Thymidine-Requiring Mutants of Phage T4

Simon, Edward H.; Tessman, Irwin

doi:10.1073/pnas.50.3.526

Cited by 140 publications

(72 citation statements)

References 10 publications

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“…A PhoA ¹ colony of SA202 was inoculated into a Tris-buffered medium (Simon and Tessman, 1963) lacking KH 2 PO 4 and containing 40 g ml ¹1 XP (5-bromo-4-chloro-3-indolyl phosphate; Sigma Chemical Co.) as the sole source of phosphate, to select for PhoA þ bacteria. phoA fusions to plasmid genes were selected by transforming an E. coli PhoA ¹ strain with the pooled plasmid DNA from the bacteria (SA202) that grew and selecting for Km R and PhoA þ phenotypes.…”

Section: Tnphoa Mutagenesismentioning

confidence: 99%

Effect of mutations in Shigella flexneri chromosomal and plasmid‐encoded lipopolysaccharide genes on invasion and serum resistance

Hong

Payne

1997

Molecular Microbiology

118

120

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SummaryThis study shows that both length and distribution of lipopolysaccharide (LPS) are important for Shigella flexneri invasion and virulence. Mutants were generated in the chromosomal LPS synthesis genes rfa, rfb, and rol, and in a plasmid-encoded O-antigen chain-length regulator, cld pHS-2 . LPS analysis showed that mutations in rfb genes and in a candidate rfaL gene either eliminated the entire O-antigen side chains or produced chains of greatly reduced length. Mutation in a previously unidentified gene, rfaX, affected the LPS core region and resulted in reduced amounts of O-antigen. Mutants defective in cld pHS-2 or rol had different distributions of O-antigen chain lengths. The results of tissue-culture cell invasion and plaque assays, the Seré ny test, and serum-sensitivity assay suggested roles for the different LPS synthesis genes in bacterial survival and virulence; rfaL, rfaX and rfb loci are required for serum resistance and intercellular spread, but not for invasion; cld pHS-2 is required for resistance to serum killing and for full inflammation in the Seré ny test, but not for invasion or intercellular spread, while rol is required for normal invasiveness and plaque formation, but not for serum resistance. Thus, O-antigen synthesis and chain-length regulation genes encoded on both the chromosome and the small plasmid pHS-2 play important roles in S. flexneri invasion and virulence.

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Section: Tnphoa Mutagenesismentioning

confidence: 99%

Effect of mutations in Shigella flexneri chromosomal and plasmid‐encoded lipopolysaccharide genes on invasion and serum resistance

Hong

Payne

1997

Molecular Microbiology

118

120

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“…coli EB53 cells were grown at 26 or 37 "C in nutrient broth with 85.6 mM NaCl (NB), NB supplemented with 50 pM of the protoporphyrin IX precursor 6-aminolevulinic acid (ALA) (Stojiljkovic & Hantke, 1992), or on NB containing 0-3 mM of the iron chelator 2,2'-dipyridyl (NBD) (Sigma) ; when appropriate, these media were solidified with 1.5 ' / o (w/v) Noble agar (Difco). E. coli SABll cells were cultivated at 26 or 37 "C in either LB broth or on solidified Tris/glucose/thymidine medium supplemented with 0-5 mM leucine and 1 mM proline (TGT) (Barghouthi et al, 1991;Simon & Tessman, 1963). TGT containing either 25 pM conalbumin (Sigma) or 5 pM haemin was solidified with 1.4% (w/v) Noble agar.…”

Section: Methodsmentioning

confidence: 99%

The haemin storage (Hms+) phenotype of Yersinia pestis is not essential for the pathogenesis of bubonic plague in mammals

LillardJr

Bearden²,

Fetherston³

et al. 1999

Microbiology

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The haemin storage (Hms+) phenotype of Yersinia pestis enables this bacillus to form greenish/brown or red colonies on haemin or Congo Red agar plates, respectively, at 26 but not 37 O C . Escherichia coli strains that contain mutations in genes essential for siderophore biosynthesis, porphyrin generation and/or haemin transport remain unable to utilize exogenous haemin as a nutritional iron or porphyrin source when transformed with the cloned Y. pestis hmsHFRS locus. Further physiological analysis of the Hms+ phenotype of Y. pestis strain KIM6+ suggests that the haemin and inorganic iron stored by the Hms system was not used nutritionally under subsequent iron-deficient conditions. In vitro analysis of the bactericidal effects of hydrogen peroxide, superoxide and nitric oxide showed that Hms' Y. pestis cells, in certain cases, were more susceptible than the Hms+ parent cells to these reactive oxygen species a t 26 and/or 37 "C.In adherence assays, a higher percentage of Hms+ cells were associated with HeLa cells and normal human neutrophils, compared to Hms' cells. However, the Hms+ phenotype did not provide any additional protection against the killing effects of neutrophils. Finally, LD, , analysis in subcutaneously infected mice showed that an Hms' strain was slightly more virulent than Hms+, indicating that the Hms phenotype is not essential for the pathogenesis of bubonic plague in mammals.

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“…Aerobactin was prepared from E. coli LG1315. Cells were grown for 48 to 72 h in Tris-succinate medium (T medium of Simon and Tessman [39] lacking glucose and FeCl3 and supplemented with 1% sodium suc- Williams (47) Williams (44) Williams (47) Williams (8) Braun (6) Braun (6,23) is study Neilands (37) is study…”

Section: Methodsmentioning

confidence: 99%

Aerobactin utilization by Neisseria gonorrhoeae and cloning of a genomic DNA fragment that complements Escherichia coli fhuB mutations

West¹,

Sparling²

1987

J Bacteriol

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Aerobactin, a dihydroxamate siderophore produced by many strains of enteric bacteria, stimulated the growth of Neisseria gonorrhoeae FA19 and F62 in iron-limiting medium. However, gonococci did not produce detectable amounts of aerobactin in the Escherichia coli LG1522 aerobactin bioassay. We probed gonococcal genomic DNA with the cloned E. coli aerobactin biosynthesis (iucABCD), aerobactin receptor (iutA), and hydroxamate utilization (flwCDB) genes. Hybridization was detected with IhuB sequences but not with the other genes under conditions which will detect 70% or greater homology. Similar results were obtained with 21 additional strains of gonococci by colony filter hybridization. A library of DNA from N. gonorrhoeae FA19 was constructed in the ph id vector XSE4, and a clone was isolated that complemented theihuB mutation in derivatives of E. coli BU736 and BN3307. These results suggest thatfJhuB is a conserved gene and may play a fundamental role in iron acquisition by N. gonorrhoeae.Most microbial iron acquisition systems consist of two components: a siderophore which is a soluble low-molecular-weight iron-chelating compound and a specific cell surface receptor for the iron-siderophore complex (33). Iron acquisition by Neisseria gonorrhoeae is different in that gonococci do not appear to produce siderophores (34, 45,46). However, gonococci, like Escherichia coli (33) and Salmonella typhimurium (26), can utilize siderophores produced by other microorganisms. Yancey and Finkelstein (48) demonstrated that aerobactin and the related dihydroxamate siderophores arthrobactin and schizokinen stimulate the growth of iron-starved gonococci.Aerobactin is produced by many strains of E. coli and several other species of gram-negative bacteria (Enterobacter [Aerobacter] aerogenes, Enterobacter cloacae, Shigella sonnei, Shigella boydii, Shigella flexneri, Salmonella arizona, Salmonella austin, and Salmonella memphis) (16,25,29,35). The aerobactin operon in E. coli consists of the genes which code for the 74,000-molecular-weight (74K) aerobactin receptor (iutA) and for four enzymes required for aerobactin biosynthesis (iucABCD [aerDBCA]) (2, 8). The proteins IucA, IucB, IucC, and IucD catalyze the synthesis of aerobactin from citrate and lysine (17). The aerobactin biosynthesis and receptor genes are arranged in a cluster flanked by ISI-like inverted repeat sequences (29) and may be either chromosomally or plasmid coded (29). In addition, several chromosomally located genes, fhuBCD, are required for uptake of iron from aerobactin and other hydroxamate siderophores such as ferrichrome, rhodotorulic acid, and coprogen (6,7,15,23,37). The fluBCD genes and theflzuA gene, which codes for the ferrichrome receptor, constitute a single operon located at 3.5 min on the E. coli chromosome; the gene order is JhuACDB (15 product is apparently a very hydrophobic membrane protein of 70,337 Mr as calculated from the nucleotide sequence (23); its exact location in the membranes of E. coli is uncertain.The mechanism of aerobactin-mediat...

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Thymidine-Requiring Mutants of Phage T4

Abstract: Infection with phage T2 can overcome the thymine requirement of a mutant of Escherichia coli.' Specifically, infection induces a thymidylate synthetase that is

Cited by 140 publications

References 10 publications

Effect of mutations in Shigella flexneri chromosomal and plasmid‐encoded lipopolysaccharide genes on invasion and serum resistance

Effect of mutations in Shigella flexneri chromosomal and plasmid‐encoded lipopolysaccharide genes on invasion and serum resistance

The haemin storage (Hms+) phenotype of Yersinia pestis is not essential for the pathogenesis of bubonic plague in mammals

Aerobactin utilization by Neisseria gonorrhoeae and cloning of a genomic DNA fragment that complements Escherichia coli fhuB mutations

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