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Biomedical subjects

A A Potter

Publications and source records attributed to A A Potter.

51 records · Page 3Linked to original sources
PubMed1989

Iron-repressible outer-membrane proteins of Pasteurella haemolytica.

The outer-membrane protein (OMP) profile of Pasteurella haemolytica grown under iron-replete and iron-restricted conditions was studied by polyacrylamide gel electrophoresis and immunoblotting. A serotype 1 isolate induced the synthesis of a new 77,000 Mr OMP in iron-restricted media while two other proteins of 100,000 Mr and 71,000 Mr were synthesized in increased amounts. None of these proteins were peptidoglycan-associated or heat-modifiable, and only the 100,000 Mr protein showed some degree of disulphide cross-linking. Kinetic analysis revealed that the iron-repressible proteins appeared in the outer membrane within 15 min of establishment of iron-restricted conditions. Analysis of P. haemolytica isolates representing serotypes 1 to 12 showed that iron-repressible OMPs of 77,000 Mr and 71,000 Mr could be induced in all 12 serotypes but that there was some variability in the expression of the 100,000 Mr protein. Immunoblotting of OMPs with convalescent sera from P. haemolytica-infected calves indicated that antibodies directed against all three iron-repressible OMPs were present, suggesting that these proteins were expressed in vivo.

Animals↗
PubMed1989

Effect of iron restriction on the outer membrane proteins of Actinobacillus (Haemophilus) pleuropneumoniae.

The outer membrane protein profile of Actinobacillus (Haemophilus) pleuropneumoniae grown under iron-restricted and iron-replete conditions was studied by polyacrylamide gel electrophoresis and immunoblotting. A virulent serotype 1 isolate synthesized a novel protein with an apparent molecular weight of 105,000 (105K) and increased the synthesis of a 76K protein under iron-restricted conditions. Both proteins were synthesized within 15 min of establishment of iron-restricted conditions. Proteins of equivalent molecular weights could also be induced by iron restriction in serotype 2, 3, 4, 5, and 7 isolates of A. pleuropneumoniae. Convalescent-phase sera from serotype 1-infected pigs contained antibodies which recognized both the 105K and 76K proteins from all six serotypes examined, indicating that these proteins were expressed in vivo and were immunologically conserved. Cells expressing the 105K and 76K proteins also displayed an enhanced ability to bind Congo red and hemin, suggesting that one or both of these proteins functioned to acquire complexed iron during in vivo growth.

Actinobacillus↗
PubMed1988

Purification of fimbriae from Pasteurella haemolytica A-1.

Pasteurella haemolytica A-1 produces large, rigid fimbriae with a diameter of 12 nm and a denisty of 1.32. Their subunit molecular weight was 35,000 as determined by SDS-polyacrylamide gel electrophoresis and immunoblotting with monoclonal antibodies raised against native fimbriae. The isoelectric point of the purified fimbriae was 4.8. While Pasteurella haemolytica whole cells plus a crude shear fraction were capable of agglutinating bovine erythrocytes, purified fimbriae exhibited no such activity.

Animals↗
PubMed1986

The effect of 2-micron DNA on survival and mutagenesis in Saccharomyces cerevisiae.

Strains of Saccharomyces cerevisiae, with and without endogenous 2-microns DNA, were studied in experiments designed to determine the effect of this plasmid on survival and mutagenesis in yeast. Comparison of the two strains exposed to ultraviolet light, 4-nitroquinoline oxide, or methyl methanesulfonate (MMS), revealed that the presence of 2-microns DNA slightly enhanced survival after exposure to each agent. Spontaneous frequencies of mutations (histidine reversion, canavanine resistance, and mitochondrial petites, but not adenine auxotrophy) were reduced by the presence of 2-microns DNA. MMS-induced His+ reversion was weak, and both strains responded similarly. No difference was found between the two strains when induced forward mutation to canavanine resistance was examined. The extent of induction of mitochondrial petites was about the same in both strains. Therefore, it appears that under these experimental conditions with these mutagens, 2-microns DNA has an effect on spontaneous mutation and survival after DNA damage but not on induced mutagenesis in S. cerevisiae.

4-Nitroquinoline-1-oxide↗
PubMed1985

A rapid method for the extraction and purification of DNA from human leukocytes.

A rapid method for the extraction and purification of DNA from human leukocytes was developed. Crude nucleic acids were obtained by sodium dodecylsulfate (SDS) lysis and potassium acetate precipitation of other cellular material, and the DNA was purified by ribonuclease digestion, diethylaminoethyl (DEAE) cellulose chromatography and ethanol precipitation. DNA obtained by this method is biologically active as reflected by its ability to act as substrate for various nucleases and T4 DNA ligase. The yield was sufficiently high that DNA from less than 1 ml of blood could be used for a number of reactions.

Chromatography, DEAE-Cellulose↗
PubMed1983

dnaB analog function associated with plasmid R100drd-1.

Plasmid R100 and a number of its derivatives were able to suppress the temperature sensitivity of strains carrying different alleles of the dnaB gene of Escherichia coli K-12. R100drd-l and pAR132 were able to rescue a strain carrying the dnaB266(Am) mutation in the absence of any known amber suppressors. This was taken as evidence for the existence of an R100drd-l dnaB analog function. The R100drd-l dnaB analog was different from those of bacteriophages P1 and P7 in that it was able to support the growth of bacteriophage lambda in a dnaB266(Am) background. The dnaB analog was also shown to be thermosensitive. The structural gene for this protein lies within the EcoRI fragment D of R100drd-l.

DNA, Bacterial↗
PubMed1983

Isolation of multiple mutants of Pseudomonas aeruginosa capable of efficient thymine salvage.

Mutants of Pseudomonas aeruginosa strain 1 which incorporate relatively large amounts of thymine into their DNA were isolated. Three steps were required to produce this phenotype. First, strains constitutive for thymine uptake and catabolism were isolated, followed by selection for mutants which could not catabolize thymine. The double mutant was then used to isolate a strain exhibiting greater sensitivity to the thymine analog 5-bromouracil. Efficient DNA-specific pulse labelling with isotopic thymine was successfully carried out with this strain.

Biological Transport↗
PubMed1983

FP2 plasmid curing in Pseudomonas aeruginosa.

A procedure for the elimination of the IncP-8 plasmid FP2 from Pseudomonas aeruginosa strain 1 was developed. The procedure consists of freezing cells, competent for transformation, in 15% glycerol at -70 degrees C for at least 48 h and screening survivors for loss of mercuric chloride resistance. Curing frequencies of 0.5% were achieved only in host cells carrying a dht mutation (unable to convert thymine to dihydrothymine).

Mutation↗
PubMed1982

Thymine metabolism in Pseudomonas aeruginosa strain 1: the presence of a salvage pathway.

Exogenous thymine was found to be taken up very slowly by Pseudomonas aeruginosa in comparison to other pyrimidines, and most of it was catabolized by the cell. The existence of a functional, although inefficient, thymine salvage pathway was demonstrated and this pathway operated more effectively when de novo thymidine nucleotide biosynthesis was inhibited by trimethoprim or methotrexate. The mechanism of thymine salvage by P. aeruginosa appears to be different from that of Escherichia coli and Pseudomonas acidovorans as thymidine was not incorporated into the DNA. Like P. acidovorans, P. aeruginosa lacked thymidine phosphorylase activity. Unsuccessful attempts were made to isolate thymine auxotrophs.

Culture Media↗
PubMed1982

Exonuclease activity from Pseudomonas aeruginosa which is missing in phenotypically restrictionless mutants.

A phenotypically restrictionless strain of Pseudomonas aeruginosa was found to lack a deoxyribonuclease specific for linear duplex DNA. The purified enzyme had an optimum pH of 8.5, required MgCl2 (10 mM) for maximum activity, and did not require ATP. Neither the degradation of heat-denatured DNA nor the degradation of bacteriophage F116 DNA was detected. The genome of bacteriophage F116 was shown to possess single-stranded terminal regions, which account for the resistance to degradation and for the ability of the phage to transfect restriction-proficient strains.

Bacteriophages↗
PubMedDate not supplied

Introduction of the plasmid pKM101-associated muc genes into Saccharomyces cerevisiae.

Bacteria-yeast shuttle plasmids containing the pKM101-associated muc genes were constructed by cloning an ARS TRP fragment into the plasmid pGW270 in both possible orientations. The insertion of Saccharomyces cerevisiae DNA into pGW270 had no effect on the mutator and protective phenotypes associated with the plasmid in Escherichia coli. Two such recombinant plasmids, pAA90 and pAA91 , were capable of efficient transformation of S. cerevisiae and were stably maintained in this organism. Hybridization experiments suggest that muc-specific mRNA was present in transformed yeast cells and a small amount was polyadenylated. The RNAs were not of a discrete size, all being smaller than the muc genes. The presence of the plasmid pAA91 , and to a lesser extent, pAA90 , in yeast resulted in a detectable increase in the reversion frequencies of three markers and in ultraviolet protection. These results are discussed in terms of studying the relationship of error-prone repair in bacteria and yeast and of developing improved yeast tester strains.

DNA Repair↗
PubMedDate not supplied

Attenuation of an avian pathogenic Escherichia coli strain due to a mutation in the rpsL gene.

The diseases caused by pathogenic Escherichia coli constitute a major economic loss to the poultry industry. The development of a live oral E. coli vaccine to prevent or reduce diseases in poultry had been the objective of our work. Four spontaneous streptomycin-dependent (str-dependent) mutants were generated from a virulent avian strain that contains a mutation in the fur region of the chromosome. Genetic analysis of the mutants indicated that the str-dependent phenotype was due to a base change of C --> T at base 272 in the rpsL gene. The mutants were tested for attenuation using the day-old chick model. Day-old birds, in groups of 20, were either challenged with 10(6) colony-forming units (CFU) of the str-dependent mutant, the parent strain (containing the fur mutation), or the wild-type strain without the fur mutation. The parent strain and the wild-type strain were highly virulent, and 80% or more of the birds died. None of the birds challenged with the str-dependent mutants died, indicating attenuation of the mutants. The protective effect of the mutant as a live vaccine against the challenge with 10(6) CFU of the wild-type strain EC317 was investigated. Vaccination by both aerosol (day 1) and oral (days 14 and 28) routes using 10(8) CFU of the str-dependent mutant (EC1598) had no effect on the occurrence of cellulitis in the birds. Two vaccinations given as aerosol on day 1 and given orally on day 14 also had no significant effect on the occurrence of systemic lesions. Three immunizations on days 1, 14, and 28 resulted in a significant reduction in the number of birds with systemic lesions. Antibody titers prior to challenge were not predictive of outcome of challenge.

Animals↗
PubMedDate not supplied

Effect of infection with hemorrhagic enteritis virus on susceptibility of turkeys to Escherichia coli.

The present study was designed to investigate hemorrhagic enteritis virus (HEV) as a predisposing factor influencing the susceptibility of young turkeys to Escherichia coli infections. In addition, the pathologic changes caused by administration of E. coli by various routes were compared. Following oral infection with HEV, groups of turkeys were inoculated with various doses of pathogenic E. coli by intravenous (IV), intra-air sac (IA), or intratracheal (IT) routes. A synergistic effect was observed in birds that were exposed to a combined HEV-E. coli challenge, resulting in higher mortality than that caused by either pathogen alone. This synergy was more evident when the bacteria were administered by the IT route than when it was administered by the two other routes. Turkeys infected with HEV and then inoculated IT with E. coli O78 had higher mortality (61%) and higher occurrence of gross body lesions (74%) than birds given E. coli alone (0% mortality and 16% gross lesions). After E. coli inoculation by the IA and IT routes, lesions observed were mainly pericarditis, perihepatitis, lung and air-sac lesions, splenic enlargement, and occasional arthritis. The incidence of lesions was affected by HEV exposure. In contrast, IV inoculation with E. coli O78 usually resulted in arthritis, and its incidence was independent of previous HEV exposure. The synergistic effect between HEV and E. coli administered IT can be used as a challenge model for testing E. coli vaccines.

Animals↗
PubMedDate not supplied

Experimental reproduction of Escherichia coli cellulitis and septicemia in broiler chickens.

Experimental reproduction of avian cellulitis was conducted by subcutaneous inoculation of 25-day-old broiler chickens with a field isolate of serogroup O78 Escherichia coli. Development of the cellulitis lesion occurred as early as 24 h post-infection. Reproduction of cellulitis occurred in 98% of inoculated birds, and E. coli was isolated from > 75% of cellulitis lesions. In addition to cellulitis, other lesions, including pericarditis, airsacculitis, osteomyelitis, arthritis, and perihepatitis, occurred in > 80% of birds inoculated with E. coli. Bacteremia occurred as early as 6 h post-infection and dramatically declined by 5 days post-infection. Seventeen of 59 (29%) birds inoculated with E. coli developed a fatal infection between 1 and 6 days post-infection, and bacteria were isolated from lesions in 98% birds. In contrast, E. coli was not isolated from lesions in birds that survived until days 7-14 post-infection. Birds that survived with cellulitis and other lesions until day 14 post-infection had a significantly lower body weight compared with the control group. This avian model of cellulitis and other lesions will be useful for studying the development of vaccination strategies for E. coli in broilers.

Animals↗