Effect of ColV plasmids on the hydrophobicity of Escherichia coli

Abstract The hydrophobicity of E. coli strains carrying or lacking the colicin V ( ColV ) plasmids, ColV , I-K94 or ColV -K30 was assayed. ColV ⁺ derivatives of strain 1829, produced by conjugation or transformation, were more hydrophobic than either the original 1829 parental strain or a Col ^- derivative formed by curing 1829 ColV -K30 of its plasmid by an SDS/high temperature growth technique. Transfer of ColV into other E. coli strains also led to increased hydrophobicity. This effect of ColV plasmids was observed for organisms grown at 37°C; ColV ⁺ and ColV^- strains did not differ in hydrophobicity of grown at 21°C. This finding and other studies suggest that sex pili may be involved in the increased hydrophobicity.     

Alkali sensitivity in plasmid-bearing strains of Escherichia coli

Introduction of the ColV,I-K94 plasmid into any of four strains of Escherichia coli gave derivatives which grew less well than the parent at alkaline pH. The alkali sensitivity of the 1829 derivative resulted from the presence of the plasmid rather than from its introduction into a less alkali tolerant variant. Of two other ColV plasmids tested, one (ColV-K30) conferred substantial alkali sensitivity whereas the other (ColV-41) had little effect. Of several other plasmids examined, R124-F2 (which confers derepressed transfer properties) resembled ColV,I-K94 in its effect on alkali sensitivity and ColV-K98 produced a marked effect. The other plasmids (F lac , R124 itself, R1 and R483ColIa) had only a small effect. For ColV plasmids, it appears to be the presence of transfer and colicin components together which leads to the reduced tolerance to alkaline pHs.     

Virulence plasmid-associated sensitivity to rifampicin and novobiocin in Escherichia coli

Introduction of the ColV, I-K94 or ColV-K30 plasmid into Escherichia coli K12 resulted in increased sensitivity to growth inhibition and killing by rifampicin and novobiocin. For increased rifampicin sensitivity, both transfer and colicin components had to be present whereas transfer components plus another unidentified ColV component were needed for novobiocin sensitivity. The Co1 factor Co1B-K98 conferred increased rifampicin sensitivity, an effect not dependent on transfer components, but this plasmid had no effect on the novobiocin sensitivity of E. coli. The effects of ColV, I-K94 on rifampicin sensitivity were fully reversed by magnesium ions but Mg²⁺ had only a slight effect on the novobiocin inhibition of the Co1V, I-K94⁺ strain and the rifampicin inhibition of the Co1B-K98⁺ strain. Prior growth at 25C greatly reduced the effects of Co1V, I-K94 on rifampicin and novobiocin sensitivity but had essentially no effect on rifampicin sensitivity of the Co1B-K98⁺ strain.     

The ColV, I-K94 plasmid and heat sensitivity of Escherichia coli

The ColV, I-K94 plasmid markedly increased the heat sensitivity of Escherichia coli K12 but had no sensitizing effect on a wild E. coli isolate. The plasmid effect on strain K12 appeared to result from ColV-encoded transfer and colicin components possibly due to their effects on membrane properties.     

Suppression by the ColV,I-K94 plasmid of a growth lesion in ompA mutants of Escherichia coli

Organisms of three independently isolated ompA mutants of Escherichia coli failed to form colonies on glucose minimal agar (glucose MA) at 44 degrees C after growth in glucose minimal salts medium at 37 degrees C, although all three strains formed colonies on nutrient agar at 44 degrees C. Supplementation of the glucose MA with individual amino acids including L-methionine and/or L-cysteine did not allow colony formation at 44 degrees C, although addition of 0.1% Casamino acids was effective; replacement of glucose with other energy sources or ammonium ions with glutamate also did not allow growth at 44 degrees C. The failure to form colonies at 44 degrees C was not due to killing of the organisms, because colonies were formed if plates of the ompA mutant initially incubated at 44 degrees C were shifted to 30 degrees C after 16 h. Introduction of the ColV, I-K94 plasmid into P678-54 ompA, 1131 ompA or an ompC ompA mutant suppressed the 44 degrees C growth lesion, but other plasmids (F lac, R483ColIa, RI, ColB-K98, R124) tested in P678-54 ompA did not. Growth of the ColV, I-K94+ derivative at 44 degrees C was due to a suppressing effect of the plasmid rather than to introduction of the plasmid into a variant with normal or altered OmpA protein. An attempt was made to ascertain which component(s) encoded by ColV, I-K94 was (were) responsible for allowing growth at 44 degrees C. Transfer components appeared unlikely to be involved and plasmids which conferred individual colicins (plus the corresponding immunity component) did not suppress.(ABSTRACT TRUNCATED AT 250 WORDS)     

Sequence rearrangements in the plasmid ColV,I-K94

ColV,I-K94 is a conjugative (F-like) plasmid specifying colicins V and I. Numerous rearrangements were observed in ColV,I-K94 plasmids from three culture collections and in derivatives which specified larger inhibition zones and increased titers of colicin V. Several of the rearrangements in ColV,I-K94 derivatives specifying higher titers of colicin V involved a 1.3 ± 0.08-kb inverted repeat sequence which was separated by a 3.3 ± 0.2-kb sequence. This was rearranged in one derivative, pKH46, such that the 3.3-kb sequence became flanked by a 2.5 ± 0.15-kb inverted repeat which was subsequently altered to form a 2.1 ± 0.1-kb inverted repeat. In two other derivatives, the 3.3-kb sequence was deleted together with all of the flanking inverted repeat (in pKH39-2) or part of each repeat (in pKH39-1). A 0.5 ± 0.1-kb sequence adjacent to one of the 1.3-kb repeats was inverted or substituted in both of these plasmids. A novel 12.9 ± 0.7-kb inverted duplication separated by a 7.3 ± 0.4-kb sequence was present in the ColV,I-K94 derivative, pKH41, and insertion of the γδ element was detected in another derivative, pKH46-1, after this nonconjugative plasmid had been mobilized by the F plasmid. The genes for colicin V and for immunity to colicin V were mapped to a 3.6-kb sequence. The copy numbers of both pKH39-2 and pKH46-1::γδ were greater than those of pKH38 from which they were derived.     

质粒在大肠杆菌对噬菌体抗性中的作用

The introduction of the ColV, I-K94 or R124 plasmid into Escherichia coli K12 resulted in resistance to certain phages. Derivatives of E. coli carrying the plasmid R124 and ColV, I-K94 were resistance to the phages T4, Mel comparing with the plasmid-free parent and the plasmid ColV, I-K94 conferred resistance to the phage Tull*. It suggested that an envelope change caused by the plasmids might be responsible for the resistance because most of the phages fell to absorb to the plasmid-bearing E. coli cells.     

Suppression by the ColV,I-K94 plasmid of inhibitor sensitivities in ompA mutants of Escherichia coli

A series of ompA mutants derived from Escherichia coli K12 strains showed increased sensitivity (compared with the ompA+ parents) to aminoglycoside antibiotics and to other cationic agents including polymyxin B. One tested mutant also showed increased sensitivity to nafcillin and fusidic acid, but not to the hydrophilic ampicillin. All these inhibitor sensitivities in the ompA mutants were suppressed by ColV, I-K94 and by certain other ColV plasmids, but not by any of the other tested large plasmids. Suppression correlated with the production of the VmpA protein, but transfer and colicin components were not needed for suppression. Further comparison of the ompA and vmpA genes and their products was made and it indicated that there is little if any homology between the genes, that the synthesis of their products is regulated by quite different mechanisms, and that regions of these gene products exposed at the cell surface show different susceptibility to protease attack after denaturation.     

Decreased DNA damage by acid and increased repair of acid-damaged DNA in acid-habituated Escherichia coli

A study of the conjugal transfer of ColV,I-K94 tn 10 from acid-treated donors suggested that acid-habituated recipients repair acid-damaged plasmid DNA better than those that are not habituated. The presence of an increased repair activity for acid-damaged DNA in habituated cells was confirmed by isolating pBR322 from acid-treated organisms; habituated cells produced more transformants when transformed by it than did non-habituated ones. Additionally, agarose gel electrophoretic studies of pBR322 DNA isolated from acid-damaged cells and tests of its transforming activity both indicated that plasmid DNA in habituated cells is less damaged by extreme acidity than is that in non-habituated organisms.     

Decreased DNA damage by acid and increased repair of acid-damaged DNA in acid-habituated Escherichia coli

A study of the conjugal transfer of ColV,I-K94 tn10 from acid-treated donors suggested that acid-habituated recipients repair acid-damaged plasmid DNA better than those that are not habituated. The presence of an increased repair activity for acid-damaged DNA in habituated cells was confirmed by isolating pBR322 from acid-treated organisms; habituated cells produced more transformants when transformed by it than did non-habituated ones. Additionally, agarose gel electrophoretic studies of pBR322 DNA isolated from acid-damaged cells and tests of its transforming activity both indicated that plasmid DNA in habituated cells is less damaged by extreme acidity than is that in non-habituated organisms.     

Acid resistance of attached organisms and its implications for the pathogenicity of plasmid-bearing Escherichia coli

Organisms of Escherichia coli attached to glass beads in a model attachment system were more resistant to acid than were unattached organisms and this applied to cultures exposed to either pH 2·5 or 3·5. Attached organisms of both Col^- and Col V, I-K94⁺ strains showed the effect but with prolonged exposure to pH 2·5, the attached Col V⁺ organisms appeared more resistant than the attached Col^- ones, possibly because they formed a thicker surface layer. It is proposed that the increased resistance of attached organisms to pH 2·5 might allow the survival, in gastric acid, of organisms attached to food particles. This would be more significant for the Col V⁺ strains because the plasmid enhances enterobacterial attachment properties. The increased resistance of attached organisms to pH 3·5 might be significant for survival of particle- or surface-bound organisms in the acidic environment of the phagocyte especially since lactate (which occurs in phagocytes) enhanced the effect of exposure to pH 3·5.     

Habituation to alkali and increased u.v.-resistance in DNA repair-proficient and -deficient strains of Escherichia coli grown at pH 9.0

Repair-deficient strains of Escherichia coli carrying polAI or recA mutations were more alkali-sensitive than was their repair-proficient parent but, like strain 1829 ColV, I-K94, they showed habituation to alkali (induction of increased resistance) when grown at pH 9.0. Occurrence of such increased alkali resistance in the recA mutant implies that habituation to alkali does not depend on induction of SOS-related repair mechanisms. Organisms of repair-proficient and repair-deficient strains also became more resistant to u.v.-irradiation after growth at pH 9.0; this increased u.v.-resistance also appeared to be RecA-independent.     

RecA-independent resistance to irradiation with u.v. light in acid-habituated Escherichia coli

Growth of Escherichia coli 1829 ColV, I-K94 at pH 5.0 led to an increase in u.v. resistance compared with cells grown at pH 7.0. This was due to a phenotypic change, since organisms grown at pH 7.0 showed increased resistance after only 2.5–5.0 min incubation at the mildly acid pH. Other E. coli K12 derivatives became more u.v.-resistant at pH 5.0 including uvrA, recA and polA1 mutants. Organisms grown at pH 5.0 also showed increased Weigle reactivation of u.v.-irradiated Λ phage and this applied to the repair-deficient mutants as well as the parent strains. Both the increased u.v. resistance of acid-habituated cells and their increased ability to bring about Weigle reactivation appear to involve RecA-independent processes and are presumably, therefore, independent of the SOS response.     

RecA-independent resistance to irradiation with u.v. light in acid-habituated Escherichia coli

Growth of Escherichia coli 1829 ColV, I-K94 at pH 5.0 led to an increase in u.v. resistance compared with cells grown at pH 7.0. This was due to a phenotypic change, since organisms grown at pH 7.0 showed increased resistance after only 2.5-5.0 min incubation at the mildly acid pH. Other E. coli K12 derivatives became more u.v.-resistant at pH 5.0 including uvrA, recA and polA1 mutants. Organisms grown at pH 5.0 also showed increased Weigle reactivation of u.v.-irradiated lambda phage and this applied to the repair-deficient mutants as well as the parent strains. Both the increased u.v. resistance of acid-habituated cells and their increased ability to bring about Weigle reactivation appear to involve RecA-independent processes and are presumably, therefore, independent of the SOS response.     

Copper sensitivity in an envelope mutant of Escherichia coli and its suppression by ColV, I-K94

A phage K3-resistant isolate from Escherichia coli P678-54 was devoid of both the OmpA and OmpC proteins but had high levels of the OmpF protein. Associated with these changes, the strain showed increased sensitivity to inhibition by detergents and greatly increased sensitivity to Cu²⁺. Introduction of the ColV, I-K94 plasmid into this mutant produced a derivative with markedly increased resistance to Cu²⁺ ions but unchanged detergent sensitivity. Analysis of membranes showed that the ColV, I-K94⁺ derivative had essentially no OmpF protein in its outer membrane. A ca 36 K outer membrane protein was present which resembled the OmpC protein in size and failure to dissociate in SDS at low temperature. It was distinct from the OmpC protein, however, in failing to allow either tetracycline uptake or the adsorption of T4-type phages. The possible significance of OmpF porin derepression (and its reversal by ColV, I-K94) for enterobacterial survival in aquatic situations is discussed.     

Temperature-Sensitive Mutants for the Replication of Plasmids in ESCHERICHIA COLI II. Properties of Host and Plasmid Mutations

Host mutations in Escherichia coli K12 selected for the temperature-sensitive replication of the bacterial plasmid colicinogenic factor E(1) (ColE(1)) exhibit a pleiotropic effect with respect to the effect of the mutation on other extra-chromosomal elements. The mutations also vary with respect to the time of incubation of the cells at 43 degrees C required for complete cessation of ColE(1) DNA synthesis. While the synthesis of the bacterial chromosome appears unaffected, supercoiled ColE(1) DNA replication stops immediately in some mutants and gradually decreases during several generations of cell growth before stopping in others. Mutations isolated in the ColE(1) plasmid resulted in only a gradual cessation of ColE(1) DNA synthesis over several generations of cell growth at 43 degrees C. Conjugal transfer of the ColE(1) and ColV factors occurs normally in the host mutants when the transfer is carried out at the permissive temperature; however, the presence of a group I mutation in the donor cell prohibited conjugal transfer of either plasmid DNA at 43 degrees C to a normal recipient cell. Similarly, the presence of this mutation in the recipient prevented the establishment of ColE(1) or ColV in the mutant recipient cell upon conjugation with a normal donor at 43 degrees C. Various host ColE(1) replication mutants carrying either ColE(1) or ColE(2) were also defective in the mitomycin C-induced production of colicin E(1) or colicin E(2) at 43 degrees C. The majority of the host mutations examined exhibited a temperature sensitivity to growth in deoxycholate in addition to the inhibition of plasmid DNA replication, suggesting a membrane alteration in these mutants when grown at the restrictive temperature.     

Habituation to acid in Escherichia coli: conditions for habituation and its effects on plasmid transfer.

Induction of acid resistance (habituation) in Escherichia coli at pH 5.0 took ca 5 min in broth at 37 degrees C and 30-60 min in minimal medium. Induction occurred at a range of pH values from 4.0 to 6.0; it was dependent on continuing protein and RNA synthesis but substantial acid resistance appeared in the presence of nalidixic acid. Acid resistance was long-lasting; organisms grown at pH 5.0 retained most of their resistance after 2 h growth at pH 7.0. Organisms grown at pH 5.0 showed increased synthesis of a number of cytoplasmic proteins compared with the level in cells grown at pH 7.0. DNA repair-deficient strains carrying recA, uvrA or polA1 mutations were more acid-sensitive than the repair-proficient parents but were able to habituate at pH 5.0. Organisms grown at pH 5.0 transferred the ColV plasmid much more effectively at acid pH than did those grown at pH 7.0 and habituated recipients appeared better able to repair incoming acid-damaged plasmid DNA than did those that were non-habituated. Induction of acid resistance at pH 5.0 may be significant for the survival of organisms exposed to periodic discharges of acid effluent in the aquatic environment and habituation may also allow plasmid transfer and repair of acid-damaged plasmid DNA during or after such exposure.     

Characteristics of a colicin plasmid in Escherichia coli strain B177 isolated from a septicemic calf

A colicin plasmid in Escherichia coli strain B177 isolated from a septicemic calf was characterized. The colicin type was identified as ColV by using reference ColV producers. The colicin plasmid was labeled with transposon Tn903 and subjected to conjugation. The transconjugants examined suggest that the colicin plasmid confers serum resistance. There was no difference in siderophore utilization ability between the transconjugants and host strain SF800. Bioassay for siderophore suggests that the colicin plasmid specifies the production of iron-chelating compounds available for the host strain.     

A Comparative Study of Two F-like Colicin Factors, ColV2 and ColV3, in Escherichia coli K-12

Of three colicin factors, each determining the synthesis of a colicin V in three wild-type Escherichia coli strains studied, two were shown to have sex-factor activities. In E. coli K-12, these activities resembled those of the F sex factor (including rapid and efficient self-transmission in exponentially growing cultures, adsorption of "male-specific" ribonucleic acid phage, production of "female phenocopies," elimination by acridine orange, and chromosomal transfer dependent upon recombination with host bacterium) and differed in this way from those of the colicin I sex factor (ColI). The two V factors, ColV2 and ColV3, differed in their efficiency of plating male-specific phage and in the pattern of transfer of chromosomal markers. Furthermore, although neither factor could stably coexist with F within the same cell, they showed markedly different exclusion effects. In general, ColV2 excluded F and ColV3 was excluded by F, irrespective of which sex factor was preestablished in the cell. An exception to this was the ability of ColV2 to stabilize in any one of a series of Hfr strains, giving rise to strains which in the majority of cases showed normal Hfr and colicinogenic properties.     

Habituation to normally lethal acidity by prior growth of Escherichia coli at a sub-lethal acid pH value

Both Col^- and ColV, I-K94⁺ strains of Escherichia coli , grown at pH 7–0, failed to grow after relatively short periods of exposure to pH 3·0 or 3·5. After growth in exposure medium initially at pH 5·0, both strains were almost unaffected by exposure to such acid pH values. Addition of catalase to nutrient agar only slightly increased plating efficiency after acid treatment and very slightly reduced the difference in survival, after acid treatment, between organisms grown from pH 5·0 and those grown from pH 7·0. Accordingly, acid resistance of organisms grown from pH 5·0 is not chiefly due to greater resistance to hydrogen peroxide already present in nutrient media.