Integrative And Conjugative Elements Research Articles

Redefining the bacteriophage mv4 site-specific recombination system and the sequence specificity of its attB and core-attP sites.

Through their involvement in the integration and excision of a large number of mobile genetic elements, such as phages and integrative and conjugative elements (ICEs), site-specific recombination systems based on heterobivalent tyrosine recombinases play a major role in genome dynamics and evolution. However, despite hundreds of these systems having been identified in genome databases, very few have been described in detail, with none from phages that infect Bacillota (formerly Firmicutes). In this study, we reanalyzed the recombination module of Lactobacillus delbrueckii subsp. bulgaricus phage mv4, previously considered atypical compared with classical systems. Our results reveal that mv4 integrase is a 369 aa protein with all the structural hallmarks of recombinases from the Tn916 family and that it cooperatively interacts with its recombination sites. Using randomized DNA libraries, NGS sequencing, and other molecular approaches, we show that the 21-bp core-attP and attB sites have structural similarities to classical systems only if considering the nucleotide degeneracy, with two 7-bp inverted regions corresponding to mv4Int core-binding sites surrounding a 7-bp strand-exchange region. We also examined the different compositional constraints in the core-binding regions, which define the sequence space of permissible recombination sites.

Isolation, Identification, Genomic Diversity, and Antimicrobial Resistance Analysis of Streptococcus suis in Hubei Province of China from 2021 to 2023

Streptococcus suis (S. suis) is a zoonotic pathogen capable of causing severe diseases in humans and pigs, including meningitis, sepsis, polyserositis, arthritis, and endocarditis. This study aimed to investigate the biological characteristics of 19 strains of S. suis isolated from diseased pigs in Hubei Province between 2021 and 2023. Through bioinformatics analysis, we investigated the serotype, MLST, pan-genome characteristics, SNP, AMR, and ICE of the 19 S. suis isolates. Among the 19 S. suis strains, ten serotypes were identified, and serotype 9 was the most prevalent (21.05%). Ten new alleles and nine new sequence types (STs) were discovered, with ST28 and ST243 emerging as the predominant STs. The results of the pan-genomic analysis of S. suis indicate that there are 943 core genes, 2259 shell genes, and 5663 cloud genes. Through SNP evolutionary analysis, we identified a strong genetic similarity between SS31 and the reference genome P1/7. The analysis of antibiotic resistance genes revealed widespread presence of erm(B) and tet(O) genes among 19 strains of S. suis. This association may be linked to the high resistance of S. suis to lincosamides, macrolides, and tetracyclines. Integrative and conjugative elements (ICEs) and integrative and mobilizable elements (IMEs) were identified in 16 strains, with a carriage rate of 84.21%, and resistance genes were identified within the ICE/IME elements of 8 strains. Antimicrobial susceptibility testing revealed that all strains showed sensitivity to vancomycin and lincomycin but resistance to tilmicosin, tiamulin, amoxicillin, and doxycycline. This study contributes to our understanding of the genomic diversity of S. suis in Hubei Province of China, providing essential data for the comprehensive prevention and control of S. suis infections in China.

Integrative and Conjugative Elements and Prophage DNA as Carriers of Resistance Genes in Erysipelothrix rhusiopathiae Strains from Domestic Geese in Poland.

Goose erysipelas is a serious problem in waterfowl breeding in Poland. However, knowledge of the characteristics of Erysipelothrix rhusiopathiae strains causing this disease is limited. In this study, the antimicrobial susceptibility and serotypes of four E. rhusiopathiae strains from domestic geese were determined, and their whole-genome sequences (WGSs) were analyzed to detect resistance genes, integrative and conjugative elements (ICEs), and prophage DNA. Sequence type and the presence of resistance genes and transposons were compared with 363 publicly available E. rhusiopathiae strains, as well as 13 strains of other Erysipelothrix species. Four strains tested represented serotypes 2 and 5 and the MLST groups ST 4, 32, 242, and 243. Their assembled circular genomes ranged from 1.8 to 1.9 kb with a GC content of 36-37%; a small plasmid was detected in strain 1023. Strains 1023 and 267 were multidrug-resistant. The resistance genes detected in the genome of strain 1023 were erm47, tetM, and lsaE-lnuB-ant(6)-Ia-spw cluster, while strain 267 contained the tetM and ermB genes. Mutations in the gyrA gene were detected in both strains. The tetM gene was embedded in a Tn916-like transposon, which in strain 1023, together with the other resistance genes, was located on a large integrative and conjugative-like element of 130 kb designated as ICEEr1023. A minor integrative element of 74 kb was identified in strain 1012 (ICEEr1012). This work contributes to knowledge about the characteristics of E. rhusiopathiae bacteria and, for the first time, reveals the occurrence of erm47 and ermB resistance genes in strains of this species. Phage infection appears to be responsible for the introduction of the ermB gene into the genome of strain 267, while ICEs most likely play a key role in the spread of the other resistance genes identified in E. rhusiopathiae.

Pseudomonas stutzeri KC carries the pdt genes for carbon tetrachloride degradation on an integrative and conjugative element

Pseudomonas stutzeri KC can rapidly degrade carbon tetrachloride (CCl4) to CO2 by a fortuitous reaction with pyridine-2,6-bis(thiocarboxylic acid), a metal chelator encoded by pdt genes. These genes were first identified after a spontaneous mutant, strain CTN1, lost the ability to degrade CCl4. Here we report the complete genome of strain KC and show that these pdt genes are located on an integrative and conjugative element (ICE), designated ICEPsstKC. Comparative genome analyses revealed homologues of pdt genes in genomes of members of other gammaproteobacterial orders. Discrepancies between the tree topologies of the deduced pdt gene products and the host phylogeny based on 16S rRNA provided evidence for horizontal gene transfer (HGT) in several sequenced strains of these orders. In addition to ICEPsstKC, HGT may be have been facilitated by other mobile genetic elements, as indicated by the location of the pdt gene cluster adjacent to fragments of other ICEs and prophages in several genome assemblies. We could here show that the majority of cells from the culture collection DSMZ had lost the ICE. The presence of the pdt gene cluster on mobile genetic elements has important implications for the bioremediation of CCl4 for bioremediation of CCl4 and needs consideration when selecting suitable strains.

Characterization of erm(B)-Carrying Integrative and Conjugative Elements Transferred from Streptococcus anginosus to Other Streptococci and Enterococci.

Integrative and conjugative elements (ICEs) are important vectors of lateral gene transfer and contribute to the evolution of bacterial pathogens. However, studies on the transfer among species and the physiological consequences of ICEs are rare. The objective of this study was to investigate the cross-species transferability of newly identified erm(B)-carried ICE in Streptococcus anginosus San95 and its physiological consequences after transfer. The erm(B)-carried ICE, characterized by a triple serine integrase module, integrated into hsdM genes, thus designated ICESan95_hsdM. Analysis of ICESan95_hsdM revealed 32 additional ICESan95-like ICEs in the available NCBI genome (n = 24) and sequence of clinical isolates (n = 8). Polymerase chain reaction (PCR) was used to evaluate the 467 clinical isolates, of which 84 were positive for core genes (integrase, relaxase, and T4SS genes) of ICESan95_hsdM. Cross-species transfer experiments demonstrated that ICESan95_hsdM could transfer from S. anginosus to different streptococcal and enterococcal recipients. Growth and competitive culture assays showed acquisition of ICESan95_hsdM incurred no fitness cost. Our work discovered a group of ICEs in Streptococci and Enterococci. For the first time, we demonstrated the broad cross-species transferability to different species or genera of ICEs with no fitness cost that enables commensal S. anginosus to deliver antimicrobial resistance genes to other streptococci and enterococci.

Type II and IV toxin-antitoxin systems coordinately stabilize the integrative and conjugative element of the ICESa2603 family conferring multiple drug resistance in Streptococcus suis.

Integrative and conjugative elements (ICEs) play a vital role in bacterial evolution by carrying essential genes that confer adaptive functions to the host. Despite their importance, the mechanism underlying the stable inheritance of ICEs, which is necessary for the acquisition of new traits in bacteria, remains poorly understood. Here, we identified SezAT, a type II toxin-antitoxin (TA) system, and AbiE, a type IV TA system encoded within the ICESsuHN105, coordinately promote ICE stabilization and mediate multidrug resistance in Streptococcus suis. Deletion of SezAT or AbiE did not affect the strain's antibiotic susceptibility, but their duple deletion increased susceptibility, mainly mediated by the antitoxins SezA and AbiEi. Further studies have revealed that SezA and AbiEi affect the genetic stability of ICESsuHN105 by moderating the excision and extrachromosomal copy number, consequently affecting the antibiotic resistance conferred by ICE. The DNA-binding proteins AbiEi and SezA, which bind palindromic sequences in the promoter, coordinately modulate ICE excision and extracellular copy number by binding to sequences in the origin-of-transfer (oriT) and the attL sites, respectively. Furthermore, AbiEi negatively regulates the transcription of SezAT by binding directly to its promoter, optimizing the coordinate network of SezAT and AbiE in maintaining ICESsuHN105 stability. Importantly, SezAT and AbiE are widespread and conserved in ICEs harbouring diverse drug-resistance genes, and their coordinated effects in promoting ICE stability and mediating drug resistance may be broadly applicable to other ICEs. Altogether, our study uncovers the TA system's role in maintaining the genetic stability of ICE and offers potential targets for overcoming the dissemination and evolution of drug resistance.

Whole-genome sequence and resistance determinants of four Elizabethkingia anophelis clinical isolates collected in Hanoi, Vietnam

Four isolates of the opportunistic pathogen Elizabethkingia anophelis were identified for the first time in a Vietnamese hospital and underwent antimicrobial susceptibility testing and genomic characterization by whole-genome sequencing. Complete, fully circularized genome sequences were obtained for all four isolates. Average Nucleotide Identity analysis and single nucleotide polymorphism phylogenetic analysis on the core genome showed that three of the four isolates were genetically distinct, ruling out the hypothesis of a single strain emergence. Antibiotic susceptibility testing highlighted multi-resistant phenotypes against most antimicrobial families, including beta-lactams, carbapenems, aminoglycosides, quinolones, macrolides, amphenicols, rifamycins and glycopeptides. Additionally, in silico genomic analysis was used to correlate the phenotypic susceptibility to putative resistance determinants, including resistance genes, point mutations and multidrug efflux pumps. Nine different resistance genes were located inside a single resistance pocket predicted to be a putative Integrative and Conjugative Element (ICE). This novel ICE was shared by three isolates from two different lineages and displayed similarity with ICEs previously reported in various Elizabethkingia and Chryseobacterium species. The role of such ICEs in pathogenicity, genome plasticity and antimicrobial resistance gene spread within the Flavobacteriaceae family needs to be further elucidated.

VIM-type metallo-β-lactamase (MBL)-encoding genomic islands in Pseudomonas spp. in Poland: predominance of clc-like integrative and conjugative elements (ICEs).

To characterize VIM-type metallo-β-lactamase (MBL)-encoding genomic islands (GIs) in Pseudomonas aeruginosa and P. putida group isolates from Polish hospitals from 2001-2015/16. Twelve P. aeruginosa and 20 P. putida group isolates producing VIM-like MBLs were selected from a large collection of these based on epidemiological and typing data. The organisms represented all major epidemic genotypes of these species spread in Poland with chromosomally located blaVIM gene-carrying integrons. The previously determined short-read sequences were complemented by long-read sequencing in this study. The comparative structural analysis of the GIs used a variety of bioinformatic tools. Thirty different GIs with blaVIM integrons were identified in the 32 isolates, of which 24 GIs from 26 isolates were integrative and conjugative elements (ICEs) of the clc family. These in turn were dominated by 21 variants of the GI2/ICE6441 subfamily with a total of 19 VIM integrons, each inserted in the same position within the ICE's Tn21-like transposon Tn4380. The three other ICEs formed a novel ICE6705 subfamily, lacking Tn4380 and having different VIM integrons located in another site of the elements. The remaining six non-ICE GIs represented miscellaneous structures. The presence of various integrons in the same ICE sublineage, and of the same integron in different GIs, indicated circulation and recombination of the integron-carrying genetic platforms across Pseudomonas species/genotypes. Despite the general diversity of the blaVIM-carrying GIs in Pseudomonas spp. in Poland, a clear predominance of broadly spread and rapidly evolving clc-type ICEs was documented, confirming their significant role in antimicrobial resistance epidemiology.

Rapid dissemination of host metabolism–manipulating genes via integrative and conjugative elements

Integrative and conjugative elements (ICEs) are self-transmissible mobile elements that transfer functional genetic units across broad phylogenetic distances. Accessory genes shuttled by ICEs can make significant contributions to bacterial fitness. Most ICEs characterized to date encode readily observable phenotypes contributing to symbiosis, pathogenicity, and antimicrobial resistance, yet the majority of ICEs carry genes of unknown function. Recent observations of rapid acquisition of ICEs in a pandemic lineage of Pseudomonas syringae pv. actinidae led to investigation of the structural and functional diversity of these elements. Fifty-three unique ICE types were identified across the P. syringae species complex. Together they form a distinct family of ICEs (PsICEs) that share a distant relationship to ICEs found in Pseudomonas aeruginosa. PsICEs are defined by conserved backbone genes punctuated by an array of accessory cargo genes, are highly recombinogenic, and display distinct evolutionary histories compared to their bacterial hosts. The most common cargo is a recently disseminated 16-kb mobile genetic element designated Tn6212. Deletion of Tn6212 did not alter pathogen growth in planta, but mutants displayed fitness defects when grown on tricarboxylic acid (TCA) cycle intermediates. RNA-seq analysis of a set of nested deletion mutants showed that a Tn6212-encoded LysR regulator has global effects on chromosomal gene expression. We show that Tn6212 responds to preferred carbon sources and manipulates bacterial metabolism to maximize growth.

Macrolide resistance in Mannheimia haemolytica isolates associated with bovine respiratory disease from the German national resistance monitoring program GERM-Vet 2009 to 2020.

Data collected from the German national resistance monitoring program GERM-Vet showed slowly increasing prevalence of macrolide resistance among bovine respiratory disease (BRD)-associated Pasteurellacae from cattle over the last decade. The focus of this study was to analyze the genetic basis of antimicrobial resistance (AMR) and the prevalence of multidrug-resistance (MDR)-mediating integrative and conjugative elements (ICEs) in 13 German BRD-associated Mannheimia haemolytica isolates collected between 2009 and 2020 via whole-genome sequencing. Antimicrobial susceptibility testing (AST) was performed via broth microdilution according to the recommendations of the Clinical and Laboratory Standards Institute for the macrolides erythromycin, tilmicosin, tulathromycin, gamithromycin, tildipirosin, and tylosin as well as 25 other antimicrobial agents. All isolates either had elevated MICs or were resistant to at least one of the macrolides tested. Analysis of whole-genome sequences obtained by hybrid assembly of Illumina MiSeq and Oxford Nanopore MinION reads revealed the presence of seven novel Tn7406-like ICEs, designated Tn7694, and Tn7724- Tn7729. These ICEs harbored the antimicrobial resistance genes erm(T), mef (C), mph(G), floR, catA3, aad(3")(9), aph(3')-Ia, aac(3)-IIa, strA, strB, tet(Y), and sul2 in different combinations. In addition, mutational changes conferring resistance to macrolides, nalidixic acid or streptomycin, respectively, were detected among the M. haemolytica isolates. In addition, four isolates carried a 4,613-bp plasmid with the β-lactamase gene blaROB - 1. The detection of the macrolide resistance genes erm(T), mef (C), and mph(G) together with other resistance genes on MDR-mediating ICEs in bovine M. haemolytica may explain the occurrence of therapeutic failure when treating BRD with regularly used antimicrobial agents, such as phenicols, penicillins, tetracyclines, or macrolides. Finally, pathogen identification and subsequent AST is essential to ensure the efficacy of the antimicrobial agents applied to control BRD in cattle.

Genomic epidemiology and molecular characteristics of blaNDM-1-positive carbapenem-resistant Pseudomonas aeruginosa belonging to international high-risk clone ST773 in the Gauteng region, South Africa

PurposeThe emergence of carbapenem-resistant P. aeruginosa (CRPA) harbouring acquired carbapenemase genes (blaVIM, blaIMP and blaNDM) has become a global public health threat. Three CRPA isolates included in the study had an extensively drug-resistant phenotype with susceptibility to colistin only and were positive for the blaNDM-1 gene. The current study aimed to investigate the genomic epidemiology and molecular characteristics of the blaNDM-1-positive CRPA isolates collected from the Gauteng region, South Africa.MethodsShort read whole genome sequencing (WGS) was performed to determine sequence types (STs), genetic relatedness, resistome, virulome and the genetic environment of the blaNDM-1 gene.ResultsThe WGS and phylogenetic analyses revealed that the study isolates belonged to an international high-risk clone ST773 and belonged to the same clade with eight blaNDM-1-positive ST773 isolates from Hungary, India, Nigeria, South Korea and USA. The study isolates harboured a wide repertoire of intrinsic and acquired antibiotic resistance genes (ARGs) related with mobile genetic elements, porins and efflux pumps, as well as virulence factor genes. The clade-specific ARGs (blaNDM-1, floR2/cmlA9, rmtB4, tetG) were found in a putative integrative and conjugative element (ICE) region similar to ICE6660-like.ConclusionAs ICE carrying the blaNDM-1 gene can easily spread to other P. aeruginosa isolates and other Gram-negative bacteria, the findings in this study highlight the need for appropriate management strategies and active surveillance of CRPA isolates in the Gauteng region, South Africa.

Metagenomic analysis reveals impact of acyl homoserine endolipid-like signaling molecules on the aqueous sediment resistome under florfenicol stress

Quorum sensing potentially helps microorganisms adapt to antibiotic stress encountered in the environment. This experiment investigated the effect of acyl homoserine endolipid-like signaling molecules on microbial antibiotic resistance gene structures in aqueous sediments under florfenicol stress. Additional acyl homoserine endolipid-like signaling molecules (AHLs) alter the structure of multidrug resistance genes in florfenicol-stressed sediments, particularly the multidrug resistance efflux pump gene family. Prophages and integrative and conjugative elements (ICEs) determined the resistance genes structure, and pathways related to mobile genetic elements (MGEs) transfer may play an essential role in this process. The practical application of AHLs to regulate quorum sensing systems may alter bacterial stress responses to environmental florfenicol residues, thereby reducing the development of antibiotic resistance in the environment.

Novel resistance ICEs carrying the blaFIM-1 metallo-β-lactamase gene from an ST235 Pseudomonas aeruginosa sublineage.

FIM-1 is an acquired metallo-β-lactamase identified in a multidrug-resistant Pseudomonas aeruginosa (index strain FI-14/157) of clinical origin isolated in 2007 in Florence, Italy. Here we report on a second case of infection by FIM-1-positive P. aeruginosa (FI-17645), which occurred in 2020 in the same hospital. Both FIM-1-positive strains exhibited resistance to all anti-Pseudomonas antibiotics except colistin and cefiderocol. Comparative genomic characterization revealed that the two FIM-positive strains were closely related [core genome difference, 16 single nucleotide polymorphisms (SNPs)], suggesting a local circulation of similar strains. In the FI-14/157 index strain, the blaFIM-1 gene was associated with an ISCR19-like element that likely contributed to its capture downstream an integron platform inserted aboard a Tn21-like transposon, named Tn7703.1, which was associated with a large integrative and conjugative element (ICE) named ICE7705.1, integrated into an att site located within the 3'-end of tRNAGly CCC gene of the P. aeruginosa chromosome. In strain FI-17645, blaFIM-1 was associated with a closely related ICE, named ICE7705.2, integrated in the same chromosomal site. Similar ICE platforms, lacking the blaFIM-1-containing region, were detected in other ST235 P. aeruginosa strains from different geographic areas, suggesting a common ancestry and underscoring the role of these elements in the dissemination of resistance genes in P. aeruginosa. Sequence database mining revealed two draft P. aeruginosa genomes, one from Italy and one from the USA (both isolated in 2012), including a contig with blaFIM-1, suggesting that this resistance gene could have a broader distribution than originally anticipated.

Characterization and risk assessment of novel SXT/R391 integrative and conjugative elements with multidrug resistance in Proteus mirabilis isolated from China, 2018-2020.

Proteus mirabilis can transfer transposons, insertion sequences, and gene cassettes to the chromosomes of other hosts through SXT/R391 integrative and conjugative elements (ICEs), significantly increasing the possibility of antibiotic resistance gene (ARG) evolution and expanding the risk of ARGs transmission among bacteria. A total of 103 strains of P. mirabilis were isolated from 25 farms in China from 2018 to 2020. The positive detection rate of SXT/R391 ICEs was 25.2% (26/103). All SXT/R391 ICEs positive P. mirabilis exhibited a high level of overall drug resistance. Conjugation experiments showed that all 26 SXT/R391 ICEs could efficiently transfer to Escherichia coli EC600 with a frequency of 2.0 × 10-7 to 6.0 × 10-5. The acquired ARGs, genetic structures, homology relationships, and conservation sequences of 26 (19 different subtypes) SXT/R391 ICEs were investigated by high-throughput sequencing, whole-genome typing, and phylogenetic tree construction. ICEPmiChnHBRJC2 carries erm (42), which have never been found within an SXT/R391 ICE in P. mirabilis, and ICEPmiChnSC1111 carries 19 ARGs, including clinically important cfr, blaCTX-M-65, and aac(6')-Ib-cr, making it the ICE with the most ARGs reported to date. Through genetic stability, growth curve, and competition experiments, it was found that the transconjugant of ICEPmiChnSCNNC12 did not have a significant fitness cost on the recipient bacterium EC600 and may have a higher risk of transmission and dissemination. Although the transconjugant of ICEPmiChnSCSZC20 had a relatively obvious fitness cost on EC600, long-term resistance selection pressure may improve bacterial fitness through compensatory adaptation, providing scientific evidence for risk assessment of horizontal transfer and dissemination of SXT/R391 ICEs in P. mirabilis.IMPORTANCEThe spread of antibiotic resistance genes (ARGs) is a major public health concern. The study investigated the prevalence and genetic diversity of integrative and conjugative elements (ICEs) in Proteus mirabilis, which can transfer ARGs to other hosts. The study found that all of the P. mirabilis strains carrying ICEs exhibited a high level of drug resistance and a higher risk of transmission and dissemination of ARGs. The analysis of novel multidrug-resistant ICEs highlighted the potential for the evolution and spread of novel resistance mechanisms. These findings emphasize the importance of monitoring the spread of ICEs carrying ARGs and the urgent need for effective strategies to combat antibiotic resistance. Understanding the genetic diversity and potential for transmission of ARGs among bacteria is crucial for developing targeted interventions to mitigate the threat of antibiotic resistance.

Detection of CTX-M-15 ESBL in XDR Haemophilus parainfluenzae from a urethral swab.

Haemophilus parainfluenzae is an opportunistic pathogen causing respiratory tract infection and sexually transmitted diseases. The emergence of multidrug resistance in this species is particularly worrisome, especially since the recent description of CTX-M-15 ESBL-producing isolates in Spain. The aim of this study was to characterize a CTX-M-15-producing H. parainfluenzae clinical isolate, HP01, obtained from a urethral swab. MICs were determined with gradient strips for this isolate. Hydrolysis assays were performed with the β LACTA test. Genomic DNA from HP01 was subjected to Illumina and Oxford Nanopore sequencing to investigate the genetic environment of blaCTX-M-15. Phylogenetic analysis was performed with available H. parainfluenzae genomes from the NCBI database, including CTX-M-15 producers. HP01, an XDR isolate, was resistant to penicillin, third-generation cephalosporins, fluoroquinolones, macrolides, cyclines and co-trimoxazole and susceptible only to carbapenems and rifampicin. HP01 carried blaTEM-1, blaCTX-M-15, tet(M), catS and mef(E)/mel and harboured amino acid substitutions in PBP3, PBP5, GyrA, ParC and FolA implicated in resistance. Genomic analysis revealed that blaCTX-M-15 was carried by a Tn3-like transposon inserted into a novel integrative and conjugative element (ICE), ICEHpaSLS, present on the chromosome and belonging to the ICEHin1056 family described in Haemophilus influenzae. The tet(M)-MEGA element was also detected on the chromosome. No plasmid was found. The phylogenetic analysis showed that four H. parainfluenzae producing CTX-M-15 clustered in the same clade. Here we report the description of an XDR H. parainfluenzae producing blaCTX-M-15 isolated from a urethral swab. The blaCTX-M-15 gene was inserted into an ICE structure similar to those recently described in CTX-M-15 producers in Spain. The emergence of XDR H. parainfluenzae producing blaCTX-M-15 is a matter of great concern. Careful surveillance is required to prevent its spread.

Genetic and Functional Characterization of a Salicylate 1-monooxygenase Located on an Integrative and Conjugative Element (ICE) in Pseudomonas stutzeri AJR13.

Pseudomonas stutzeri strain AJR13 was isolated for growth on the related compounds biphenyl (BPH) and diphenylmethane (DPM). The BPH and DPM degradative pathway genes are present on an integrative and conjugative element (ICE) in the chromosome. Examination of the genome sequence of AJR13 revealed a gene encoding a salicylate 1-monooxygenase (salA) associated with the ICE even though AJR13 did not grow on salicylate. Transfer of the ICE to the well-studied Pseudomonas putida KT2440 resulted in a KT2440 strain that could grow on salicylate. Knockout mutagenesis of the salA gene on the ICE in KT2440 eliminated the ability to grow on salicylate. Complementation of the knockout with the cloned salA gene restored growth on salicylate. Transfer of the cloned salA gene under control of the lac promoter to KT2440 resulted in a strain that could grow on salicylate. Heterologous expression of the salA gene in E. coli BL21 DE3 resulted in the production of catechol from salicylate, confirming that it is indeed a salicylate 1-monooxygenase. Interestingly, transfer of the cloned salA gene under control of the lac promoter to AJR13 resulted in a strain that could now grow on salicylate, suggesting that gene expression for the downstream catechol pathway is intact.

Genomic analysis reveals the population structure and antimicrobial resistance of avian Pasteurella multocida in China.

To investigate the population structure and antimicrobial resistance (AMR) of avian Pasteurella multocida in China. Utilizing WGS analysis, we explored the phylogeny using a dataset of 546 genomes, comprising avian P. multocida isolates from China (n = 121), the USA (n = 165), Australia(n = 153), Bangladesh (n = 3) and isolates of other hosts from China (n = 104). We examined the integrative and conjugative element (ICE) structures and the distribution of their components carrying resistance genes, and reconstructed the evolutionary history of A:L1:ST129 (n = 110). The population structure of avian P. multocida in China was dominated by the A:L1:ST129 clone with limited genetic diversity. A:L1:ST129 isolates possessed a broader spectrum of resistance genes at comparatively higher frequencies than those from other hosts and countries. The novel putative ICEs harboured complex resistant clusters that were prevalent in A:L1:ST129. Bayesian analysis predicted that the A:L1:ST129 clone emerged around 1923, and evolved slowly. A:L1:ST129 appears to possess a host predilection towards avian species in China, posing a potential health threat to other animals. The complex AMR determinants coupled with high frequencies may strengthen the population dominance of A:L1:ST129. The extensive antimicrobial utilization in poultry farming and the mixed rearing practices could have accelerated AMR accumulation in A:L1:ST129. ICEs, together with their resistant clusters, significantly contribute to resistance gene transfer and facilitate the adaptation of A:L1:ST129 to ecological niches. Despite the genetic stability and slow evolution rate, A:L1:ST129 deserves continued monitoring due to its propensity to retain resistance genes, warranting global attention to preclude substantial economic losses.

A new Salmonella enterica serovar that was isolated from a wild sparrow presents a distinct genetic, metabolic and virulence profile

Salmonella enterica is a ubiquitous and clinically-important bacterial pathogen, able to infect and cause different diseases in a wide range of hosts. Here, we report the isolation and characterization of a new S. enterica serovar (13,23:i:-; S. Tirat-Zvi), belonging to the Havana supper-lineage that was isolated from a wild house sparrow (Passer domesticus) in Israel. Whole genome sequencing and complete assembly of its genome indicated a plasmid-free, 4.7 Mb genome that carries the Salmonella pathogenicity islands 1–6, 9, 19 and an integrative and conjugative element (ICE), encoding arsenic resistance genes. Phenotypically, S. Tirat-Zvi isolate TZ282 was motile, readily formed biofilm, more versatile in carbon source utilization than S. Typhimurium and highly tolerant to arsenic, but impaired in host cell invasion. In-vivo infection studies indicated that while S. Tirat-Zvi was able to infect and cause an acute inflammatory enterocolitis in young chicks, it was compromised in mice colonization and did not cause an inflammatory colitis in mice compared to S. Typhimurium. We suggest that these phenotypes reflect the distinctive ecological niche of this new serovar and its evolutionary adaptation to passerine birds, as a permissive host. Moreover, these results further illuminate the genetic, phenotypic and ecological diversity of S. enterica pathovars.

Diversity and evolution of an abundant ICEclc family of integrative and conjugative elements in Pseudomonas aeruginosa.

Microbial populations swiftly adapt to changing environments through horizontal gene transfer. While the mechanisms of gene transfer are well known, the impact of environmental conditions on the selection of transferred gene functions remains less clear. We investigated ICEs, specifically the ICEclc-type, in Pseudomonas aeruginosa clinical isolates. Our findings revealed co-evolution between ICEs and their hosts, with ICE transfers occurring within strains. Gene functions carried by ICEs are positively selected, including potential virulence factors and heavy metal resistance. Comparison to publicly available P. aeruginosa genomes unveiled widespread antibiotic-resistance determinants within ICEclc clades. Thus, the ubiquitous ICEclc family significantly contributes to P. aeruginosa's adaptation and fitness in diverse environments.

ICEberg 3.0: functional categorization and analysis of the integrative and conjugative elements in bacteria.

ICEberg 3.0 (https://tool2-mml.sjtu.edu.cn/ICEberg3/) is an upgraded database that provides comprehensive insights into bacterial integrative and conjugative elements (ICEs). In comparison to the previous version, three key enhancements were introduced: First, through text mining and manual curation, it now encompasses details of 2065 ICEs, 607 IMEsand 275 CIMEs, including 430 with experimental support. Secondly, ICEberg 3.0 systematically categorizes cargo gene functions of ICEs into six groups based on literature curation and predictive analysis, providing a profound understanding of ICEs'diverse biological traits. The cargo gene prediction pipeline is integrated into the online tool ICEfinder 2.0. Finally, ICEberg 3.0 aids the analysis and exploration of ICEs from the human microbiome. Extracted and manually curated from 2405 distinct human microbiome samples, the database comprises 1386 putative ICEs, offering insights into the complex dynamics of Bacteria-ICE-Cargo networks within the human microbiome. With the recent updates, ICEberg 3.0 enhances its capability to unravel the intricacies of ICE biology, particularly in the characterization and understanding of cargo gene functions and ICE interactions within the microbiome. This enhancement may facilitate the investigation of the dynamic landscape of ICE biology and its implications for microbial communities.