Showing papers by "Marilyn C. Roberts published in 1997"

Distribution and mobility of the tetracycline resistance determinant tetQ.

Abstract: We tested 34 American Type Culture Collection (ATCC) and 168 clinical bacterial isolates, from the human urogenital and oral tracts and streptococci isolated from cows with mastitis, for the presence of the tetQ gene using a polymerase chain reaction (PCR) assay and DNA-DNA hybridization. The identities of PCR products were confirmed by Southern blot hybridization of whole-cell DNA. Eleven of the ATCC strains were positive for tetQ, including five Bacteroides spp., five Prevotella spp. and a single isolate of Mitsuokella multiacidus. Twenty-eight (29%) of the 95 clinical Gram-negative isolates carried the tetQ gene, while eight (11%) of the 73 clinical Gram-positive isolates carried the tetQ gene. This is the first description of tetQ in Gram-positive species. All isolates except one Peptostreptococcus sp. carried tetQ integrated into the chromosome. The tetQ gene could be transferred from Prevotella bivia, Bacteroides ovatus, Bacteroides fragilis, Bacteroides vulgatus and Bacteroides distasonis into an Enterococcus faecalis recipient at frequencies of 10(-7)-10(-9) per recipient. In contrast, tetQ failed to transfer from two isolates of Prevotella intermedia, two isolates of Porphyromonas gingivalis, one isolate of Mobiluncus curtisii and one isolate of Peptostreptococcus sp. The latter two are Gram-positive species. The PCR assay was used to screen 198 proteinase K-treated biopsies of prostate, periprostate and bladder from 84 men with prostatitis. Thirty-four (40%) of the patients had one or more positive samples, suggesting that the PCR assay could be of value in screening patient material directly for the presence of bacteria.

Mycobacterium tuberculosis efpA encodes an efflux protein of the QacA transporter family.

Abstract: The Mycobacterium tuberculosis H37Rv efpA gene encodes a putative efflux protein, EfpA, of 55,670 Da. The deduced EfpA protein was similar in secondary structure to Pur8, MmrA, TcmA, LfrA, EmrB, and other members of the QacA transporter family (QacA TF) which mediate antibiotic and chemical resistance in bacteria and yeast. The predicted EfpA sequence possessed all transporter motifs characteristic of the QacA TF, including those associated with proton-antiport function and the motif considered to be specific to exporters. The 1,590-bp efpA open reading frame was G+C rich (65%), whereas the 40-bp region immediately upstream had an A+T bias (35% G+C). Reverse transcriptase-PCR assays indicated that efpA was expressed in vitro and in situ. Putative promoter sequences were partially overlapped by the A+T-rich region and by a region capable of forming alternative secondary structures indicative of transcriptional regulation in analogous systems. PCR single-stranded conformational polymorphism analysis demonstrated that these upstream flanking sequences and the 231-bp, 59 coding region are highly conserved among both drug-sensitive and multiply-drug-resistant isolates of M. tuberculosis. The efpA gene was present in the slow-growing human pathogens M. tuberculosis, Mycobacterium leprae, and Mycobacterium bovis and in the opportunistic human pathogens Mycobacterium avium and Mycobacterium intracellular. However, efpA was not present in 17 other opportunistically pathogenic or nonpathogenic mycobacterial species.

Genetic mobility and distribution of tetracycline resistance determinants.

Abstract: Since 1953, tetracycline-resistant bacteria have been found increasingly in humans, animals, food and the environment. Tetracycline resistance is normally due to the acquisition of new genes and is primarily due to either energy-dependent efflux of tetracycline or protection of the ribosomes from its action. Gram-negative efflux genes are frequently associated with conjugative plasmids, whereas Gram-positive efflux genes are often found on small mobilizable plasmids or in the chromosome. The ribosomal protection genes are generally associated with conjugative transposons which have a preference for the chromosome. Recently, tetracycline resistance genes have been found in the genera Mycobacterium, Nocardia, Streptomyces and Treponema. The Tet M determinant codes for a ribosomal protection protein which can be found in Gram-positive, Gram-negative, cell-wall-free, aerobic, anaerobic, pathogenic, opportunistic and normal flora species. This promiscuous nature may be correlated with its location on a conjugative transposon and its ability to cross most biochemical and physical barriers found in bacteria. The Tet B efflux determinant is unlike other efflux gene products because it confers resistance to tetracycline, doxycycline and minocycline and has the widest host range of all Gram-negative efflux determinants. We have hypothesized that mobility and the environment of the bacteria may help influence the ultimate host range of specific tet genes. If we are to reverse the trend towards increasingly antibiotic-resistant pathogenic bacteria, we will need to change how antibiotics are used in both human and animal health as well as food production.

Genomic homogeneity of the AHU/IA-1,2 phenotype of Neisseria gonorrhoeae during its disappearance from an urban population.

Abstract: BACKGROUND Isolates of Neisseria gonorrhoeae requiring arginine, hypoxanthine, and uracil (AHU) appeared in Denmark in 1946, were preponderant in Seattle during the 1970s, were associated with disseminated gonococcal infections (DGI), and were primarily of the IA-1,2 serovar. GOALS To investigate the disappearance of the AHU/IA-1,2 phenotype and to examine by pulsed-field gel electrophoresis (PFGE) the genomic homogeneity of this unique phenotype isolated from Seattle-King County during the past decade. STUDY DESIGN This retrospective study used data extracted from previous publications for the period 1971 through 1984, and from existing records at the Neisseria Reference Laboratory, University of Washington for the period 1985 through 1996. Samples (n = 68) of AHU/IA-1,2 isolates from 1984 to 1985 and 1988 to 1993 were analyzed using endonucleases NheI and XbaI. For comparison, 10 AHU/IB isolates were included in the study. RESULTS AHU isolates, predominantly IA-1,2 strains accounted for 52% of the gonococcal isolates for the period 1971 through 1974, 40% for 1974 through 1976, 16% for 1984, and then declined from 7% in 1986 to 0% for each of the last 3 years, 1994 through 1996. Isolates with < or = 1 band difference after digestion with either NheI or XbaI were considered to belong to a single closely related pattern. Pulsed-field gel electrophoresis (PFGE) type designation was made from the combination of NheI and XbaI patterns. These criteria yielded 5 NheI and 8 XbaI PFGE patterns, and 11 PFGE types based on the combination of NheI and XbaI pattern. The most frequently occurring NheI and XbaI patterns accounted for 74% and 57% of isolates, respectively. One type persisted throughout the decade and accounted for 54% of the 68 isolates. Analysis of the 10 AHU/IB isolates yielded 7 NheI and 8 XbaI patterns that gave 9 types that were distinct from the types found in the AHU/IA-1,2 strains. CONCLUSION The AHU/IA-1,2 phenotype first documented 50 years ago in Denmark still shows a high degree of genomic homogeneity during the past decade in Seattle. The implementation of screening, decreased rates of sexual exposure in the acquired immune deficiency syndrome era, or other factors may explain its apparent elimination in Seattle-King County.