Homology-facilitated plasmid transfer in Haemophilus influenzae

Abstract

The 8 kbp plasmid pAT4 transformed Haemophilus influenzae Rd cells at low frequencies. Transformation was increased up to 100 times, however, when the recipient cells carried a DNA segment in either their chromosome or in a resident plasmid that was homologous to at least part of plasmid pAT4. Linearized plasmid DNA molecules did not transform cells without DNA homology; they efficiently transformed homology recipients, but only when the cuts had been made in the region of shared homology. In most cases examined the circular donor plasmid had been reconstituted from the transforming DNA; in some cases the reconstituted plasmid carried a mutation initially present in the recipient chromosome, provided the transforming plasmid had been linearized in the region of shared homology. Plasmid reconstitution was not observed in recA1 cells. We conclude that homology-facilitated plasmid transformation (transfer) is similar to that reported for Bacillus subtilis and Streptococcus pneumoniae.

Plasmid Transformation of Azotobacter vinelandii OP

“…The frequency of pKT210 transformation of strain OP was two orders of magnitude lower than the observed plasmid transformation frequency of naturally competent H. injluenzae (Stuy & Walter, 1986), B. subtilis (Contente & Dubnau, 1979) or S. sanguis (Behnke, 1981), indicating that naturally competent A. vinelandii discriminated against heterologous plasmid DNA transformation more than these other bacteria. As was found with H. injluenzae, competent A .…”

“…The phenomenon of homology-facilitated transformation has been well studied in several eubacterial natural transformation systems. The very low frequency of transformation of H. injluenzae by linear plasmid DNA (one transformant per lo7 plasmid molecules; Stuy & Walter, 1986) appears to be a result of the requirement that homologous DNA sequences be present in the recipient cell and the transforming plasmid (Stuy & Walter, 1986). This is certainly a consequence of the fact that plasmid DNA is converted to a single-stranded form during uptake Pifer & Smith, 1985).…”

Cloning, characterization, and DNA base sequence of the high-level streptomycin resistance gene strA1 of Haemophilus influenzae Rd

“…A rec-1 mutant (15) was given to us by J. Setlow. Plasmid pAT4 is a 7.5-kb Ampr Tetr Haemophilus plasmid (22) (Fig. 1).…”

“…Standard methods have been described (18)(19)(20)(21)23). E. coli cells were made competent by the CaCl2 technique (12).…”

Haemophilus influenzae: antibiotic susceptibility

“…Although transformation is known to occur in Haemophilus spp., the efficiency of this mechanism of genetic exchange is very low in the absence of homologous plasmid DNA sequences (1). However, the conjugative resistance plasmids are not infrequently integrated into the chromosome (102 Non-beta-lactamase-producing strains of ampicillin-resistant H. influenzae are also known to occur (45,60,112 (122). However, the incidence of beta-lactamase-producing encapsulated strains had increased to 14% by 1981 in Great Britain (81).…”