Plasmid-linked ampicillin resistance in haempohilus influenza type b

Abstract

Four ampicillin-resistant, f,-lactamase-producing strains of Haemophilus influenzae type b were examined for the presence of plasmid deoxyribonucleic acid (DNA). Three resistant strains contained a 30 x 106-dalton (30 Mdal) plasmid and one resistant strain contained a 3-Mdal plasmid. The ampicillin-sensitive Haemophilus strains examined did not contain plasmid DNA. Transformation of a sensitive H. influenzae strain to ampicillin resistance with isolated plasmid DNA preparations revealed that the structural gene for ,B-lactamase resided on both plasmid species. DNA-DNA hybridization studies showed that the 30-Mdal Haemophilus plasmid contained the ampicillin translocation DNA segment (TnA) found on some R-factors of enteric origin. The significance of this finding is discussed in relation to the possible origin of the H. influenzae plasmids. Disease due to ampicillin-resistant strains of Haemophilus influenzae type b has become an increasingly serious and widespread problem (1, 3, 25). Ampicillin-resistant strains have been isolated from cases of epiglottitis, pneumonia, otitis media, and meningitis, including three fatal cases of infant meningitis (15, 17). The primary cause of ampicillin resistance appears to be the production of a d-lactamase shown in one strain to possess a broad-spectrum penicillinase-cephalosporinase activity (9). Thorne and Farrar (J. Infect. Dis., in press) recently have demonstrated transfer of the ampicillin resistance from one strain to an ampicillin-sensitive strain during mixed incubation, presumably by conjugation. The sudden emergence of ampicillin resistance in H. influenzae due to a transferable #-lactamase gene suggested to us, and others (9,15), that an R-factor might be involved. In this study we report the isolation of plasmid deoxyribonucleic acid (DNA) from four ampicillin-resistant strains of H. influenzae. Three strains contained a 30 x 108-dalton (30 Mdal) plasmid and one strain contained a 3.0-Mdal plasmid, and evidence is presented indicating that the ,3-lactamase gene is located on those plasmid species. Resistance to ampicillin mediated by the TEM-type ,B-lactamase in enteric bacteria is found on a segment of DNA that can be translocated from replicon to replicon (13). This same segment is found on the plasmid DNA isolated from resistant H. influenzae.

Heat-labile enterotoxin production in isolates from a shipboard outbreak of human diarrheal illness

“…Purified plasmid deoxyribonucleic acid (DNA) was isolated from toxigenic and nontoxigenic 025:NM strains by centrifugation of cleared lysates in CsClethidium bromide gradients. Cleared lysates were prepared as described by Elwell et al (7). Gradients were centrifuged at 35,000 rpm and 15°C for 40 to 48 h. Bacteriology.…”

Characterization of the cytochrome c gene from the starch‐fermenting yeast Schwanniomyces occidentalis and its expression in bakers's yeast

“…Large-scale (Elwell et at., 1975) and small-scale alkaline (Maniatis et al, 1982) plasmid preparations were carried out as described. Singlestranded DNA templates were prepared as described (Messing, 1983).…”

The Effect of Tetracycline upon the Spread of Bacterial Resistance from Calves to Man1

“…Isolates with resistance characteristics of R135 (resistant to gentamicin) were crossed in vitro with a rifampin resistant E. coli K12(J62). Plasmid DNA was extracted from E. coli K12(J62) and subjected to DNA homology studies (Elwell et al, 1975).…”

“…DNA tlomology Studies. The method used to determine homology between plasmids with a determinant coding for gentamicin resistance and R135 has been described elsewhere (Elwell et al, 1975;Crosa et al, 1973). Briefly, tritium labelled R135 DNA (approximately 2,500 nucleotides in length) was mixed with plasmid DNA fragments (approximately 2,500 nucleotides in length) isolated from the human subjects.…”