil), and IncR were predicted in the genome. The draft genome analysis revealed the occurrence of genetic mobile elements positioned around the ARGs, suggesting the ease of dissemination via horizontal gene transfer. CONCLUSIONS: This study reports a comprehensive pathogenomic analysis of MDR-KP isolated from a hospitalized patient. These findings could be relevant for future studies investigating the diversity of antimicrobial resistance and virulence in Egypt.

Deciphering the reduction of antibiotic resistance genes (ARGs) during medium-chain fatty acids production from waste activated sludge: Driven by inhibition of ARGs transmission and shift of microbial community.

Medium-chain fatty acids (MCFAs) production from waste activated sludge (WAS) by chain extension (CE) is a promising technology. However, the effects and mechanisms of CE process on the fate of antibiotic resistance genes (ARGs) remain unclear. In this study, the results showed that the removal efficiency of ARGs was 81.15 % in CE process, suggesting its efficacy in reducing environmental risks. Further, the observed decrease in mobile genetic elements (MGEs) indicated that CE process restricted the horizontal gene transfer (HGT). Complementing this, the increase in soluble organic matters and extracellular 16 S rDNA confirmed that MCFAs production caused bacterial damage. Decreased intracellular ARGs and increased extracellular ARGs further revealed that MCFAs production impaired ARGs hosts, thereby limiting the vertical gene transfer (VGT) of ARGs. Shift of microbial community combined with co-occurrence network analysis demonstrated that functional bacteria without host potential for ARGs were enriched, but potential ARGs and MGEs hosts decreased, showing the role of functional bacterial phylogeny and selection pressure of MCFAs in reducing ARGs. Finally, partial least squares path model was used to systematic verify the mechanism of ARGs removal in CE process, which was attributed to the inhibition of ARGs transmission (HGT and VGT) and shift of microbial community.

Multitasking functions of bacterial extracellular DNA in biofilms.

Bacterial biofilms are intricate ecosystems of microbial communities that adhere to various surfaces and are enveloped by an extracellular matrix composed of polymeric substances. Within the context of bacterial biofilms, extracellular DNA (eDNA) originates from cell lysis or is actively secreted, where it exerts a significant influence on the formation, stability, and resistance of biofilms to environmental stressors. The exploration of eDNA within bacterial biofilms holds paramount importance in research, with far-reaching implications for both human health and the environment. An enhanced understanding of the functions of eDNA in biofilm formation and antibiotic resistance could inspire the development of strategies to combat biofilm-related infections and improve the management of antibiotic resistance. This comprehensive review encapsulates the latest discoveries concerning eDNA, encompassing its origins, functions within bacterial biofilms, and significance in bacterial pathogenesis.

Comparative genomic analyses of Cutibacterium granulosum provide insights into genomic diversity.

Cutibacterium granulosum, a commensal bacterium found on human skin, formerly known as Propionibacterium granulosum, rarely causes infections and is generally considered non-pathogenic. Recent research has revealed the transferability of the multidrug-resistant plasmid pTZC1 between C. granulosum and Cutibacterium acnes, the latter being an opportunistic pathogen in surgical site infections. However, there is a noticeable lack of research on the genome of C. granulosum, and the genetic landscape of this species remains largely uncharted. We investigated the genomic features and evolutionary structure of C. granulosum by analyzing a total of 30 Metagenome-Assembled Genomes (MAGs) and isolate genomes retrieved from public databases, as well as those generated in this study. A pan-genome of 6,077 genes was identified for C. granulosum. Remarkably, the 'cloud genes' constituted 62.38% of the pan-genome. Genes associated with mobilome: prophages, transposons [X], defense mechanisms [V] and replication, recombination and repair [L] were enriched in the cloud genome. Phylogenomic analysis revealed two distinct mono-clades, highlighting the genomic diversity of C. granulosum. The genomic diversity was further confirmed by the distribution of Average Nucleotide Identity (ANI) values. The functional profiles analysis of C. granulosum unveiled a wide range of potential Antibiotic Resistance Genes (ARGs) and virulence factors, suggesting its potential tolerance to various environmental challenges. Subtype I-E of the CRISPR-Cas system was the most abundant in these genomes, a feature also detected in C. acnes genomes. Given the widespread distribution of C. granulosum strains within skin microbiome, our findings make a substantial contribution to our broader understanding of the genetic diversity, which may open new avenues for investigating the mechanisms and treatment of conditions such as acne vulgaris.

Genomic insights into the evolution and adaptation of secondary metabolite gene clusters in fungicolous species Cladobotryum mycophilum ATHUM6906.

Mycophilic or fungicolous fungi can be found wherever fungi exist since they are able to colonize other fungi, which occupy a diverse range of habitats. Some fungicolous species cause important diseases on Basidiomycetes, and thus, they are the main reason for the destruction of mushroom cultivations. Nonetheless, despite their ecological significance, their genomic data remain limited. Cladobotryum mycophilum is one of the most aggressive species of the genus, destroying the economically important Agaricus bisporus cultivations. The 40.7 Mb whole genome of the Greek isolate ATHUM6906 is assembled in 16 fragments, including the mitochondrial genome and 2 small circular mitochondrial plasmids, in this study. This genome includes a comprehensive set of 12,282 protein coding, 56 rRNA, and 273 tRNA genes. Transposable elements, CAZymes, and pathogenicity related genes were also examined. The genome of C. mycophilum contained a diverse arsenal of genes involved in secondary metabolism, forming 106 biosynthetic gene clusters, which renders this genome as one of the most BGC abundant among fungicolous species. Comparative analyses were performed for genomes of species of the family Hypocreaceae. Some BGCs identified in C. mycophilum genome exhibited similarities to clusters found in the family Hypocreaceae, suggesting vertical heritage. In contrast, certain BGCs showed a scattered distribution among Hypocreaceae species or were solely found in Cladobotryum genomes. This work provides evidence of extensive BGC losses, horizontal gene transfer events, and formation of novel BGCs during evolution, potentially driven by neutral or even positive selection pressures. These events may increase Cladobotryum fitness under various environmental conditions and potentially during host-fungus interaction.

Insights into the role of extracellular polymeric substances (EPS) in the spread of antibiotic resistance genes.

Antibiotic resistance genes (ARG) are prevalent in aquatic environments. Discharge from wastewater treatment plants is an important point source of ARG release into the environment. It has been reported that biological treatment processes may enhance rather than remove ARG because of their presence in sludge. Attenuation of ARG in biotechnological processes has been studied in depth, showing that many microorganisms can secrete complex extracellular polymeric substances (EPS). These EPS can serve as multifunctional elements of microbial communities, involving aspects, such as protection, structure, recognition, adhesion, and physiology. These aspects can influence the interaction between microbial cells and extracellular ARG, as well as the uptake of extracellular ARG by microbial cells, thus changing the transformative capability of extracellular ARG. However, it remains unclear whether EPS can affect horizontal ARG transfer, which is one of the main processes of ARG dissemination. In light of this knowledge gap, this review provides insight into the role of EPS in the transmission of ARGs; furthermore, the mechanism of ARG spread is analyzed, and the molecular compositions and functional properties of EPS are summarized; also, how EPS influence ARG mitigation is addressed, and factors impacting how EPS facilitate ARG during wastewater treatment are summarized. This review provides comprehensive insights into the role of EPS in controlling the transport and fate of ARG during biodegradation processes at the mechanistic level.

Phage tailspike modularity and horizontal gene transfer reveals specificity towards E. coli O-antigen serogroups.

BACKGROUND: The interaction between bacteriophages and their hosts is intricate and highly specific. Receptor-binding proteins (RBPs) of phages such as tail fibers and tailspikes initiate the infection process. These RBPs bind to diverse outer membrane structures, including the O-antigen, which is a serogroup-specific sugar-based component of the outer lipopolysaccharide layer of Gram-negative bacteria. Among the most virulent Escherichia coli strains is the Shiga toxin-producing E. coli (STEC) pathotype dominated by a subset of O-antigen serogroups. METHODS: Extensive phylogenetic and structural analyses were used to identify and validate specificity correlations between phage RBP subtypes and STEC O-antigen serogroups, relying on the principle of horizontal gene transfer as main driver for RBP evolution. RESULTS: We identified O-antigen specific RBP subtypes for seven out of nine most prevalent STEC serogroups (O26, O45, O103, O104, O111, O145 and O157) and seven additional E. coli serogroups (O2, O8, O16, O18, 4s/O22, O77 and O78). Eight phage genera (Gamaleya-, Justusliebig-, Kaguna-, Kayfuna-, Kutter-, Lederberg-, Nouzilly- and Uetakeviruses) emerged for their high proportion of serogroup-specific RBPs. Additionally, we reveal sequence motifs in the RBP region, potentially serving as recombination hotspots between lytic phages. CONCLUSION: The results contribute to a better understanding of mosaicism of phage RBPs, but also demonstrate a method to identify and validate new RBP subtypes for current and future emerging serogroups.

Analysis of genetic structure and function of clustered regularly interspaced short palindromic repeats loci in 110 Enterococcus strains.

Clustered regularly interspaced short palindromic repeats (CRISPR) and their CRISPR-associated proteins (Cas) are an adaptive immune system involved in specific defenses against the invasion of foreign mobile genetic elements, such as plasmids and phages. This study aims to analyze the gene structure and to explore the function of the CRISPR system in the Enterococcus genome, especially with regard to drug resistance. The whole genome information of 110 enterococci was downloaded from the NCBI database to analyze the distribution and the structure of the CRISPR-Cas system including the Cas gene, repeat sequences, and spacer sequence of the CRISPR-Cas system by bioinformatics methods, and to find drug resistance-related genes and analyze the relationship between them and the CRISPR-Cas system. Multilocus sequence typing (MLST) of enterococci was performed against the reference MLST database. Information on the drug resistance of Enterococcus was retrieved from the CARD database, and its relationship to the presence or absence of CRISPR was statistically analyzed. Among the 110 Enterococcus strains, 39 strains (35.45%) contained a complete CRISPR-Cas system, 87 CRISPR arrays were identified, and 62 strains contained Cas gene clusters. The CRISPR system in the Enterococcus genome was mainly type II-A (59.68%), followed by type II-C (33.87%). The phylogenetic analysis of the cas1 gene sequence was basically consistent with the typing of the CRISPR-Cas system. Of the 74 strains included in the study for MLST typing, only 19 (25.68%) were related to CRISPR-Cas typing, while the majority of the strains (74.32%) of MLST typing were associated with the untyped CRISPR system. Additionally, the CRISPR-Cas system may only be related to the carrying rate of some drug-resistant genes and the drug-resistant phenotype. In conclusion, the distribution of the enterococcus CRISPR-Cas system varies greatly among different species and the presence of CRISPR loci reduces the horizontal transfer of some drug resistance genes.

Assembly and analytical validation of a metagenomic reference catalog of human gut microbiota based on co-barcoding sequencing.

Human gut microbiota is associated with human health and disease, and is known to have the second-largest genome in the human body. The microbiota genome is important for their functions and metabolites; however, accurate genomic access to the microbiota of the human gut is hindered due to the difficulty of cultivating and the shortcomings of sequencing technology. Therefore, we applied the stLFR library construction method to assemble the microbiota genomes and demonstrated that assembly property outperformed standard metagenome sequencing. Using the assembled genomes as references, SNP, INDEL, and HGT gene analyses were performed. The results demonstrated significant differences in the number of SNPs and INDELs among different individuals. The individual displayed a unique species variation spectrum, and the similarity of strains within individuals decreased over time. In addition, the coverage depth analysis of the stLFR method shows that a sequencing depth of 60X is sufficient for SNP calling. HGT analysis revealed that the genes involved in replication, recombination and repair, mobilome prophages, and transposons were the most transferred genes among different bacterial species in individuals. A preliminary framework for human gut microbiome studies was established using the stLFR library construction method.

The dynamics and transmission of antibiotic resistance associated with plant microbiomes.

Antibiotic resistance genes (ARGs) have been widely found and studied in soil and water environments. However, the propagation of ARGs in plant microbiomes has attracted insufficient attention. Plant microbiomes, especially the rhizosphere microorganisms, are closely connected with water, soil, and air, which allows ARGs to spread widely in ecosystems and pose a threat to human health after entering the human body with bacteria. Therefore, it is necessary to deeply understand and explore the dynamics and the transmission of ARGs in rhizosphere microorganisms and endophytes of plants. In this review, the transmission and influencing factors of ARGs in the microorganisms associated with plants, especially the influence of root exudates on plant microbiomes, are analyzed. Notably, the role of intrinsic genes of plants in determining root exudates and their potential effects on ARGs are proposed and analyzed. The important role of phyllosphere microorganisms and endophytes in the transmission of ARGs and co-resistance of antibiotics and other substances are also emphasized. The proliferation and transmission of ARGs associated with plant microbiomes addressed in this review is conducive to revealing the fate of ARGs in plant microorganisms and alleviating ARG pollution.

A comprehensive genomic analysis provides insights on the high environmental adaptability of Acinetobacter strains.

Acinetobacter is ubiquitous, and it has a high species diversity and a complex evolutionary pattern. To elucidate the mechanism of its high ability to adapt to various environment, 312 genomes of Acinetobacter strains were analyzed using the phylogenomic and comparative genomics methods. It was revealed that the Acinetobacter genus has an open pan-genome and strong genome plasticity. The pan-genome consists of 47,500 genes, with 818 shared by all the genomes of Acinetobacter, while 22,291 are unique genes. Although Acinetobacter strains do not have a complete glycolytic pathway to directly utilize glucose as carbon source, most of them harbored the n-alkane-degrading genes alkB/alkM (97.1% of tested strains) and almA (96.7% of tested strains), which were responsible for medium-and long-chain n-alkane terminal oxidation reaction, respectively. Most Acinetobacter strains also have catA (93.3% of tested strains) and benAB (92.0% of tested strains) genes that can degrade the aromatic compounds catechol and benzoic acid, respectively. These abilities enable the Acinetobacter strains to easily obtain carbon and energy sources from their environment for survival. The Acinetobacter strains can manage osmotic pressure by accumulating potassium and compatible solutes, including betaine, mannitol, trehalose, glutamic acid, and proline. They respond to oxidative stress by synthesizing superoxide dismutase, catalase, disulfide isomerase, and methionine sulfoxide reductase that repair the damage caused by reactive oxygen species. In addition, most Acinetobacter strains contain many efflux pump genes and resistance genes to manage antibiotic stress and can synthesize a variety of secondary metabolites, including arylpolyene, ß-lactone and siderophores among others, to adapt to their environment. These genes enable Acinetobacter strains to survive extreme stresses. The genome of each Acinetobacter strain contained different numbers of prophages (0-12) and genomic islands (GIs) (6-70), and genes related to antibiotic resistance were found in the GIs. The phylogenetic analysis revealed that the alkM and almA genes have a similar evolutionary position with the core genome, indicating that they may have been acquired by vertical gene transfer from their ancestor, while catA, benA, benB and the antibiotic resistance genes could have been acquired by horizontal gene transfer from the other organisms.

Preceding Host History of Conjugative Resistance Plasmids Affects Intra- and Interspecific Transfer Potential from Biofilm.

Conjugative plasmids can confer antimicrobial resistance (AMR) to their host bacterium. The plasmids disperse even between distantly related host species, rescuing the host from otherwise detrimental effects of antibiotics. Little is known about the role of these plasmids in the spread of AMR during antibiotic treatment. One unstudied question is whether the past evolutionary history of a plasmid in a particular species creates host specificity in its rescue potential or if interspecific coevolution can improve interspecific rescues. To study this, we coevolved the plasmid RP4 under three different host settings; solely Escherichia coli or Klebsiella pneumoniae, or alternating between both of them. The ability of evolved plasmids in bacterial biofilm to rescue susceptible planktonic host bacteria of either the same or different species during beta-lactam treatment was tested. The interspecific coevolution seemed to decrease rescue potential for the RP4 plasmid, while the K. pneumoniae evolved plasmid became more host specific. Large deletion in the region encoding the mating pair formation (Tra2) apparatus was detected in the plasmids evolved with K. pneumoniae. This adaptation resulted in the exapted evolution of resistance against a plasmid-dependent bacteriophage PRD1. Further, previous studies have suggested that mutations in this region completely abolish the plasmid's ability to conjugate; however, our study shows it is not essential for conjugation but rather affects the host-specific conjugation efficiency. Overall, the results suggest that previous evolutionary history can result in the separation of host-specific plasmid lineages that may be further amplified by unselected exaptations such as phage resistance. IMPORTANCE Antimicrobial resistance (AMR) is a major global public health threat which can rapidly spread in microbial communities via conjugative plasmids. Here, we advance with evolutionary rescue via conjugation in a more natural setting, namely, biofilm, and incorporate a broad-host range plasmid RP4 to test whether intra- and interspecific host histories affect its transfer potential. Escherichia coli and Klebsiella pneumoniae hosts were seen to elicit different evolutionary influences on the RP4 plasmid, leading to clear differences in the rescue potential and underlining the significant role of the plasmid-host interactions in the spread of AMR. We also contradicted previous reports that established certain conjugal transfer genes of RP4 as essential. This work enhances the understanding of how plasmid host range evolve in different host settings and further, the potential effects it may have on the horizontal spread of AMR in complex environments such as biofilms.

An insight overview on COVID-19 mRNA vaccines: Advantageous, pharmacology, mechanism of action, and prospective considerations.

The worldwide spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has urged scientists to present some novel vaccine platforms during this pandemic to provide a rather prolonged immunity against this respiratory viral infection. In spite of many campaigns formed against the administration of mRNA-based vaccines, those platforms were the most novel types, which helped us meet the global demand by developing protection against COVID-19 and reducing the development of severe forms of this respiratory viral infection. Some societies are worry about the COVID-19 mRNA vaccine administration and the potential risk of genetic integration of inoculated mRNA into the human genome. Although the efficacy and long-term safety of mRNA vaccines have not yet been fully clarified, obviously their application has switched the mortality and morbidity of the COVID-19 pandemic. This study describes the structural features and technologies used in producing of COVID-19 mRNA-based vaccines as the most influential factor in controlling this pandemic and a successful pattern for planning to produce other kind of genetic vaccines against infections or cancers.

Uncovering Bacterial Hosts of Class 1 Integrons in an Urban Coastal Aquatic Environment with a Single-Cell Fusion-Polymerase Chain Reaction Technology.

Horizontal gene transfer (HGT) is a key driver of bacterial evolution via transmission of genetic materials across taxa. Class 1 integrons are genetic elements that correlate strongly with anthropogenic pollution and contribute to the spread of antimicrobial resistance (AMR) genes via HGT. Despite their significance to human health, there is a shortage of robust, culture-free surveillance technologies for identifying uncultivated environmental taxa that harbor class 1 integrons. We developed a modified version of epicPCR (emulsion, paired isolation, and concatenation polymerase chain reaction (PCR)) that links class 1 integrons amplified from single bacterial cells to taxonomic markers from the same cells in emulsified aqueous droplets. Using this single-cell genomic approach and Nanopore sequencing, we successfully assigned class 1 integron gene cassette arrays containing mostly AMR genes to their hosts in coastal water samples that were affected by pollution. Our work presents the first application of epicPCR for targeting variable, multigene loci of interest. We also identified the Rhizobacter genus as novel hosts of class 1 integrons. These findings establish epicPCR as a powerful tool for linking taxa to class 1 integrons in environmental bacterial communities and offer the potential to direct mitigation efforts toward hotspots of class 1 integron-mediated dissemination of AMR.

Comparative genome identification of accessory genes associated with strong biofilm formation in Vibrio parahaemolyticus.

Vibrio parahaemolyticus biofilms on the seafood processing plant surfaces are a potential source of seafood contamination and subsequent food poisoning. Strains differ in their ability to form biofilm, but little is known about the genetic characteristics responsible for biofilm development. In this study, pangenome and comparative genome analysis of V. parahaemolyticus strains reveals genetic attributes and gene repertoire that contribute to robust biofilm formation. The study identified 136 accessory genes that were exclusively present in strong biofilm forming strains and these were functionally assigned to the Gene Ontology (GO) pathways of cellulose biosynthesis, rhamnose metabolic and catabolic processes, UDP-glucose processes and O antigen biosynthesis (p < 0.05). Strategies of CRISPR-Cas defence and MSHA pilus-led attachment were implicated via Kyoto Encyclopedia of Genes and Genomes (KEGG) annotation. Higher levels of horizontal gene transfer (HGT) were inferred to confer more putatively novel properties on biofilm-forming V. parahaemolyticus. Furthermore, cellulose biosynthesis, a neglected potential virulence factor, was identified as being acquired from within the order Vibrionales. The cellulose synthase operons in V. parahaemolyticus were examined for their prevalence (22/138, 15.94 %) and were found to consist of the genes bcsG, bcsE, bcsQ, bcsA, bcsB, bcsZ, bcsC. This study provides insights into robust biofilm formation of V. parahaemolyticus at the genomic level and facilitates: identification of key attributes for robust biofilm formation, elucidation of biofilm formation mechanisms and development of potential targets for novel control strategies of persistent V. parahaemolyticus.

Antibiotic and heavy metal resistance genes in sewage sludge survive during aerobic composting.

Municipal sewage sludge has been generated in increasing amounts with the acceleration of urbanization and economic development. The nutrient rich sewage sludge can be recycled by composting that has a great potential to produce stabilized organic fertilizer and substrate for plant cultivation. However, little is known about the metals, pathogens and antibiotic resistance transfer risks involved in applying the composted sludge in agriculture. We studied changes in and relationships between heavy metal contents, microbial communities, and antibiotic resistance genes (ARGs), heavy metal resistance genes (HMRGs) and mobile genetic elements (MGEs) in aerobic composting of sewage sludge. The contents of most of the analyzed heavy metals were not lower after composting. The bacterial α-diversity was lower, and the community composition was different after composting. Firmicutes were enriched, and Proteobacteria and potential pathogens in the genera Arcobacter and Acinetobacter were depleted in the composted sludge. The differences in bacteria were possibly due to the high temperature phase during the composting which was likely to affect temperature-sensitive bacteria. The number of detected ARGs, HMRGs and MGEs was lower, and the relative abundances of several resistance genes were lower after composting. However, the abundance of seven ARGs and six HMRGs remained on the same level after composting. Co-occurrence analysis of bacterial taxa and the genes suggested that the ARGs may spread via horizontal gene transfer during composting. In summary, even though aerobic composting is effective for managing sewage sludge and to decrease the relative abundance of potential pathogens, ARGs and HMRGs, it might include a potential risk for the dissemination of ARGs in the environment.

Evolutionary Overview of Aquaporin Superfamily.

Aquaporins (AQPs) are present not only in three domains of life, bacteria, eukaryotes, and archaea, but also in viruses. With the accumulating arrays of AQP superfamily, the evolutional relationship has attracted much attention with multiple publications on "the genome-wide identification and phylogenetic analysis" of AQP superfamily. A pair of NPA boxes forming a pore is highly conserved throughout the evolution and renders key residues for the classification of AQP superfamily into four groups: AQP1-like, AQP3-like, AQP8-like, and AQP11-like. The complexity of AQP family has mostly been achieved in nematodes and subsequent evolution has been directed toward increasing the number of AQPs through whole-genome duplications (WGDs) to extend the tissue specific expression and regulation. The discovery of the intracellular AQP (iAQP: AQP8-like and AQP11-like) and substrate transports by the plasma membrane AQP (pAQP: AQP1-like and AQP3-like) have accelerated the AQP research much more toward the transport of substrates with complex profiles. This evolutionary overview based on a simple classification of AQPs into four subfamilies will provide putative structural, functional, and localization information and insights into the role of AQP as well as clues to understand the complex diversity of AQP superfamily.

Plant cultivar determined bacterial community and potential risk of antibiotic resistance gene spread in the phyllosphere.

The global increased antibiotic resistance level in pathogenic microbes has posed a significant threat to human health. Fresh vegetables have been recognized to be an important vehicle of antibiotic resistance genes (ARGs) from environments to human beings. Phyllosphere ARGs have been indicated to be changed with plant species, yet the influence of plant cultivar on the phyllospheric resistome is still unclear. Here, we detected the ARGs and bacterial communities in the phyllosphere of two cultivars of cilantros and their corresponding soils using high-throughput quantitative PCR technique and bacterial 16S rRNA gene-based high-throughput sequencing, respectively. We further identified the potential bacterial pathogens and analyzed the effects of plant cultivar on ARGs, mobile genetic elements (MGEs), microbiome and potential bacterial pathogens. The results showed that the cultivars did not affect the ARG abundance and composition, but significantly shaped the abundance of MGEs and the composition structure of bacteria in the phyllosphere. The relative abundance of potential bacterial pathogens was significantly higher in the phyllosphere than that in soils. Mantel test showed that the ARG patterns were significantly correlated to the patterns of potential bacterial pathogens. Our results suggested that the horizontal gene transfer of ARGs in the phyllosphere might be different between the two cultivars of cilantro and highlighted the higher risk of phyllospheric microorganisms compared with those in soils. These findings extend our knowledge on the vegetable microbiomes, ARGs, and potential pathogens, suggesting more agricultural and hygiene protocols are needed to control the risk of foodborne ARGs.

Genomic and functional analysis of phage-mediated horizontal gene transfer in Pseudomonas syringae on the plant surface.

Many strains of Pseudomonas colonise plant surfaces, including the cherry canker pathogens, Pseudomonas syringae pathovars syringae and morsprunorum. We have examined the genomic diversity of P. syringae in the cherry phyllosphere and focused on the role of prophages in transfer of genes encoding Type 3 secreted effector (T3SE) proteins contributing to the evolution of virulence. Phylogenomic analysis was carried out on epiphytic pseudomonads in the UK orchards. Significant differences in epiphytic populations occurred between regions. Nonpathogenic strains were found to contain reservoirs of T3SE genes. Members of P. syringae phylogroups 4 and 10 were identified for the first time from Prunus. Using bioinformatics, we explored the presence of the gene encoding T3SE HopAR1 within related prophage sequences in diverse P. syringae strains including cherry epiphytes and pathogens. Results indicated that horizontal gene transfer (HGT) of this effector between phylogroups may have involved phage. Prophages containing hopAR1 were demonstrated to excise, circularise and transfer the gene on the leaf surface. The phyllosphere provides a dynamic environment for prophage-mediated gene exchange and the potential for the emergence of new more virulent pathotypes. Our results suggest that genome-based epidemiological surveillance of environmental populations will allow the timely application of control measures to prevent damaging diseases.