R factors were first demonstrated in Japan in 1959 when it
was shown that resistance to several antibiotics could be transferred as a unit
between strains of Shigella and Escherichia coli by conjugation. Many surveys
since then in all parts of the world have shown that R factors are now common
and widespread. Resistance to sulphonamides, tetracycline, chloramphenicol,
ampicillin, streptomycin, kanamycin, neomycin and gentamicin have all been
found to be commonly extra-chromosomal in enterobacteria and transferable to
drug-sensitive organisms in vitro or in vivo.
The actual incidence of drug resistance in particular species varies when different populations are studied, but is found to be increasing in most pathogenic species throughout the world; the incidence of strains with multiple resistances is similarly increasing. It is not known how or where extrachromosomal (plasmid-borne) resistance genes originate, but whether they originate in chromosomal or in plasmid DNA, they are certainly liable to appear in R factors soon after a new antibiotic comes into general use. The exact way in which R factors are built up is the subject of some dispute, but it is clear that simple transfer factors that can pick up resistance genes are relatively common in enterobacteria and that such transfer factors can combine with non-transmissible resistance plasmids to produce transmissible R factors. R factors can recombine with non-transmissible resistance plasmids or with other R factors to produce complex R factors that contain genes for resistance to several antibiotics, so that under suitable conditions transferable multiple-resistance R factors can be built up.
Resistance to as many as seven or eight different antimicrobial drugs has been found to be carried in a single R factor. This process need not occur in a single bacterial species. R factors and transfer factors can transfer themselves into a wide range of commensal and pathogenic bacteria in vitro, e.g. Escherichia, Salmonella, Shigella, Klebsiella, Proteus, Serratia, Pasteurella, Yersinia, Vibrio and Pseudomonas. Once drug resistance appears in any of these species it may be picked up and built into multiple-resistance R factors which may then be distributed to cells of other Gram-negative species.
It is relatively easy to study the build-up and spread of R factors in vitro under the selective pressure of antibiotics, but it is harder to analyze in vivo. Conditions in the normal gut are pot very suitable for conjugation: alkaline pH, bile salts, fatty acids and anaerobic conditions my all inhibit conjugation; pili are better formed by actively growing cells than by stationary phase cells; and since the number of suitable recipient cells in the gut is considerably lower than that of other organisms such as Bacteroides and Gram-positive species, opportunities for a donor cell to come into contact with a suitable recipient are not frequent.
It is often dace to show spread of R factors to other bacterial species in the gut of a normal animal; the R factors are perpetuated mainly by linear paw mission to the progeny at cell division. When an antibiotic is given, however, the numbers of drug-sensitive organisms are rapidly reduced drug-resistant organisms are selected and multiply to take their place and opportunities for the transfer of R factors are improved.
Any originally drug-sensitive organisms that receive R factors are also selected while those that do not are killed. It is much easier to show R factor spread in vivo when antibiotic selection is used; the result of giving an antibiotic may be a dramatic increase in the incidence of R factors in the gut flora. Although R factors may have been present originally in only a small pro-portion of the cells of a single species, anti-biotic treatment may select for them so that they become present in a high proportion of cells in several species.
Since R factors may contain multiple resistance genes the effect of giving a single antibiotic may be to ensure that the majority of the enterobacteria in the gut, commensal or pathogenic, come to contain an R factor that confers resistance, not just to the anti-biotic given, but to several other drugs as well. R factors were not demonstrated until 1959 in Japan, 1962 in Britain and 1966 in the United States, although many of the basic antibiotics had already been in use for many years before these dates. Whatever the reasons for this initial delay in appearance, R factors are now common in commensal and pathogenic drug- resistant enterobacteria from the gut, and from infections caused by organisms derived from the gut, e.g. urinary tract infection and Gram-negative septicaemia.
It seems that this is an inevitable consequence of the widespread use of antibiotics. The frequent exposure of a human or animal population to a variety of antibiotics leads to the appearance of extrachromosomal resistance genes in the gut enterobacteria. Transfer factorsthat can pick up these genes acquire a new survival value both to themselves and their host bacteria, and a continuing exposure to antibiotics leads to an increasing frequency of R factors in a variety of bacterial species.
Infections caused by pathogens containing R factors may be very difficult to treat and as workers in human and animal medicine are faced with increasing drug resistance in pathogenic species, there is a tendency towards the administration of a greater range of drugs. In turn, this results in the build-up of more complex R factors that become more widely distributed to commensal and pathogenic organisms. The dangers of transferable (infectious) drug resistance are obvious.
If the incidence of multiple-resistance R factors continues
to increase, infections by coliform organisms will become more difficult to
treat. Many of these infections are usually mild and do not require antibiotic
therapy, e.g. most cases of salmonella
food poisoning and Shigella sonnet dysentery, but antibiotic therapy is of
great importance in others. In typhoid fever, for example, treatment with
chloramphenicol is often life-saving and we have few effective substitutes. R
factors with chloramphenicol resistance had been common in food-poisoning
salmonellae for many years but did not appear in Salmonella typhi until 1972.
Since then, epidemics of chloramphenicol-resistant typhoid
fever in Mexico and Asia have had higher mortality rates and have proved more
difficult to control. Cholera and bacillary dysentery would become much harder
to control if anti-microbial drugs were to lose their efficacy. The antibiotic
treatment of Escherichia coli enteritis in infants is becoming more difficult
because of the increase in R factors in human enteropathogenic strains and drug
resistance is a growing problem in the therapy of urinary tract infection.
There is an increasing frequency of 'opportunist'
infections caused by drug-resistant organisms that are normally of low
virulence. Some opportunist pathogens belong to species that are naturally
resistant to many antibiotics. e.g. Candida, but others are strains of
Gram-negative species such as Pseudomonas, Serratia or Klebsiella that have
acquired R factors.
A further danger that
has attracted a great deal of attention is the transfer to man of R factors
evolved in farm animals. E. S. Anderson and others in the Enteric Reference
Laboratory, London, studied strains of salmonellae typhimurium isolated from
calves during a succession of outbreaks of enteritis among calves in 1963-66.
Infection spread rapidly because of overcrowding and poor hygiene as intensive
rearing was introduced, and large amounts of antibiotics were used in
unsuccessful attempts to control the outbreak. An increasing proportion of all
salmonellae isolated from cattle were found to be salmonellae typhimurium type
29 and these organisms acquired transferable resistance to increasing numbers
of antibiotics as the outbreak progressed.
The Multiple-resistance R factors became common not only in
the infecting salmonellae typhimurium type 29 but also in commensal Escherichia
coli from the same batches of calves. The outbreak was eventually brought under
control only by alterations in the arrangements for breeding and transporting
calves to reduce the amount of cross-infection between the animals. Human
infection with salmonellae typhimurium is usually a result of food-poisoning;
the main reservoir of infection is in farm animals and the disease is spread to
man by contaminated meat or other food. The outbreak of infection in calves was
paralleled by a rise in the incidence of human food-poisoning due to salmonellae
typhimurium type 29 containing the same R factors.
There is no doubt that these strains were derived from the
calves as many were resistant to furazolidone, a drug used only in veterinary
practice, as well as to other agents used in both man and animals. There is a
risk that such R factors, evolved in farm animals, will spread to human
commensal Esch. coli and then be transferred to human pathogens that cause more
serious infections than food-poisoning. R. factors may be transmitted to man by
commensal bacteria from animals as well as by pathogens. The rate of transfer
of food-poisoning salmonellae to man is considerable despite careful
precautions in food preparation; the rate of transfer of non-pathogenic
organisms must be greater. Although animal commensals may be less likely to
establish themselves in the human gut.
R factors are common in Escherichia coli and Salmonella
strains isolated from pigs and chickens as well as from calves. Penicillin or
tetracycline may be given to young animals as a feed supplement in order to
produce a faster growth rate. The practice leads to the selection of R factors
that contain genes for tetracycline resistance and for penicillinase
production. Gram-negative bacilli are naturally resistant to penicillin but are
susceptible to ampicillin since ampicillin is also inactivated by penicillinase,
the use of penicillin as a feed supplement leads to production of ampicillin
resistance in the Gram-negative flora of the gut.
The use of antibiotics as feed supplements is restricted by law in Britain and only certain antimicrobial drugs not used in human medicine may now be used. Antibiotics are also widely prescribed for the treatment of infections in farm animals and the drugs used in veterinary practice are largely the same as those used in man. In 1967 the total amount of antibiotic used in human medicine was estimated at 240 tons and the total amount in agriculture, both for therapy and feed supplements, was 170 tons. One of the main differences between veterinary and medical practice is a tendency for treatment of a whole animal population rather than an individual human patient. In certain cases the whole population is treated in the hope that the infected animals will be cured and infection will be prevented in the others. In other circumstances, farmers are accustomed to administer low doses of antibiotics to a whole population of animals that are judged to be under 'stress' and particularly liable to develop infection.
When R factor resistance is present, the mass use of antibiotics fails to prevent the spread of infection or to cure infected animals and results in a very high incidence of the R factor in the gut flora of the whole batch of animals. It is probable that sensible limitation of antibiotic usage in man and animals could prevent. Further increase in R factors and perhaps reduce their incidence. Many R factors are unstable and tend to lose resistance genes when the selective pressure is removed. R factors are lost spontaneously from a small proportion of cells in a culture since plasmid replication and segregation are not always precisely synchronous with chromosome replication and segregation.
Cells that lose an R factor may have a slight metabolic advantage because they no longer have the bur-den of producing plasmid nucleic acids and proteins. In the absence of antibiotics, drug-resistant organisms have no selective advantage and may be slowly outgrown by drug-sensitive organisms. Moreover, R factors that evolve in one species may not thrive in another.
An R factor that is well adjusted to existence in Escherichia may be unstable in Proteus strains and be lost at a high rate, or it may transfer itself to other organisms much less efficiently. Similarly, commensal or pathogenic organisms that are adapted to the gut of a calf, pig or chicken may not be established readily in man; they may be present only transiently and conjugate seldom with the human commensal flora. Such factors as these may be helping to contain the spread of R factors. The results of some experiments with populations of animals encourage the hope that R factors slowly disappear when no antibiotics are given.