Resistance to antibiotics is an increasing problem in the developing world.
Resistance genes in the normal gut flora may act as a reservoir from which
pathogens may acquire resistance. Trimethoprim is an important drug in the
treatment of a wide range of infections but resistance has increased in recent years
particularly in the developing world.
A survey was conducted in Vellore, south India, to determine the rate of carriage
of antibiotic resistant enterobacteria in the normal gut flora of a sample of the
population. Very high rates of resistance were encountered to the widely used
antimicrobials, ampicillin, chloramphenicol and trimethoprim. The trimethoprim
resistant strains were analyzed further, 35% were capable of plasmid-mediated
transfer of trimethoprim resistance by different plasmid types, as identified by
restriction endonuclease digestion. Resistance to a variety of other agents was cotransferred. Transferable trimethoprim resistance was shown, by DNA
hybridization, to result from the presence of three different drug resistant
dihydrofolate reductase (dhfr) genes. The dhfrV was present in 50% of
transconjugants, associated with either the transposon Tn21 or only with the
integrase-like open reading frame (orf) of this transposon; 31% possessed the dhfrl,
associated with the integrase orf of Tn7 and 19% had dhfrlV
The dihydrofolate reductase (DHFR) enzyme type IV has only ever been identified
in this area of India, in 1984. The enzyme mediates only low level resistance, as
measured by conventional tests, but it is inducible, a unique property in the
DHFRs. It has persisted in this area despite a seemingly poor resistance
mechanism. The DHFR typelV enzymes characterized during this survey were also
inducible. It was demonstrated that the induction mechanism will produce over
100-fold greater resistance if the cells are challenged prior to determination of
resistance level. This mechanism was shown to be mediated by a lack of thymine
and to be dependant on cell phase and density. A plasmid characterized in this
study also mediated resistance to ampicillin as a result of the presence of the
TEM-1 ß-lactamase. This enzyme was also inducible, a property not observed
before for this enzyme or for any other plasmid mediated ß-lactamase in Gram-negative bacteria, producing increased resistance to the widely used amoxycillinclavulanic acid combination. The induction of both enzymes was cross-reactive,
both trimethoprim and the ß-lactam drugs induced both mechanisms. The
metabolic impact of the ß-lactams must be triggering the induction mechanism.
This cannot, however, be by the same pathway as trimethoprim.
The high rates of carriage of normal gut flora resistant to antimicrobials are
probably the result of selection by antimicrobials which are freely available without
prescription combined with poor hygiene and sanitation. The evolution of such
unusual induction mechanisms may have resulted from the prevailing low level of
antimicrobials or frequent exposure to low levels of drug as a consequence of self - dosing. High rates of resistance in normal flora and the evolution of such resistance
mechanisms is serious because of their impact on the development of resistance and
in the testing for resistance, when such induction mechanisms may disguise true
The DNA sequence of the dhfrIV was also determined and was shown to be only
distantly related to the chromosomal dhfr and other plasmid mediated dhfrs.