Integrative and conjugative elements of Pasteurella multocida: Prevalence and signatures in population evolution.

Pasteurella multocida (P. multocida) is a bacterial pathogen responsible for a range of infections in humans and various animal hosts, causing significant economic losses in farming. Integrative and conjugative elements (ICEs) are important horizontal gene transfer elements, potentially enabling host bacteria to enhance adaptability by acquiring multiple functional genes. However, the understanding of ICEs in P. multocida and their impact on the transmission of this pathogen remains limited. In this study, 42 poultry-sourced P. multocida genomes obtained by high-throughput sequencing together with 393 publicly available P. multocida genomes were used to analyse the horizontal transfer of ICEs. Eighty-two ICEs were identified in P. multocida, including SXT/R391 and Tn916 subtypes, as well as three subtypes of ICEHin1056 family, with the latter being widely prevalent in P. multocida and carrying multiple resistance genes. The correlations between insertion sequences and resistant genes in ICEs were also identified, and some ICEs introduced the carbapenem gene blaOXA-2 and the bleomycin gene bleO to P. multocida. Phylogenetic and collinearity analyses of these bioinformatics found that ICEs in P. multocida were transmitted vertically and horizontally and have evolved with host specialization. These findings provide insight into the transmission and evolution mode of ICEs in P. multocida and highlight the importance of understanding these elements for controlling the spread of antibiotic resistance.

RATA: A novel class A carbapenemase with broad geographic distribution and potential for global spread.

Carbapenem resistance's global proliferation poses a significant public health challenge. The primary resistance mechanism is carbapenemase production. In this study, we discovered a novel carbapenemase, RATA, located on the chromosome of Riemerella anatipestifer isolates. This enzyme shares ≤52 % amino acid sequence identity with other known ß-lactamases. Antimicrobial susceptibility tests and kinetic assays demonstrated that RATA could hydrolyze not only penicillins and extended-spectrum cephalosporins but also monobactams, cephamycins, and carbapenems. Furthermore, its activity was readily inhibited by ß-lactamase inhibitors. Bioinformatic analysis revealed 46 blaRATA-like genes encoding 27 variants in the NCBI database, involving 21 different species, including pathogens, host-associated bacteria, and environmental isolates. Notably, blaRATA-positive strains were globally distributed and primarily collected from marine environments. Concurrently, taxonomic analysis and GC content analysis indicated that blaRATA orthologue genes were predominantly located on the chromosomes of Flavobacteriaceae and shared a similar GC content as Flavobacteriaceae. Although no explicit mobile genetic elements were identified by genetic environment analysis, blaRATA-2 possessed the ability of horizontal transfer in R. anatipestifer via natural transformation. This work's data suggest that RATA is a new chromosome-encoded class A carbapenemase, and Flavobacteriaceae from marine environments could be the primary reservoir of the blaRATA gene.

Genome-based assessment of antimicrobial resistance reveals the lineage specificity of resistance and resistance gene profiles in Riemerella anatipestifer from China.

Riemerella anatipestifer (R. anatipestifer) is an important pathogen that causes severe systemic infections in domestic ducks, resulting in substantial economic losses for China's waterfowl industry. Controlling R. anatipestifer with antibiotics is extremely challenging due to its multidrug resistance. Notably, large-scale studies on antimicrobial resistance (AMR) and the corresponding genetic determinants in R. anatipestifer remain scarce. To solve this dilemma, more than 400 nonredundant R. anatipestifer isolates collected from 22 provinces in China between 1994 and 2021 were subjected to broth dilution antibiotic susceptibility assays, and their resistance-associated genetic determinants were characterized by whole-genome sequencing. While over 90% of the isolates was resistant to sulfamethoxazole, kanamycin, gentamicin, ofloxacin, norfloxacin, and trimethoprim, 88.48% of the isolates was resistant to the last-resort drug (tigecycline). Notably, R. anatipestifer resistance to oxacillin, norfloxacin, ofloxacin, and tetracycline was found to increase relatively over time. Genome-wide analysis revealed the alarmingly high prevalence of blaOXA-like (93.05%) and tet(X) (90.64%) genes and the uneven distribution of resistance genes among lineages. Overall, this study reveals a serious AMR situation regarding R. anatipestifer in China, with a high prevalence and high diversity of antimicrobial resistance genes, providing important data for the rational use of antibiotics in veterinary practice.IMPORTANCERiemerella anatipestifer (R. anatipestifer), an important waterfowl pathogen, has caused substantial economic losses worldwide, especially in China. Antimicrobial resistance (AMR) is a major challenge in controlling this pathogen. Although a few studies have reported antimicrobial resistance in R. anatipestifer, comprehensive data remain a gap. This study aims to address the lack of information on R. anatipestifer AMR and its genetic basis. By analyzing more than 400 isolates collected over two decades, this study reveals alarming levels of resistance to several antibiotics, including drugs of last resort. The study also revealed the lineage-specificity of resistance profiles and resistance gene profiles. Overall, this study provides new insights and updated data support for understanding AMR and its genetic determinants in R. anatipestifer.

Genome-wide association study reveals serovar-associated genetic loci in Riemerella anatipestifer.

BACKGROUND: The disease caused by Riemerella anatipestifer (R. anatipestifer, RA) results in large economic losses to the global duck industry every year. Serovar-related genomic variation, such as the O-antigen and capsular polysaccharide (CPS) gene clusters, has been widely used for serotyping in many gram-negative bacteria. RA has been classified into at least 21 serovars based on slide agglutination, but the molecular basis of serotyping is unknown. In this study, we performed a pan-genome-wide association study (Pan-GWAS) to identify the genetic loci associated with RA serovars. RESULTS: The results revealed a significant association between the putative CPS synthesis gene locus and the serological phenotype. Further characterization of the CPS gene clusters in 11 representative serovar strains indicated that they were highly diverse and serovar-specific. The CPS gene cluster contained the key genes wzx and wzy, which are involved in the Wzx/Wzy-dependent pathway of CPS synthesis. Similar CPS loci have been found in some other species within the family Weeksellaceae. We have also shown that deletion of the wzy gene in RA results in capsular defects and cross-agglutination. CONCLUSIONS: This study indicates that the CPS synthesis gene cluster of R. anatipestifer is a serotype-specific genetic locus. Importantly, our finding provides a new perspective for the systematic analysis of the genetic basis of the R anatipestifer serovars and a potential target for establishing a complete molecular serotyping scheme.

Capsular polysaccharide determines the serotyping of Riemerella anatipestifer.

IMPORTANCE: Riemerella anatipestifer (R. anatipestifer) is one of the most important veterinary pathogens with at least 21 serotypes. However, the exact polysaccharide(s) that determine R. anatipestifer serotype is still unknown. This study has provided a preliminary exploration of the relationship between capsular polysaccharides and serotyping in R. anatipestifer and suggests possible directions for further investigation of the genetic basis of serotypes in this bacterium.

Identification of a novel carbapenem-hydrolysing class D ß-lactamase RAD-1 in Riemerella anatipestifer.

OBJECTIVES: To elucidate the role of a novel carbapenem-hydrolysing class D ß-lactamase (RAD-1) from Riemerella anatipestifer. METHODS: We applied WGS and bioinformatic analysis to screen putative ß-lactamase genes in R. anatipestifer SCVM0004. A putative class D ß-lactamase gene was cloned into pET24a and transferred into Escherichia coli BL21 (DE3) for antibiotic susceptibility determination and protein purification. Meanwhile, the purified native protein was used to determine the enzymatic activities. RESULTS: A class D ß-lactamase, RAD-1, was identified in the genome of R. anatipestifer SCVM0004. It was distinct from all characterized class D ß-lactamases (≤42% amino acid sequence identity). Searching in GenBank showed that blaRAD-1 was widely disseminated among R. anatipestifer. Genomic environment analysis indicated that the chromosomal structures of blaRAD-1-located regions were relatively conserved. Expression of RAD-1 in E. coli results in elevated MICs for various ß-lactam antibiotics, including penicillins, extended-spectrum cephalosporins, a monobactam and carbapenems. Moreover, kinetic analysis of purified RAD-1 revealed: (i) high-level activity against penicillins; (ii) highest affinity for carbapenems; (iii) moderate hydrolysis of extended-spectrum cephalosporins and a monobactam; and (iv) no activity for oxacillin and cefoxitin. CONCLUSIONS: This study identified a novel chromosomally located class D carbapenemase RAD-1 (Bush-Jacoby functional group 2def) in R. anatipestifer SCVM0004. Moreover, bioinformatic analysis confirmed that the RAD-1 was widely prevalent and conserved in R. anatipestifer.

EBR-5, a Novel Variant of Metallo-ß-Lactamase EBR from Multidrug-Resistant Empedobacter stercoris.

A novel chromosome-encoded metallo-ß-lactamase (MBL) EBR variant, namely, EBR-5, was identified in a multidrug-resistant Empedobacter stercoris strain SCVM0123 that was isolated from chicken anal swab samples. EBR-5 shared 82.13% amino acid identity with the previously known EBR-1. The expression of EBR-5 in Escherichia coli reduced susceptibility to expanded-spectrum cephalosporins and carbapenems. Compared with blaEBR-1, the recombinant strain harboring blaEBR-5 exhibited higher minimum inhibitory concentrations of piperacillin, cefotaxime, and meropenem. Despite the genetic diversity, EBR-5 and EBR-1 possessed similar kinetic parameters, except for cefepime, cefotaxime, cefoxitin, cephalothin, and meropenem, which were hydrolyzed more by EBR-5. In addition to blaEBR-1, a whole-genome sequencing analysis of SCVM0123 also revealed a plasmid-mediated blaRAA-1 gene. This study underlines the importance of E. stercoris monitoring, as it could be a potential reservoir of these ß-lactamase genes. IMPORTANCE Carbapenemases are one of the greatest threats to clinical therapy, as they could confer resistance by hydrolyzing carbapenems and other ß-lactam antimicrobials. In this study, we identified a novel metallo-ß-lactamase EBR variant, namely, EBR-5, in Empedobacter stercoris. The biochemical properties, substrate hydrolysis abilities, and inhibition profiles of EBR-5 were reported. Through whole-genome sequencing and bioinformatic analyses, we revealed for the first time that the ESBL gene blaRAA-1 was located on a plasmid. This study extends the database of class B metallo-ß-lactamases. Meanwhile, E. stercoris could be a major reservoir of blaEBR-5 and blaRAA-1, which have potential to spread to pathogens.

Phenotypic and genotypic characterization of antimicrobial resistance profiles in Salmonella isolated from waterfowl in 2002-2005 and 2018-2020 in Sichuan, China.

Salmonella enterica is a widespread foodborne pathogen with concerning antimicrobial resistance (AMR). Waterfowl are a major source of Salmonella transmission, but there are few systematic studies on Salmonella prevalence in waterfowl species. In this study, 126 Salmonella isolates (65 collected in 2018-2020 and 61 collected in 2002-2005) were obtained from waterfowl samples in Sichuan, China. Their serotypes, pulsed-field gel electrophoresis (PFGE) types, and phenotypic and genotypic AMR profiles were systematically examined. The isolates were distributed in 7 serotypes, including serovars Enteritidis (46.0%), Potsdam (27.8%), Montevideo (7.9%), Cerro (6.3%), Typhimurium (4.8%), Kottbus (4.0%) and Apeyeme (3.2%). Their PFGE characteristics were diverse; all isolates were distributed in four groups (cutoff value: 60.0%) and 20 clusters (cutoff value: 80.0%). Moreover, all isolates were multidrug resistant, and high rates of AMR to lincomycin (100.0%), rifampicin (100.0%), sulfadiazine (93.7%), erythromycin (89.7%), ciprofloxacin (81.0%), and gentamicin (75.4%) were observed. Finally, 49 isolates were subjected to whole-genome sequencing, and a wide variety of AMR genes were found, including multiple efflux pump genes and specific resistance genes. Interestingly, the tet(A)/tet(B) and catII resistance genes were detected in only isolates obtained in the first collection period, while the gyrA (S83F, D87N and D87G) and gyrB (E466D) mutations were detected at higher frequencies in the isolates obtained in the second collection period, supporting the findings that isolates from different periods exhibited different patterns of resistance to tetracycline, chloramphenicol and nalidixic acid. In addition, various incompatible plasmid replicon fragments were detected, including Col440I, Col440II, IncFIB, IncFII, IncX1, IncX9, IncI1-I and IncI2, which may contribute to the horizontal transmission of AMR genes and provide competitive advantages. In summary, we demonstrated that the Salmonella isolates prevalent in Sichuan waterfowl farms exhibited diverse serotypes, multiple AMR phenotypes and genotypes, and AMR changes over time, indicating their potential risks to public health.

Emergence of plasmid-mediated tigecycline, ß-lactam and florfenicol resistance genes tet(X), blaOXA-347 and floR in Riemerella anatipestifer isolated in China.

Bacterial antimicrobial resistance (AMR) continues to develop, with the horizontal transfer of antibiotic resistance genes (ARGs) through plasmids playing a major role. Recently, the antimicrobial resistance of R. anatipestifer has become increasingly severe, jeopardizing the development of the poultry industry. In this study, we used PromethION to determine the whole genome sequence of R. anatipestifer RCAD0416, a multidrug-resistant isolate from China. We detected a plasmid in the isolate. We named the plasmid pRCAD0416RA-1; the plasmid was 37356 bp in size with 36 putative open reading frames and included the blaOXA-347, floR, tet(X), ermF, ereD, and AadS resistance genes. Most resistance genes might be obtained from R. anatipestifer HXb2. Mobile elements and floR might be transmitted by plasmid pB18-2 from Acinetobacter indicus, and the ICEPg6Chn1 mobile elements can be transmitted from Proteus genomosp. The plasmid pRCAD0416RA-1 was transferred to Escherichia coli K-12 × 7232 via electroporation. Subsequent antimicrobial sensitivity tests (AST) showed a noticeable levels of antimicrobial resistance to ß-lactams (4-8 fold), tigecycline (8 fold), and florfenicol (8 fold). These types of antibiotics are in common clinical use. The purpose of this article is to elucidate the basic characteristics of pRCAD0416RA-1 and the level of resistance mediated by blaOXA-347, floR, and tet(X).

Characterization of RASA-1, a novel class A extended-spectrum beta-lactamase from Riemerella anatipestifer.

A novel chromosomally-located ß-lactamase gene, blaRASA-1, was identified in Riemerella anatipestifer RA-CH-1. The RASA-1, encoded by blaRASA-1, was a class A extended-spectrum ß-lactamase (ESBL), which shared 42.7% and 40.5% identities with the RAA-1 and CGA-1 ß-lactamase, respectively. Overexpression of RASA-1 in Escherichia coli confers broad resistance to ß-lactams and the purified native RASA-1 revealed ESBL-like hydrolysis activity. Blasting in GenBank showed that blaRASA-1 was exclusively detected in Riemerella anatipestifer. Moreover, sequence analysis revealed that this gene was located within the multi-resistance region of Riemerella anatipestifer genome.

Tracing genetic signatures of bat-to-human coronaviruses and early transmission of North American SARS-CoV-2.

Highly pathogenic coronaviruses, including SARS-CoV-2, SARS-CoV and MERS-CoV, are thought to be transmitted from bats to humans, but the viral genetic signatures that contribute to bat-to-human transmission remain largely obscure. In this study, we identified an identical ribosomal frameshift motif among the three bat-human pairs of viruses and strong purifying selection after jumping from bats to humans. This represents genetic signatures of coronaviruses that are related to bat-to-human transmission. To further trace the early human-to-human transmission of SARS-CoV-2 in North America, a geographically stratified genome-wide association study (North American isolates and the remaining isolates) and a retrospective study were conducted. We determined that the single nucleotide polymorphisms (SNPs) 1,059.C > T and 25,563.G > T were significantly associated with approximately half of the North American SARS-CoV-2 isolates that accumulated largely during March 2020. Retrospectively tracing isolates with these two SNPs was used to reconstruct the early, reliable transmission history of North American SARS-CoV-2, and European isolates (February 26, 2020) showed transmission 3 days earlier than North American isolates and 17 days earlier than Asian isolates. Collectively, we identified the genetic signatures of the three pairs of coronaviruses and reconstructed an early transmission history of North American SARS-CoV-2. We envision that these genetic signatures are possibly diagnosable and predic markers for public health surveillance.

Distribution and association of antimicrobial resistance and virulence traits in Escherichia coli isolates from healthy waterfowls in Hainan, China.

There are rising concerns about microbes harboring antibiotic resistance genes (ARGs) and virulence-associated genes (VAGs) in humans and food-producing animals. Moreover, ARGs are considered as emerging environmental pollutants, posing probable life-threatening complications in humans and animals. Commensal Escherichia coli (E. coli) strain can carry a large number of VAGs, which may become opportunistic pathogen. The objective of this study was to determine the prevalence and possible association of ARGs and VAGs in E. coli isolates from clinically healthy waterfowls in China's tropical island, Hainan. For this purpose, 311 non-repeating E. coli isolates were evaluated for phenotypic drug resistance linked with ARGs. Additionally, strains were examined for subsequent resistance and virulence genes by uniplex or multiplex PCR and sequencing. Overall, 89 types of antibiotic resistance patterns were analysed, while 25 ARGs and 23 VAGs were observed, of which qnrS (99.4%) and iucD (99.7%) were the most commonly found genes, respectively. Significant positive associations were observed among ARGs and VAGs (p<0.05, OR>1). The strongest association between resistance and virulence gene was observed for qnrS and iss (OR, 76.25; 95% CI, 4.02-1445.42). Our results propose that waterfowls serve as a reservoir of E. coli carrying multi ARGs and various ExPEC associated VAGs. Therefore, this study provides necessary information on the occurrence and possible associations of ARGs and VAGs in healthy waterfowls, which may act as a reference for the regulatory use of antibiotics to stop the direct or indirect spread of these resistant and potential virulent microbes to natural environment.

An Exposed Outer Membrane Hemin-Binding Protein Facilitates Hemin Transport by a TonB-Dependent Receptor in Riemerella anatipestifer.

Iron is an essential element for the replication of most bacteria, including Riemerella anatipestifer, a Gram-negative bacterial pathogen of ducks and other birds. R. anatipestifer utilizes hemoglobin-derived hemin as an iron source; however, the mechanism by which this bacterium acquires hemin from hemoglobin is largely unknown. Here, rhuA disruption was shown to impair iron utilization from duck hemoglobin in R. anatipestifer CH-1. Moreover, the putative lipoprotein RhuA was identified as a surface-exposed, outer membrane hemin-binding protein, but it could not extract hemin from duck hemoglobin. Mutagenesis studies showed that recombinant RhuAY144A, RhuAY177A, and RhuAH149A lost hemin-binding ability, suggesting that amino acid sites at tyrosine 144 (Y144), Y177, and histidine 149 (H149) are crucial for hemin binding. Furthermore, rhuR, the gene adjacent to rhuA, encodes a TonB2-dependent hemin transporter. The function of rhuA in duck hemoglobin utilization was abolished in the rhuR mutant strain, and recombinant RhuA was able to bind the cell surface of R. anatipestifer CH-1 ΔrhuA rather than R. anatipestifer CH-1 ΔrhuR ΔrhuA, indicating that RhuA associates with RhuR to function. The sequence of the RhuR-RhuA hemin utilization locus exhibits no similarity to those of characterized hemin transport systems. Thus, this locus is a novel hemin uptake locus with homologues distributed mainly in the Bacteroidetes phylum. IMPORTANCE In vertebrates, hemin from hemoglobin is an important iron source for infectious bacteria. Many bacteria can obtain hemin from hemoglobin, but the mechanisms of hemin acquisition from hemoglobin differ among bacteria. Moreover, most studies have focused on the mechanism of hemin acquisition from mammalian hemoglobin. In this study, we found that the RhuR-RhuA locus of R. anatipestifer CH-1, a duck pathogen, is involved in hemin acquisition from duck hemoglobin via a unique pathway. RhuA was identified as an exposed outer membrane hemin-binding protein, and RhuR was identified as a TonB2-dependent hemin transporter. Moreover, the function of RhuA in hemoglobin utilization is RhuR dependent and not vice versa. The homologues of RhuR and RhuA are widely distributed in bacteria in marine environments, animals, and plants, representing a novel hemin transportation system of Gram-negative bacteria. This study not only was important for understanding hemin uptake in R. anatipestifer but also enriched the knowledge about the hemin transportation pathway in Gram-negative bacteria.

Evolutionarily missing and conserved tRNA genes in human and avian.

Viral infection heavily relies on host transfer RNA (tRNA) for viral RNA decoding. Counterintuitively, not all tRNA species based on anticodon are matched to all 64-triplet codons during evolution. Life solves this problem by cognate tRNA species via wobbling decoding. We found that 14 out of 64 tRNA genes in humans and the main avian species (chicken and duck) were parallelly missing, including 8 tRNA-A34NN and 6 tRNA-G34NN species. By analyzing the conservation of key motifs in tRNA genes, we found that box A and B served as intragenic tRNA promoters were evolutionally conserved among human, chicken, and duck. Thus, decoding viral RNA by similar wobbling strategies and tRNA transcripts may be parallelly used by human, chicken, and duck. We envisioned that many basic mechanisms regarding viral RNA decoding were possibly conserved in these hosts and may consequently promote cross-species infection. Transfer RNAs (tRNAs) are essentially required for gene decoding. Despite the universal nature of genetic codon, not all tRNA genes are common to all organisms. Here, we would like to discuss fundamental problems and possible effects arising from the evolutionarily missing and conserved tRNA genes in human, chicken, and duck (Alkatib et al., 2012; Ou et al., 2019; Rogalski et al., 2008). Among these three organisms, viruses especially the avian influenza virus can crossly infect (Pepin et al., 2010). For multi-host viruses, similar viral RNA decoding strategies may be parallelly used by different hosts. Because viral cross-species infection heavily relies on host tRNAs of different species for viral RNA decoding (Ou et al., 2020; van Weringh et al., 2011). We envisioned that many basic mechanisms regarding viral RNA decoding were possibly conserved in these three hosts and may consequently promote cross-species infection.

Emergence of Escherichia coli isolates producing NDM-1 carbapenemase from waterfowls in Hainan island, China.

Carbapenems are traditionally recognized to be the last resort drugs to treat infections due to MDR organisms such as E. coli. As such, the emergence of New Delhi metallo-ß-lactamase-producing E. coli strains have become a challenging threat to the public health. In this regard, we examined the molecular characteristics of carbapenem-resistant E. coli (CRE) isolated from waterfowls in China's tropical island, Hainan. A total of 311 single E. coli strains were obtained from 20 various farms of healthy ducks and geese in 2 districts of Hainan island. The CRE strains were initially identified via phenotypic resistance and modified Hodge test. PCR assay and subsequent nucleotide sequencing were used to detect different types of carbapenemase encoding genes (blaNDM, blaVIM, blaIMP, blaOXA and blaKPC). In addition, MLST and PFGE analyses were also performed. Among the 311 E. coli strains, 8 strains were detected to produce a single type of carbapenemase i.e. NDM-1 (2.6%). A total of 5 sequence types (STs) were observed, of which ST10 was the most prevalent accounting for 37.5% (3/8). Moreover, these 8 isolates yielded 6 different PFGE clusters but showed approximately related PFGE types, suggesting the propagation of similar clone between the farms. This is the first report on the identification of NDM-1-producing E. coli from waterfowls in Hainan island, China. Our results emphasize the need for better efforts to control the further spread of NDM-1-producing E. coli strains in this tropical island.

Pan-genome analysis of Riemerella anatipestifer reveals its genomic diversity and acquired antibiotic resistance associated with genomic islands.

Riemerella anatipestifer is a gram-negative bacterium that leads to severe contagious septicemia in ducks, turkeys, chickens, and wild waterfowl. Here, a pan-genome with 32 R. anatipestifer genomes is re-established, and the mathematical model is calculated to evaluate the expansion of R. anatipestifer genomes, which were determined to be open. Average nucleotide identity (ANI) and phylogenetic analysis preliminarily clarify intraspecies variation and distance. Comparative genomic analysis of R. anatipestifer found that horizontal gene transfer events, which provide an expressway for the recruitment of novel functionalities and facilitate genetic diversity in microbial genomes, play a key role in the process of acquiring and transmitting antibiotic-resistance genes in R. anatipestifer. Furthermore, a new antibiotic-resistance gene cluster was identified in the same loci in 14 genomes. The uneven distribution of virulence factors was also confirmed by our results. Our study suggests that the ability to acquire foreign genes (such as antibiotic-resistance genes) increases the adaptability of R. anatipestifer, and the virulence genes with little mobility are highly conserved in R. anatipestifer.

First Report of Integrative Conjugative Elements in Riemerella anatipestifer Isolates From Ducks in China.

We report for the first time the occurrence of integrative conjugative elements (ICEs) in Riemerella anatipestifer (R.anatipestifer) isolated from diseased ducks in China. For this purpose, a total of 48 genome sequences were investigated, which comprised 30 publicly available R. anatipestifer genome sequences, and 18 clinical isolates genomes sequences. Two ICEs, named ICERanRCAD0133-1 and ICERanRCAD0179-1 following the classic nomenclature system, were identified in R. anatipestifer through the use of bioinformatics tools. Comparative analysis revealed that three ICEs in Ornithobacterium rhinotracheale showed a high degree of conservation with the core genes of ICERanRCAD0133-1, while 13 ICEs with high similarity to ICERanRCAD0179-1 were found in Bacteroidetes. Based on the definition of ICE family, ICERanRCAD0179-1 was grouped in CTnDOT/ERL family; however, ICERanRCAD0133-1, which had no significant similarity with known ICEs, might be classified into a novel ICE family. The sequences of ICERanRCAD0133-1 and ICERanRCAD0179-1 were 70890 bp and 49166 bp in length, had 33.14 and 50.34% GC content, and contained 77 CDSs and 51 CDSs, respectively. Cargo genes carried by these two ICEs were predicted to encode: R-M systems, IS elements, a putative TonB-dependent receptor, a bacteriocin/lantibiotic efflux ABC transporter, a tetracycline resistance gene and more. In addition, phylogenetic analyses revealed that ICERanRCAD0179-1 and related ICEs were derived from a common ancestor, which may have undergone divergence prior to integartation into the host bacterial chromosome, and that the core genes co-evolved via a related evolutionary process or experienced only a low degree of recombination events during spread from a common CTnDOT/ERL family ancestor. Collectively, this study is the first identification and characterization of ICEs in R. anatipestifer; and provides new insights into the genetic diversity, evolution, adaptation, antimicrobial resistance, and virulence of R. anatipestifer.

Comparative analysis reveals the Genomic Islands in Pasteurella multocida population genetics: on Symbiosis and adaptability.

BACKGROUND: Pasteurella multocida (P. multocida) is a widespread opportunistic pathogen that infects human and various animals. Genomic Islands (GIs) are one of the most important mobile components that quickly help bacteria acquire large fragments of foreign genes. However, the effects of GIs on P. multocida are unknown in the evolution of bacterial populations. RESULTS: Ten avian-sourced P. multocida obtained through high-throughput sequencing together with 104 publicly available P. multocida genomes were used to analyse their population genetics, thus constructed a pan-genome containing 3948 protein-coding genes. Through the pan-genome, the open evolutionary pattern of P. multocida was revealed, and the functional components of 944 core genes, 2439 accessory genes and 565 unique genes were analysed. In addition, a total of 280 GIs were predicted in all strains. Combined with the pan-genome of P. multocida, the GIs accounted for 5.8% of the core genes in the pan-genome, mainly related to functional metabolic activities; the accessory genes accounted for 42.3%, mainly for the enrichment of adaptive genes; and the unique genes accounted for 35.4%, containing some defence mechanism-related genes. CONCLUSIONS: The effects of GIs on the population genetics of P. multocida evolution and adaptation to the environment are reflected by the proportion and function of the pan-genome acquired from GIs, and the large quantities of GI data will aid in additional population genetics studies.

Segmentation and Multi-Scale Convolutional Neural Network-Based Classification of Airborne Laser Scanner Data.

The classification of point clouds is a basic task in airborne laser scanning (ALS) point cloud processing. It is quite a challenge when facing complex observed scenes and irregular point distributions. In order to reduce the computational burden of the point-based classification method and improve the classification accuracy, we present a segmentation and multi-scale convolutional neural network-based classification method. Firstly, a three-step region-growing segmentation method was proposed to reduce both under-segmentation and over-segmentation. Then, a feature image generation method was used to transform the 3D neighborhood features of a point into a 2D image. Finally, feature images were treated as the input of a multi-scale convolutional neural network for training and testing tasks. In order to obtain performance comparisons with existing approaches, we evaluated our framework using the International Society for Photogrammetry and Remote Sensing Working Groups II/4 (ISPRS WG II/4) 3D labeling benchmark tests. The experiment result, which achieved 84.9% overall accuracy and 69.2% of average F1 scores, has a satisfactory performance over all participating approaches analyzed.