We collected serotyped isolates of salmonella from reference laboratories in the United States, tested their susceptibility to antibiotics, and extracted plasmids from isolates that were resistant to a different combination of antibiotics from each of three serotypes. Restriction-endonuclease digestion showed that within each of the three groups, plasmid molecules from animal and human isolates were often identical or nearly identical. One serotype-plasmid combination appeared to be endemic in cattle in 20 states and infected 26 persons in two states. The human cases, which were not recognizably related except for their common plasmids, appeared to be clustered in time but geographically dispersed, like cases in previous outbreaks spread by food products. These findings suggest that resistance plasmids may be extensively shared between animal and human bacteria, and that spread of multiresistant strains of salmonella among animals and human beings, as observed in Britain, may have been undetected in the United States for lack of comparable surveillance.

Bacteria of different genera isolated at nine medical centers in different parts of the United States and at one center in Venezuela during the first decade of gentamicin usage carried the gentamicin resistance gene 2"-aminoglycoside nucleotidyltransferase on the same transferable plasmid. Such widespread dissemination of a newly observed resistance gene on one plasmid suggests that a new resistance gene may emerge once on a single plasmid, which then carries it to other centers and other plasmids. The resistance gene might, therefore, be contained if detected early.

During the first 6 years after appearing in one hospital, a 92-kilobase conjugative plasmid, pBWHl, which encoded resistance to chloramphenicol and sulfonamides and determined TEM-1 beta-lactamase and 2"-aminoglycoside nucleotidyltransferase, underwent a variety of molecular changes. It was most prevalent initially in isolates of Klebsiella pneumoniae, then in isolates of Serratia marcescens, and finally, after nearly disappearing, in isolates of Enterobacter cloacae. Evolutionary changes in the plasmid did not account for its shifts in species distribution, since the original molecule was found in isolates of each species. The late resurgence of pBWHl occurred after a copy of its original molecule entered a distinctive ornithine decarboxylase-negative strain of E. cloacae, new to the hospital. The resulting transconjugant strain, chromosomally resistant to topical silver salts and to cephalosporins, and with the addition of pBWH1-encoded aminoglycoside resistance, spread in the hospital by causing an outbreak of sepsis in the burn unit, where these were commonly used antibacterial agents. Thus, an endemic plasmid became prevalent in a new host species because one of its genes supplemented the fitness of an uncommon strain of the species for a particular clinical niche.The percentages of clinical isolates of different bacterial species resistant to specific antibiotics are frequently calculated to delineate the usefulness of these drugs. Such tabulations will reflect in large part the distribution of resistance plasmids among strains of the species being surveyed, since a majority of resistance genes are carried by plasmids (4). The distribution of the plasmids may be determined by properties of the plasmids themselves such as their transferability, host range, or stability or the selective advantages conferred by their resistance genes or other cryptic genes (4,5). Also, properties of the host strains such as their fitness to occupy niches in which they come in contact with antibiotics might contribute to the observed distribution of plasmids.The surveillance of antibiotic resistance plasmids has been aided by plasmid fingerprinting, using restriction endonucleases (6,9,10,15,24,27)

We present a genetic and physical characterization of the IncM plasmid pBWH1. A physical map was constructed for the enzymes EcoRI, BamHI, Sall, BglII, HindIII, MstII, and XhoI. A series of deletions and a series of subclones of pBWH1 were constructed and used to determine the locations on this map of the transfer region; the replication region; and the genes determining resistance to beta-lactamg, chlotaamphenicol, the sulfonamides, and gentamicin. We compared 51 different isolates, including isolates which had lost individual antibiotic resistances or the transfer phenotype, and showed that variations occureed in all regions of the plasmid genome. Frequently, correlations could be made between phenotypic variation and variation of the EcoRI fragments which contained the gene determining that phenotype.The plasmid pBWH1 is a 92-kilobase IncM plasmid that has a broad host range for gram-negative members of the family Enterobacteriacae and carries four antibiotic resistance genes. These genes encode resistance to the aminoglycosides gentamicin, tobramicin, and kanamycin; to ampicillin and carbenicillin; to chloramphenicol; and to the sulfonamides (16). We have been monitoring the evolution of this plasmid and its derivatives at one location. Over a 10-year period, we obtained over 2,000 different isolates of strains carrying this plasmid as we followed the epidemic spread of pBWH1 at the Brigham and Women's Hospital in Boston.We examined phenotypic changes and restriction fragment variations. Analysis of these variations in pBWH1 may extend an important technique in the field of molecular evolution. Restriction fragment variation of, for example, metazoan mitochondria is currently being used to ascertain phylogenetic relationships (7, 13). The results of these experiments can be compared with the relationships inferred from comparative morphology and the fossil record; but rholecular data are available only for the present. The molecular evolution of bacterial plasmids is much faster, with significant changes appearing over relatively brief periods. Thus it is possible to monitor changing populations of plasmids through real time, as well as to extensively sample a population at any given time.pBWH1 has been found in strains of six different genera of the family Enterobacteriacae. Screening of many of these isolates has shown that the full phenotype of pBWH1, as well as a standard pattern of EcoRI restriction enzyme fragments, has persisted from 1976 to the present. A number of isolates, however, have shoWn significant phenotypic variations, losing resistance to an antibiotic or the ability to transfer the plasmid. There have also been many variations in the pattern of EcoRI fragments. We refer to these isolates as the pBWH1 series of plasmids. In this report we present a genetic and restriction enzyme map of pBWH1 and discuss variations in natural isolates in relationship to this map. We * Corresponding author.show that the variations of EcoRI restriction enzyme sites indicate that variations occur throughout th...