基因转移因子———一类在海洋中广泛存在的介导水平基因转移的新型生物因子

基因转移因子(Gene Transfer Agent,GTA)是一种由细菌释放的、形态和有尾病毒类似的生物颗粒。GTA颗粒携带的遗传物质是宿主基因组的随机小片段而不包含编码GTA自身的基因或病毒基因组。根据4个模式菌株释放的GTA的研究,GTA具有高效的,种间介导基因水平转移的功能。近年来大规模细菌基因组测序,发现编码GTA的基因簇广泛存在于海洋细菌基因组上,GTA是在海洋环境中发生水平基因转移的重要模式。本文在总结4个模式菌株释放的GTA的认识的基础上,着重描述海洋主要类群的细菌释放的GTA的特征,讨论在海洋生态系统中,GTA对水平基因转移的贡献,并对未来的研究进行了展望。     

Release and uptake of gene transfer agent by Rhodopseudomonas capsulata.

Many strains of Rhodopseudomonas capsulata are capable of exchanging genetic information via a recently discovered gene transfer process involving the release and subsequent uptake from the medium of particles containing genetic information (gene transfer agents, GTAs). No viral activities are observed to be associated with this system. An assay has been developed to quantitate gene transfer in R. capsulata. Conditions are described for which the number of cells acquiring a new genetic trait is direcly proportional to the number of GTAs and independent of the number of receipient cells. These conditions were used for the assay of the uptake and release of GTAs by cells. The maximum fraction of recipients that acquire a given genetic marker is approximately 4 X 10(-4). Free GTA appears in a growing culture in one or two abrupt waves near the time of transition from exponential to stationary phase. During these waves, the titer of GTA for a given marker may reach 2 X 103/ml. A comparison of the frequency of single- and double-marker transfers suggests that most of the cells in early-stationary-phase cultures are active recipients. The ultraviolet inactivation spectrum of GTA resembles that of the small ribonucleic acid phages. The inactivation cross section section beta, for GTA was calculated to be 1.7 X 10(-16) cm2/photon at 265 nm.     

Prophage-like gene transfer agents-novel mechanisms of gene exchange for Methanococcus, Desulfovibrio, Brachyspira, and Rhodobacter species

Gene transfer agents (GTAs) are novel mechanisms for bacterial gene transfer. They resemble small, tailed bacteriophages in ultrastructure and act like generalized transducing prophages. In contrast to functional prophages, GTAs package random fragments of bacterial genomes and incomplete copies of their own genomes. The packaged DNA content is characteristic of the GTA and ranges in size from 4.4 to 13.6kb. GTAs have been reported in species of Brachyspira, Methanococcus, Desulfovibrio, and Rhodobacter. The best studied GTAs are VSH-1 of the anaerobic, pathogenic spirochete Brachyspira hyodysenteriae and RcGTA of the nonsulfur, purple, photosynthetic bacterium Rhodobacter capsulatus. VSH-1 and RcGTA have likely contributed to the ecology and evolution of these bacteria. The existence of GTAs in phylogenetically diverse bacteria suggests GTAs may be more common in nature than is now appreciated.     

The gene transfer agent-like particle of the marine phototrophic bacterium Rhodovulum sulfidophilum

Gene transfer agents (GTAs) are shaped like bacteriophage particles but have many properties that distinguish them from bacteriophages. GTAs play a role in horizontal gene transfer in nature and thus affect the evolution of prokaryotic genomes. In the course of studies on the extracellular production of designed RNAs using the marine bacterium Rhodovulum sulfidophilum, we found that this bacterium produces a GTA-like particle. The particle contains DNA fragments of 4.5 kb, which consist of randomly fragmented genomic DNA from the bacterium. This 4.5-kb DNA production was prevented while quorum sensing was inhibited. Direct observation of the particle by transmission electron microscopy revealed that the particle resembles a tailed phage and has a head diameter of about 40 nm and a tail length of about 60 nm. We also identified the structural genes for the GTA in the genome. Translated amino acid sequences and gene positions are closely related to those of the genes that encode the Rhodobacter capsulatus GTA. This is the first report of a GTA-like particle from the genus Rhodovulum. However, gene transfer activity of this particle has not yet been confirmed. The differences between this particle and other GTAs are discussed.     

Gene transfer agents, bacteriophages, and bacteriocins of Rhodopseudomonas capsulata.

Thirty-three wild type strains of Rhodopseudomonas capsulata were examined for ability to engage in genetic recombination through mediation by "gene transfer agent" (GTA) particles. The genetic exchange assays were based on capacity of strains to produce or receive GTA required for restoration of photosynthetic growth competence to a non-photosynthetic "white" mutant or for acquisition of resistance to rifampicin. A majority of the strains could either produce or receive GTA, and it was demonstrated that the agent is species specific. Possible relations between GTA and bacteriophages or bacteriocins were investigated. Sixteen types of virulent phages active on Rps. capsulata were isolated and their host ranges determined. Tests for transduction by the phages gave uniformly negative results. The viruses showed strict species specificity, but there was no apparent correlation between capacity of the Rps. capsulata strains to donate or receive GTA and susceptibility to the phages. A comparable survey disclosed that most of the bacterial strains were sensitive to or capable of producing bacteriocins; the latter also appear to be unrelated to GTA activity. The collection of bacterial strains was also screened for detection of lysogenic properties. None of the isolates is a "true" lysogen, but phages were detected in cultures of two strains, which may be "phage carriers" or pseudolysogens.     

A Gene Transfer Agent and a Dynamic Repertoire of Secretion Systems Hold the Keys to the Explosive Radiation of the Emerging Pathogen Bartonella

Gene transfer agents (GTAs) randomly transfer short fragments of a bacterial genome. A novel putative GTA was recently discovered in the mouse-infecting bacterium Bartonella grahamii. Although GTAs are widespread in phylogenetically diverse bacteria, their role in evolution is largely unknown. Here, we present a comparative analysis of 16 Bartonella genomes ranging from 1.4 to 2.6 Mb in size, including six novel genomes from Bartonella isolated from a cow, two moose, two dogs, and a kangaroo. A phylogenetic tree inferred from 428 orthologous core genes indicates that the deadly human pathogen B. bacilliformis is related to the ruminant-adapted clade, rather than being the earliest diverging species in the genus as previously thought. A gene flux analysis identified 12 genes for a GTA and a phage-derived origin of replication as the most conserved innovations. These are located in a region of a few hundred kb that also contains 8 insertions of gene clusters for type III, IV, and V secretion systems, and genes for putatively secreted molecules such as cholera-like toxins. The phylogenies indicate a recent transfer of seven genes in the virB gene cluster for a type IV secretion system from a cat-adapted B. henselae to a dog-adapted B. vinsonii strain. We show that the B. henselae GTA is functional and can transfer genes in vitro. We suggest that the maintenance of the GTA is driven by selection to increase the likelihood of horizontal gene transfer and argue that this process is beneficial at the population level, by facilitating adaptive evolution of the host-adaptation systems and thereby expansion of the host range size. The process counters gene loss and forces all cells to contribute to the production of the GTA and the secreted molecules. The results advance our understanding of the role that GTAs play for the evolution of bacterial genomes.     

Gene transfer agents,bacteriophages, and bacteriocins of Rhodopseudomonas capsulata

Thirty-three wild type strains of Rhodopseudomonas capsulata were examined for ability to engage in genetic recombination through mediation by “gene transfer agent” (GTA) particles. The genetic exchange assays were based on capacity of strains to produce or receive GTA required for restoration of photosynthetic growth competence to a non-photosynthetic “white” mutant or for acquisition of resistance to rifampicin. A majority of the strains could either produce or receive GTA, and it was demonstrated that the agent is species specific. Possible relations between GTA and bacteriophages or bacteriocins were investigated. Sixteen types of virulent phages active on Rps. capsulata were isolated and their host ranges determined. Tests for transduction by the phages gave uniformly negative results. The viruses showed strict species specificity, but there was no apparent correlation between capacity of the Rps. capsulata strains to donate or receive GTA and susceptibility to the phages. A comparable survey disclosed that most of the bacterial strains were sensitive to or capable of producing bacteriocins; the latter also appear to be unrelated to GTA activity. The collection of bacterial strains was also screened for detection of lysogenic properties. None of the isolates is a “true” lysogen, but phages were detected in cultures of two strains, which may be “phage carriers” or pseudolysogens.     

Occurrence and expression of gene transfer agent genes in marine bacterioplankton

Genes with homology to the transduction-like gene transfer agent (GTA) were observed in genome sequences of three cultured members of the marine Roseobacter clade. A broader search for homologs for this host-controlled virus-like gene transfer system identified likely GTA systems in cultured Alphaproteobacteria, and particularly in marine bacterioplankton representatives. Expression of GTA genes and extracellular release of GTA particles ( approximately 50 to 70 nm) was demonstrated experimentally for the Roseobacter clade member Silicibacter pomeroyi DSS-3, and intraspecific gene transfer was documented. GTA homologs are surprisingly infrequent in marine metagenomic sequence data, however, and the role of this lateral gene transfer mechanism in ocean bacterioplankton communities remains unclear.     

Gene transfer agents: phage-like elements of genetic exchange

Horizontal gene transfer is important in the evolution of bacterial and archaeal genomes. An interesting genetic exchange process is carried out by diverse phage-like gene transfer agents (GTAs) that are found in a wide range of prokaryotes. Although GTAs resemble phages, they lack the hallmark capabilities that define typical phages, and they package random pieces of the producing cell's genome. In this Review, we discuss the defining characteristics of the GTAs that have been identified to date, along with potential functions for these agents and the possible evolutionary forces that act on the genes involved in their production.     

Complete-fosmid and fosmid-end sequences reveal frequent horizontal gene transfers in marine uncultured planktonic archaea

The extent of horizontal gene transfer (HGT) among marine pelagic prokaryotes and the role that HGT may have played in their adaptation to this particular environment remain open questions. This is partly due to the paucity of cultured species and genomic information for many widespread groups of marine bacteria and archaea. Molecular studies have revealed a large diversity and relative abundance of marine planktonic archaea, in particular of Thaumarchaeota (also known as group I Crenarchaeota) and Euryarchaeota of groups II and III, but only one species (the thaumarchaeote Candidatus Nitrosopumilus maritimus) has been isolated in pure culture so far. Therefore, metagenomics remains the most powerful approach to study these environmental groups. To investigate the impact of HGT in marine archaea, we carried out detailed phylogenetic analyses of all open reading frames of 21 archaeal 16S rRNA gene-containing fosmids and, to extend our analysis to other genomic regions, also of fosmid-end sequences of 12 774 fosmids from three different deep-sea locations (South Atlantic and Adriatic Sea at 1000 m depth, and Ionian Sea at 3000 m depth). We found high HGT rates in both marine planktonic Thaumarchaeota and Euryarchaeota, with remarkable converging values estimated from complete-fosmid and fosmid-end sequence analysis (25 and 21% of the genes, respectively). Most HGTs came from bacterial donors (mainly from Proteobacteria, Firmicutes and Chloroflexi) but also from other archaea and eukaryotes. Phylogenetic analyses showed that in most cases HGTs are shared by several representatives of the studied groups, implying that they are ancient and have been conserved over relatively long evolutionary periods. This, together with the functions carried out by these acquired genes (mostly related to energy metabolism and transport of metabolites across membranes), suggests that HGT has played an important role in the adaptation of these archaea to the cold and nutrient-depleted deep marine environment.     

Inquiry-based training improves teaching effectiveness of biology teaching assistants

Graduate teaching assistants (GTAs) are used extensively as undergraduate science lab instructors at universities, yet they often have having minimal instructional training and little is known about effective training methods. This blind randomized control trial study assessed the impact of two training regimens on GTA teaching effectiveness. GTAs teaching undergraduate biology labs (n = 52) completed five hours of training in either inquiry-based learning pedagogy or general instructional “best practices”. GTA teaching effectiveness was evaluated using: (1) a nine-factor student evaluation of educational quality; (2) a six-factor questionnaire for student learning; and (3) course grades. Ratings from both GTAs and undergraduates indicated that indicated that the inquiry-based learning pedagogy training has a positive effect on GTA teaching effectiveness.     

Interstrain transfer of the prophage ϕNM2 in staphylococcal strains

Staphylococcus aureus is a successful pathogen in part because the bacterium can adapt rapidly to selective pressures imparted by the external environment. Horizontal gene transfer (HGT) plays an integral role in the evolution of bacterial genomes, and phage transduction is likely to be the most common and important HGT mechanism for S. aureus. Phage can transfer not only its own genome DNA but also host bacterial DNA with or without pathogenicity islands to other bacteria. Here, we demonstrate that the staphylococcal prophage ?NM2 could transfer between strains Newman and NCTC8325/NCTC8325-4 by simulating a natural situation in laboratory without mitomycin C or ultra-violet light treatment. This transference may be caused by direct contact between Newman and NCTC8325/NCTC8325-4 instead of phage particles released in Newman culture’s supernatant. The rates of successful horizontal genetic transfer in recipients NCTC8325 and NCTC8325-4 were 2.1% and 1.8%, respectively. Prophage ?NM2 was integrated with one direction at an intergenic region between rpmF and isdB in all 17 lysogenic isolates. Phage particles were spontaneously released from lysogenic strains again and had no noticeable influence on the growth of host cells. The results reported herein provide insight into how mobile genetic elements such as prophages can lead to the emergence of genetic diversity among S. aureus strains.     

Shuffling bacterial metabolomes

Horizontal gene transfer (HGT) has a far more significant role than gene duplication in bacterial evolution. This has recently been illustrated by work demonstrating the importance of HGT in the emergence of bacterial metabolic networks, with horizontally acquired genes being placed in peripheral pathways at the outer branches of the networks.     

Characterization of the gene transfer agent made by an overproducer mutant of Rhodopseudomonas capsulata.

Most wild-type isolates of the photosynthetic bacterium Rhodopseudomonas capsulata spontaneously produce nucleoprotein particles that act as vectors of genetic exchange in this species. The low yield of these particles, termed gene transfer agents, has made their characterization difficult. We have utilized a new screening technique to select a mutant, Y262, which produces about three orders of magnitude more GTA² per culture. The GTA produced by Y262 seem to be identical to those of wild type by immunological, genetic and sedimentation analyses, and they have been further characterized by a variety of techniques. GTA resemble tailed bacteriophage, although GTA with a head diameter of 30 nm are smaller than most morphologically similar viruses. DNA isolated from GTA is in the form of linear duplex molecules of about 3 × 10⁶ molecular weight. There is no phage-like DNA in GTA particles as judged by C_ot and restriction endonuclease analyses. Instead, GTA DNA shows the same kinetic complexity and restriction endonuclease digestion pattern as the R. capsulata genome. The GTA of ten independently isolated wild-type strains of R. capsulata show a high degree of immunological cross-reactivity; thus this genetic exchange system, while not a species-wide trait, does seem to be the product of a conservative evolutionary process.     

Plasmid-mediated horizontal gene transfer is a coevolutionary process

Conjugative plasmids are key agents of horizontal gene transfer (HGT) that accelerate bacterial adaptation by vectoring ecologically important traits between strains and species. However, although many conjugative plasmids carry beneficial traits, all plasmids exert physiological costs-of-carriage on bacteria. The existence of conjugative plasmids, therefore, presents a paradox because non-beneficial plasmids should be lost to purifying selection, whereas beneficial genes carried on plasmids should be integrated into the bacterial chromosome. Several ecological solutions to the paradox have been proposed, but none account for co-adaptation of bacteria and conjugative plasmids. Drawing upon evidence from experimental evolution, we argue that HGT via conjugation can only be fully understood in a coevolutionary framework.     

An ancient horizontal gene transfer between mosquito and the endosymbiotic bacterium Wolbachia pipientis

The extent and biological relevance of horizontal gene transfer (HGT) in eukaryotic evolution remain highly controversial. Recent studies have demonstrated frequent and large-scale HGT from endosymbiotic bacteria to their hosts, but the great majority of these transferred genes rapidly become nonfunctional in the recipient genome. Here, we investigate an ancient HGT between a host metazoan and an endosymbiotic bacterium, Wolbachia pipientis. The transferred gene has so far been found only in mosquitoes and Wolbachia. In mosquitoes, it is a member of a gene family encoding candidate receptors required for malaria sporozoite invasion of the mosquito salivary gland. The gene copy in Wolbachia has substantially diverged in sequence from the mosquito homolog, is evolving under purifying selection, and is expressed, suggesting that this gene is also functional in the bacterial genome. Several lines of evidence indicate that the gene may have been transferred from eukaryotic host to bacterial endosymbiont. Regardless of the direction of transfer, however, these results demonstrate that interdomain HGT may give rise to functional, persistent, and possibly evolutionarily significant new genes.