Investigation of the effects of antifolates on Escherichia coli 1810.

The Gel electrophoresis procedure was employed in the investigation of the effect of Trimethoprim (TMP), Sulphadiazine (SD) and p-Aminobenzoic acid (pABA) on the outer membrane and peptidoglycan (OM-PG) associated proteins in the test bacteria. The test organisms were the sensitive Escherichia coli NCTC K-12 (E.coli K-12) and Escherichia coli 1810 (E.coli 1810) which was highly resistant to TMP. The checkerboard determinations indicated synergism with SD:TMP combinations against both E.coli strains. Viable counts also indicated that SD exhibited synergism with TMP after 2 h and throughout the next 5 h in both E.coli strains. The results on outer membrane (OM) permeability showed that 1/4 and 3/4 MICs of either SD or TMP alone or combined against both E.coli strains altered the OM permeability. It was unlikely that 3/4 MICs of either SD or TMP alone and combined altered the cytoplasmic membrane (CM) permeability of either E.coli strain. Light microscopy (LM) showed that cells of both E.coli strains were both longer and wider after exposure to their respective near MICs of either SD or TMP alone and combined. At 3/400 of their MICs both strains appeared similar to the control cells. Electron Microscopy (EM) of E.coli 1810 grown in the presence of either 300 upsilon g/ml of SD or TMP alone and combined indicated marked structural damage. No effect on the OM or ultrastructure of E.coli 1810 was observed with 3/400 MICs of either SD or TMP alone and combined. It should be noted that this concentration affected the permeability of sensitive cells. Antibacterial concentrations which affected the ultrastructure of cells of both E.coli strains as determined by EM, were also shown by the permeability probe to alter the OM and also partially inhibited the growth of E.coli 1810 cultures. Since the action of SD and TMP, singly and in combination, on the cell structure of E.coli 1810 took place only at concentrations approaching the respective MICs of the antibacterials, then this effect on cell structure was concluded to be a normal component of their antibacterial action. OM-PG associated protein bands separated from both E.coli strains by SDS-PAGE after exposure to either 1/2 MICs of SD or TMP alone and combined showed an absence of higher and medium MW protein bands with E.coli K-12 and medium MW bands with E.coli 1810. Accumulation of low MW protein bands occurred with both E.coli strains when compared with the control cells. The combination of SD plus TMP produced a greater effect on the pattern of protein bands with the E.coli K-12 than with the E.coli 1810. Several protein bands were absent from the SDS-PAGE results obtained with E.coli K-12 but only 2 protein bands were absent from the resistant E.coli 1810 after exposure to either SD or TMP alone and in combination. LM results showed that 1/2 MIC of pABA produced longer and wider cells when compared with the control cells of both E.coli strains. OM permeability provided evidence that 1/2 MIC of pABA altered the OM permeability of E.coli K-12 but not E.coli 1810. It was unlikely that 1/2 MIC of pABA altered the CM permeability of either E.coli strain. Gel electrophoresis showed that 1/2 MIC pABA affected the OM-PG associated proteins of both E.coli strains. The number of absent protein bands from the sensitive E.coli K-12 was greater than for the resistant E.coli 1810. In addition, the number of higher MW protein bands was greater and the number of lower MW protein bands was less for the resistant E.coli 1810 than the sensitive E.coli K-12.

Grantor dc:publisher.institution

Robert Gordon University

Year dc:date.issued

1999

Author dc:creator

• Eumkeb, Griangsak

Advisor dc:contributor.advisor

• R.M.E. Richards, J.Z. Xing, D.G. Chapman and I.X. Liu

• Antifolates
• Escherichia coli 1810
• Trimethoprim
• Sulphadiazine
• P-aminobenzoic acid
• Outer membrane
• Peptidoglycan
• Protein bands

Language dc:language

en

Identifier

oai:rgu-repository.worktribe.com:2807375
https://doi.org/10.48526/rgu-wt-2807375

OAI identifier oai:identifier

oai:rgu-repository.worktribe.com:2807375