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1.
Davies RL  Lee I 《Journal of bacteriology》2004,186(17):5741-5752
The OmpA (or heat-modifiable) protein is a major structural component of the outer membranes of gram-negative bacteria. The protein contains eight membrane-traversing beta-strands and four surface-exposed loops. The genetic diversity and molecular evolution of OmpA were investigated in 31 Mannheimia (Pasteurella) haemolytica, 6 Mannheimia glucosida, and 4 Pasteurella trehalosi strains by comparative nucleotide sequence analysis. The OmpA proteins of M. haemolytica and M. glucosida contain four hypervariable domains located at the distal ends of the surface-exposed loops. The hypervariable domains of OmpA proteins from bovine and ovine M. haemolytica isolates are very different but are highly conserved among strains from each of these two host species. Fourteen different alleles representing four distinct phylogenetic classes, classes I to IV, were identified in M. haemolytica and M. glucosida. Class I, II, and IV alleles were associated with bovine M. haemolytica, ovine M. haemolytica, and M. glucosida strains, respectively, whereas class III alleles were present in certain M. haemolytica and M. glucosida isolates. Class I and II alleles were associated with divergent lineages of bovine and ovine M. haemolytica strains, respectively, indicating a history of horizontal DNA transfer and assortative (entire gene) recombination. Class III alleles have mosaic structures and were derived by horizontal DNA transfer and intragenic recombination. Our findings suggest that OmpA is under strong selective pressure from the host species and that it plays an important role in host adaptation. It is proposed that the OmpA protein of M. haemolytica acts as a ligand and is involved in binding to specific host cell receptor molecules in cattle and sheep. P. trehalosi expresses two OmpA homologs that are encoded by different tandemly arranged ompA genes. The P. trehalosi ompA genes are highly diverged from those of M. haemolytica and M. glucosida, and evidence is presented to suggest that at least one of these genes was acquired by horizontal DNA transfer.  相似文献   

2.
Mannheimia haemolytica and Bibersteinia (Pasteurella) trehalosi are the most common bacterial isolates that cause pulmonary diseases in ruminants worldwide. The disease is determined by specific serotypes found in cattle and small ruminants. The molecular epidemiology of strains involved in disease is important in the control of outbreaks as well as in the preparation of vaccines. This study aimed to detect the instability and variations of bacterial strains that may affect the analysis of epidemic strains, or the stability of vaccinal strains. Eight strains of M. haemolytica belonging to serotypes A1 and A2 and three B. trehalosi strains of the T3 and T4 serotypes were used. Strains were subjected to pulsed field gel electrophoresis (PFGE) and capsular and phenotypic typing at each round of a total of 50 successive subcultures. Remarkable stability was found in all selected strains of B. trehalosi in contrast to M. haemoltyica, in which strains of both serotypes showed pattern variations produced by PFGE and capsular and phenotypic analysis. Objective criteria for M. haemolytica and B. trehalosi typing are consequently addressed.  相似文献   

3.
The mosaic structure and molecular evolution of the leukotoxin operon (lktCABD) was investigated by nucleotide sequence comparison of the lktC, lktB, and lktD genes in 23 Mannheimia (Pasteurella) haemolytica, 6 Mannheimia glucosida, and 4 Pasteurella trehalosi strains. Sequence variation in the lktA gene has been described previously (R. L. Davies et al., J. Bacteriol. 183:1394-1404, 2001). The leukotoxin operon of M. haemolytica has a complex mosaic structure and has been derived by extensive inter- and intraspecies horizontal DNA transfer and intragenic recombination events. However, the pattern of recombination varies throughout the operon and among the different evolutionary lineages of M. haemolytica. The lktA and lktB genes have the most complex mosaic structures with segments derived from up to four different sources, including M. glucosida and P. trehalosi. In contrast, the lktD gene is highly conserved in M. haemolytica. The lktC, lktA, and lktB genes of strains representing the major ovine lineages contain recombinant segments derived from bovine or bovine-like serotype A2 strains. These findings support the previous conclusion that host switching of bovine A2 strains from cattle to sheep has played a major role in the evolution of the leukotoxin operon in ovine strains of M. haemolytica. Homologous segments of donor and recipient alleles are identical, or nearly identical, indicating that the recombinational exchanges occurred relatively recent in evolutionary terms. The 5' and 3' ends of the operon are highly conserved in M. haemolytica, which suggests that multiple horizontal exchanges of the complete operon have occurred by a common mechanism such as transduction. Although the lktA and lktB genes both have complex mosaic structures and high nucleotide substitution rates, the amino acid diversity of LktB is significantly lower than that of LktA due to a higher degree of evolutionary constraint against amino acid replacement. The recombinational exchanges within the leukotoxin operon have had greatest effect on LktA and probably provide an adaptive advantage against the host antibody response by generating novel antigenic variation at surface-exposed sites.  相似文献   

4.
To date, a number of Myoviridae bacteriophages that infect Aeromonadaceae have been identified and characterized. However, the genome sequences of Aeromonas phages that not belong to the Myoviridae have not been investigated yet. Herein, we report the complete genome sequence of Aeromonas phage phiAS7, which belongs to the Podoviridae and infects Aeromonas salmonicida subsp. salmonicida.  相似文献   

5.
【目的】从环境中分离获得希瓦氏菌烈性噬菌体,并对其性质进行研究。【方法】以4株希瓦氏菌为宿主菌,采用双层平板法从污水样品中分离得到奥奈达希瓦氏菌MR-1烈性噬菌体M1;观察噬菌斑特征;利用超速离心法浓缩M1颗粒,进一步用氯化铯密度梯度离心纯化;采用透射电子显微镜观察纯化的M1颗粒;提取M1核酸,通过核酸酶处理分析其核酸类型及结构;绘制一步生长曲线。【结果】噬菌体M1在双层平板上形成圆形的噬菌斑,清晰透明,边缘光滑,直径为2.3 mm-2.5 mm;经电镜观察,噬菌体M1头部呈二十面体,直径约为55 nm,尾长约为170 nm,尾部可收缩,属于肌尾噬菌体科(Myoviridae);通过酶切分析表明噬菌体M1核酸为线形双链DNA;一步生长曲线显示该噬菌体感染后完成一个复制循环所需要的时间约为15-20 min。【结论】噬菌体M1属肌尾噬菌体科,研究结果为后续研究病毒在地球微生物成岩过程中所起的作用提供了实验材料。  相似文献   

6.
Mannheimia haemolytica, Pasteurella multocida, and Bibersteinia trehalosi have been identified in the lungs of pneumonic bighorn sheep (BHS; Ovis canadensis). Of these pathogens, M. haemolytica has been shown to consistently cause fatal pneumonia in BHS under experimental conditions. However, M. haemolytica has been isolated by culture less frequently than the other bacteria. We hypothesized that the growth of M. haemolytica is inhibited by other bacteria in the lungs of BHS. The objective of this study was to determine whether P. multocida inhibits the growth of M. haemolytica. Although in monoculture both bacteria exhibited similar growth characteristics, in coculture with P. multocida there was a clear inhibition of growth of M. haemolytica. The inhibition was detected at mid-log phase and continued through the stationary phase. When cultured in the same medium, the growth of M. haemolytica was inhibited when both bacteria were separated by a membrane that allowed contact (pore size, 8.0 μm) but not when they were separated by a membrane that limited contact (pore size, 0.4 μm). Lytic bacteriophages or bactericidal compounds could not be detected in the culture supernatant fluid from monocultures of P. multocida or from P. multocida-M. haemolytica cocultures. These results indicate that P. multocida inhibits the growth of M. haemolytica by a contact- or proximity-dependent mechanism. If the inhibition of growth of M. haemolytica by P. multocida occurs in vivo as well, it could explain the inconsistent isolation of M. haemolytica from the lungs of pneumonic BHS.  相似文献   

7.

Aims

This study aimed to characterize the impact of lytic and temperate bacteriophages on the genetic and phenotypic diversity of Mannheimia haemolytica from feedlot cattle.

Methods and Results

Strictly lytic phages were not detected from bovine nasopharyngeal (n = 689) or water trough (n = 30) samples, but Myoviridae‐ or Siphoviridae‐like phages were induced from 54 of 72 M. haemolytica strains by mitomycin C, occasionally from the same strain. Phages with similar restriction fragment length polymorphism profiles (RFLP ≥70% relatedness) shared common host serotypes 1 or 2 (< 0·000 1). Likewise, phages with similar RFLP tended to occur in genetically related host bacteria (70–79% similarity). Host range assays showed that seven phages from host serotypes 1, 2 and 6 lysed representative strains of serotypes 1, 2 or 8. The genome of vB_MhM_1152AP from serotype 6 was found to be collinear with P2‐like phage φMhaA1‐PHL101.

Conclusions

Prophages are a significant component of the genome of M. haemolytica and contribute significantly to host diversity. Further characterization of the role of prophage in virulence and persistence of M. haemolytica in cattle could provide insight into approaches to control this potential respiratory pathogen.

Significance and Impact of the Study

This study demonstrated that prophages are widespread within the genome of M. haemolytica isolates and emphasized the challenge of isolating lytic phage as a therapeutic against this pathogen.  相似文献   

8.
The molecular evolution of the leukotoxin structural gene (lktA) of Mannheimia (Pasteurella) haemolytica was investigated by nucleotide sequence comparison of lktA in 31 bovine and ovine strains representing the various evolutionary lineages and serotypes of the species. Eight major allelic variants (1.4 to 15.7% nucleotide divergence) were identified; these have mosaic structures of varying degrees of complexity reflecting a history of horizontal gene transfer and extensive intragenic recombination. The presence of identical alleles in strains of different genetic backgrounds suggests that assortative (entire gene) recombination has also contributed to strain diversification in M. haemolytica. Five allelic variants occur only in ovine strains and consist of recombinant segments derived from as many as four different sources. Four of these alleles consist of DNA (52.8 to 96.7%) derived from the lktA gene of the two related species Mannheimia glucosida and Pasteurella trehalosi, and four contain recombinant segments derived from an allele that is associated exclusively with bovine or bovine-like serotype A2 strains. The two major lineages of ovine serotype A2 strains possess lktA alleles that have very different evolutionary histories and encode divergent leukotoxins (5.3% amino acid divergence), but both contain segments derived from the bovine allele. Homologous segments of donor and recipient alleles are identical or nearly identical, indicating that the recombination events are relatively recent and probably postdate the domestication of cattle and sheep. Our findings suggest that host switching of bovine strains from cattle to sheep, together with inter- and intraspecies recombinational exchanges, has played an important role in generating leukotoxin diversity in ovine strains. In contrast, there is limited allelic diversity of lktA in bovine strains, suggesting that transmission of strains from sheep to cattle has been less important in leukotoxin evolution.  相似文献   

9.
Large Pseudomonas phages isolated from barley rhizosphere   总被引:1,自引:0,他引:1  
Abstract: Five bacteriophages infecting common fluorescent pseudomonads ( Pseudomonas fluorescens and Pseudomonas putida ) were isolated from barley rhizosphere soil. Morphological and molecular characteristics of the phages are described together with selected phage-host interactions. All phages belonged to the Myoviridae family with isometrical heads on contractile tails; 4 of them were unusually large and had complex protein and DNA profiles. The large phages had estimated genome sizes of 200 kb or more. Restriction enzyme analyses and DNA-DNA hybridizations showed that all isolates represented different phage species. None of the isolates were observed to establish lysogeny with the main host strain, P. putida MM1. The large phages multiplied slowly on their hosts, producing very small plaques; one-step growth experiments with one of the large phages (Psp 4) hence demonstrated a long latent period (2.5 h) and a very small burst size (10 particles). One of the large phages (Psp 3) was abundant in the rhizosphere (approx. 104 pfu g−1 soil) and had a particularly broad host range which extended to both fluorescent ( Pseudomonas aeruginosa, P. fluorescens, P. putida and Pseudomonas chlororaphis ) and non-fluorescent (Pseudomonas stutzeri) Pseudomonas spp. occurring in soil. The ecological importance of the large Pseudomonas phages must be further studied, but their slow multiplication rates suggested a possible mechanism of balanced phage-host co-existence in the rhizosphere.  相似文献   

10.
The purpose of our study was to evaluate the inclusion of seven experimental phages into the international phage set for subtyping Listeria monocytogenes. The seven additional phages included the broad-host-range virulent Myoviridae phage A511 (M. J. Loessner, Appl. Environ. Microbiol. 57:1912-1918, 1991), three temperate phages from the Danish subsystem for typing serotype 1/2 strains (12682, 6223, and 5775) (P. Gerner-Smidt, V.T. Rosdahl, and W. Frederiksen, APMIS 101:160-167, 1993), and three temperate phages isolated by this laboratory in France (9425, 1313, and 197). A panel of 395 Listeria monocytogenes isolates (including 180 that were non-phage typeable by the international set) were used in the study for a comparison of the lytic spectra of the various bacteriophages. These results showed that the inclusion of five of the experimental phages contributed greatly to the overall typeability and discriminatory power of the system, especially for strains within serogroup 1/2.  相似文献   

11.
Seventeen virulent bacteriophages specific to Pseudomonas aeruginosa strains were isolated by screening various environmental samples. These isolated bacteriophages were grouped based on results obtained from restriction fragment analysis of phage genomes, random amplification of polymorphic DNA (RAPD) typing, morphology observations under transmission electron microscope, and host range analysis. All 17 bacteriophages are double-stranded DNA viruses and can be divided into 5 groups based on DNA restriction profiles. A set of 10-mer primers was used in RAPD typing of phages, and similar conclusions were obtained as for restriction fragment analysis. One phage was randomly selected from each of the 5 groups for morphology observations. Four of them had an icosahedral head with a long contractile tail, belonging to the Myoviridae family, and one phage had an icosahedral head with a short tail, thereby belonging to the Podoviridae family. Host range experiments were conducted on 7 laboratory strains and 12 clinical strains of P.?aeruginosa. The results showed that 13 phages had the same infection profile, killing 8 out of 19 tested P.?aeruginosa strains, and the remaining 4 phages had different and unique infection profiles. This study highlights the diversity of bacteriophages specific to P.?aeruginosa in the environment.  相似文献   

12.
Among dsDNA tailed bacteriophages (Caudovirales), members of the Myoviridae family have the most sophisticated virion design that includes a complex contractile tail structure. The Myoviridae generally have larger genomes than the other phage families. Relatively few "dwarf" myoviruses, those with a genome size of less than 50 kb such as those of the Mu group, have been analyzed in extenso. Here we report on the genome sequencing and morphological characterization of a new group of such phages that infect a diverse range of Proteobacteria, namely Aeromonas salmonicida phage 56, Vibrio cholerae phages 138 and CP-T1, Bdellovibrio phage φ1422, and Pectobacterium carotovorum phage ZF40. This group of dwarf myoviruses shares an identical virion morphology, characterized by usually short contractile tails, and have genome sizes of approximately 45 kb. Although their genome sequences are variable in their lysogeny, replication, and host adaption modules, presumably reflecting differing lifestyles and hosts, their structural and morphogenesis modules have been evolutionarily constrained by their virion morphology. Comparative genomic analysis reveals that these phages, along with related prophage genomes, form a new coherent group within the Myoviridae. The results presented in this communication support the hypothesis that the diversity of phages may be more structured than generally believed and that the innumerable phages in the biosphere all belong to discrete lineages or families.  相似文献   

13.
Lysogeny has long been proposed as an important long-term maintenance strategy for autochthonous soil bacteriophages (phages). Whole genome sequence data indicate that prophage-derived sequences pervade prokaryotic genomes, but the connection between inferred prophage sequence and an active temperate phage is tenuous. Thus, definitive evidence of phage production from lysogenic prokaryotes will be critical in determining the presence and extent of temperate phage diversity existing as prophage within bacterial genomes and within environmental contexts such as soils. This study optimized methods for systematic and definitive determination of lysogeny within a collection of autochthonous soil bacteria. Twenty bacterial isolates from a range of Delaware soil environments (five from each soil) were treated with the inducing agents mitomycin C (MC) or UV light. Six isolates (30%) carried inducible temperate phages as evidenced by an increase in virus direct counts. The magnitude of induction response was highly dependent upon specific induction conditions, and corresponding burst sizes ranged from 1 to 176. Treatment with MC for 30 min yielded the largest induction responses for three of the six lysogens. Morphological analysis revealed that four of the lysogens produced lambda-like Siphoviridae particles, whereas two produced Myoviridae particles. Additionally, pulsed-field gel electrophoresis data indicated that two of the six lysogens were polylysogens, producing more than one distinct type of phage particle. These results suggest that lysogeny is relatively common among soil bacteria.  相似文献   

14.
The bioluminescence assay system using Vibrio harveyi reporter strains were used to examine quorum-sensing autoinducer (AI) activity from Mannheimia haemolytica A1 cell-free culture supernatant. We showed that M. haemolytica A1 cell-free culture supernatant contains molecules that can stimulate the quorum-sensing system that regulates the expression of the luciferase operon in V. harveyi. Specifically, M. haemolytica A1 can stimulate only the quorum system 2 but not system 1, suggesting that the culture supernatant only contains molecules similar to AI-2 of V. harveyi. The bioluminescence assay was also used to show that culture supernatants from related Pasteurellaceae organisms, Pasteurella multocida, Pasteurella trehalosi, Actinobacillus suis and Actinobacillus pleuropneumoniae, also contain AI-2-like molecules. This is consistent with the presence of a luxS homolog in the genomes of P. multocida and A. pleuropneumoniae. A luxS homolog was cloned by PCR from M. haemolytica A1 using sequencing data from the ongoing genome sequencing project. The cloned luxS(M.h.) was able to complement AI-2 production in the Escherichia coli DH5alpha luxS mutant. This is the first report of a quorum-sensing activity in M. haemolytica A1 and suggests that this bacterium utilizes this mechanism to regulate expression of genes under specific conditions.  相似文献   

15.
In recent years, antimicrobial-resistant Pseudomonas aeruginosa strains have increased in the veterinary field. Therefore, phage therapy has received significant attention as an approach for overcoming antimicrobial resistance. In this context, we isolated and characterized four Pseudomonas bacteriophages. Phylogenetic analysis showed that the isolated phages are novel Myoviridae Pbunavirus PB1-like phages with ØR12 belonging to a different clade compared with the other three. These phages had distinct lytic activity against 22 P. aeruginosa veterinary isolates. The phage cocktail composed from the PB1-like phages clearly inhibited the occurrence of the phage-resistant variant, suggesting that these phages could be useful in phage therapy.  相似文献   

16.
Study of two recently isolated giant bacteriophages Lu11 and OBP that are active on Pseudomonas putida var. Manila and Pseudomonas fluorescens, respectively, demonstrated their similarity in morphology, genome size, and size of phage particles, with giant bacteriophages of Pseudomonas aeruginosa assigned to the supergroup of phiKZ-like phages of the family Myoviridae designated in this manner according to the best studied phage phiKZ that belongs to the species of this group widely distributed in nature. Comparison of major polypeptide sizes of mature particles suggests the similarity of certain proteins in the phages examined. In OBP particles visualized with an electron microscope, an "inner body" was detected, which points to the specific DNA package intrinsic to phages of phiKZ group. In the meantime, phages Lul11 and OBP do not exhibit resemblance among themselves or with any of earlier described phiKZ-like phages in respect to other traits; particularly, they have no detectable DNA homology. Note that phage Lu11 of P. putida var. Manila exhibits very slight homology with phage Lin68 of the family of P. aeruginosa phiKZ-like phages detected only in blot hybridization. This suggests the possible involvement of these phages in interspecies recombination ("gene shuffling") between phages of various bacterial species. Results of partial sequencing of phage genomes confirmed the phylogenetic relatedness of phage OBP to phages of the phiKZ-supergroup, whereas phage Lu11 most probably belongs to a novel species that is not a member of supergroup phiKZ composition. The results of the study are discussed in terms of the evolution of these phages.  相似文献   

17.
18.
Mannheimia haemolytica is a leading causative agent of pasteurellosis in ruminants. Genome of M. haemolytica strains from different hosts has been sequenced worldwide to understand its pathogenesis. There are only few reports on the isolation of M. haemolytica in India with limited information on its molecular characteristics. The present study focuses on genome sequence analysis of a M. haemolytica strain isolated from pneumonic sheep. Mannheimia haemolytica A2 strain NIVEDI/MH/1 was isolated and identified by species and serotype-specific PCRs. Whole genome sequencing was performed using the Ion Torrent Personal Genome Machine. A comparative genomic analysis was performed to understand the virulence determinants of the Indian strain and its phylogenetic relationship with other global strains. Sequence data revealed a draft genome of 2,211,426 bp size with 41.3% GC content, assembled into 17 contigs, and contained 2379 genes. Five genomic islands identified in the genome showed high sequence identity with other respiratory pathogens of the Pasteurellaceae family. Phylogenetic analysis showed M. haemolytica A2 NIVEDI/MH/1 is very close to a M. haemolytica A2 strain from pneumonic calf. Further, the analysis revealed the presence of virulence, metal-, and multidrug resistance genes needed for pathogenesis and survival of the bacteria during infection. Also, we identified the presence of type I-C and type II-C of CRISPR-Cas arrays in the present sequenced genome. The study emphasizes the role of M. haemolytica in respiratory infections of ruminants in the Indian subcontinent and indicates the role of vertical and horizontal gene pools in pathogenicity and survivability of the bacteria.  相似文献   

19.
Study of two recently isolated giant bacteriophages Lu11 and OBP that are active on Pseudomonas putida var. Manila and Pseudomonas fluorescens, respectively, demonstrated their similarity in morphotype, genome size, and size of phage particles, with giant bacteriophages of Pseudomonas aeruginosa assigned to the supergroup of ?KZ-like phages of the family Myoviridae. This supergroup was designated in this manner according to the best studied phage ?KZ that belongs to the species of this group widely distributed in nature. Comparison of major polypeptide sizes of mature particles suggests similarity of certain proteins in the phages examined. In OBP particles visualized with an electron microscope, an “inner body” was detected, which points to specific DNA package intrinsic to phages of ?KZ group. In the meantime, phages Lu11 and OBP do not exhibit resemblance among themselves or with any of earlier described ?KZ-like phages in respect to detectable DNA homology. Note that phage Lu11 of P. putida var. Manila exhibits very slight homology with phage Lin68 of the family of P. aeruginosa ?KZ-like phages detected only in blot hybridization. This suggests the possible involvement of these phages in interspecies recombination (“gene shuffling”) between phages of various bacterial species. Results of partial sequencing of phage genomes confirmed the phylogenetic relatedness of phage OBP to phages of the ?KZ supergroup, whereas phage Lu11 most probably belongs to a novel species that is not a member of supergroup ?KZ composition. The results of the study are discussed in terms of the evolution of these phages.  相似文献   

20.
Flavobacterium psychrophilum is an important fish pathogen worldwide that causes cold water disease (CWD) or rainbow trout fry syndrome (RTFS). Phage therapy has been suggested as an alternative method for the control of this pathogen in aquaculture. However, effective use of bacteriophages in disease control requires detailed knowledge about the diversity and dynamics of host susceptibility to phage infection. For this reason, we examined the genetic diversity of 49 F. psychrophilum strains isolated in three different areas (Chile, Denmark, and USA) through direct genome restriction enzyme analysis (DGREA) and their susceptibility to 33 bacteriophages isolated in Chile and Denmark, thus covering large geographical (>12,000 km) and temporal (>60 years) scales of isolation. An additional 40 phage-resistant isolates obtained from culture experiments after exposure to specific phages were examined for changes in phage susceptibility against the 33 phages. The F. psychrophilum and phage populations isolated from Chile and Denmark clustered into geographically distinct groups with respect to DGREA profile and host range, respectively. However, cross infection between Chilean phage isolates and Danish host isolates and vice versa was observed. Development of resistance to certain bacteriophages led to susceptibility to other phages suggesting that “enhanced infection” is potentially an important cost of resistance in F. psychrophilum, possibly contributing to the observed co-existence of phage-sensitive F. psychrophilum strains and lytic phages across local and global scales. Overall, our results showed that despite the identification of local communities of phages and hosts, some key properties determining phage infection patterns seem to be globally distributed.  相似文献   

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