Construction of intergeneric hybrids using bacteriophage P1CM: transfer of the Klebsiella aerogenes ribitol dehydrogenase gene to Escherichia coli

Abstract

Study of many of the interesting properties of Klebsiella aerogenes is limited by the lack of a well-characterized genetic system for this organism. Our investigations of the evolution of the enzyme ribitol dehydrogenasel?EC -1.1.1.56) in K. aerogenes would be greatly facilitated by the availability of such a system, and we here report two approaches to developing one. We have isolated mutants sensitive to the coliphage P1, which will efficiently transduce genetic markers between such sensitive strains and which will thus make detailed mapping studies possible. Derivatives of K. aerogenes lysogenic for P1 can be readily isolated by using the specialized transducing particle PlCMclrlOO. Bacteria lysogenic for this phage are chloramphenicol resistant and temperature sensitive. Phage particles produced by temperature induction of such lysogens can be used to transfer K. aerogenes genes to the natural host of P1 phage, Escherichia coli. We have used this method to prepare derivatives of E. coli K-12 carrying the K. aerogenes genes conferring the ability to metabolize the pentitols ribitol and D-arabitol. We have shown that these E. coli-K. aerogenes hybrids synthesize a ribitol dehydrogenase with the properties of the K. aerogenes enzyme and have mapped the position of the transferred gene on the E. coli chromosome. The ramifications of this methodology are discussed.

Genes for D-arabinitol and ribitol catabolism from Klebsiella pneumoniae

“…Hartley and co-workers originally transferred the genes from K . pneumoniae 1033-5P14, by means of P1-transduction, into the chromosome of E. coli K-12 where the genes inserted at a similar place (near 46 min) as in E. coli C (Rigby et al, 1976). Using a secondary att2 site near the pentitol-specific genes and a phage A derivative, Neuberger & Hartley (1979) cloned the dal and rbt genes onto phage Ap rbt dal.…”

“…2). This location may explain why Rigby et al (1976) were able to transduce the dal and r6t genes from K . pneurnoniae to E. coli K-12 by means of phage P1.…”

Expansion of the host range of coliphage P1 and gene transfer from enteric bacteria to other gram-negative bacteria

“…Moreover, the optimal multiplicity of infection is 0.1 to 0.5 for transduction between members of the same strain, whereas it is 5 to 10 for intergeneric transduction (14). P1CM or Plclrl00CM could be used to isolate P1-sensitive clones from bacteria that are resistant to kanamycin and P1 phage, as already achieved with M. xanthus (9), Pasteurella (13), and K. aerogenes (17).…”

“…Most of the bacterial strains used here are important in agriculture or in microbial industry: K. aerogenes W70 produces special enzymes, arylsulfatase (18), histidase (16), pentitol-degradating enzymes (17), or pullulanase (2,11); K.…”

Lysogenic conversion in Klebsiella pneumoniae: system which requires active immunity regulation for expression of the conversion phenomenon

“…Many examples of lysogenic conversion have already been described in various bacterial species (2, 4, 14-16, 18, 21-23). However, to our knowledge, lysogenic conversion has never been described in Klebsiella pneumoniae, a species related both physiologically and genetically to Escherichia coli (26,31), even though this microorganism has been intensively studied, particularly in view of its nitrogen-fixing capability (10,11,29), its pathogenicity (5,19,30), and its frequent antibiotic resistance (17).…”