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Genetic characterization of lactose fermentation and antibiotic resistance in plasmid-harboring enteric bacteria isolated from amazon amphibians

 

 

Luzia Mitie IoshimotoI; Lucile Maria Floeter-WinterII; Carlos Eduardo WinterII; Willy BeçakI; Maria Heloiza T. AffonsoII

IServiço de Genética, Instituto Butantan, Av. Vital Brasil, 1500, 05504 São Paulo, SP, Brasil. Send correspondence to L.M.I.
IIDepartamento de Parasitologia, Instituto de Ciências Biomédicas, USP, 05508 São Paulo, SP, Brasil

 

 

ABSTRACT

Aerobic gram-negative bacteria were isolated from the feces of 20 wild amphibians. The animals were collected from regions flooded by the waters of the Tocantins river during the building of the Tucuruí-PA hydroelectric plant. Of the 420 isolates tested for the presence of plasmids, 123 (29%) showed at least one plasmid DNA band.
Among the 123 plasmid containing isolates, 89% were lactose fermenting, 41% showed resistance to ampicillin, 6.5% to streptomycin and 6.5% to tetracycline, at levels ≥ 10 mg/ml.
The most prevalent genera identified among the 123 bacterial isolates were the Citrobacter (67%), followed by Enterobacter (16%), Klebsiella (6%), Providencia (3%), Escherichia (3%), Proteus (2%), Aeromonas (2%) and Salmonella (1%).
As most of the detected plasmids were large enough to carry genes for conjugal transfer, the relationship between the presence of plasmids and characteristics such as lactose fermentation and drug resistance was analyzed by mating to E. coli. The presence of these genes was also examined by DNA-DNA hybridization, using specific probes.

Keywords: Lactose; Antibiotic resistance; Enteric bacteria; Amazon amphibians.

 

 

REFERENCES

Affonso, M.H.T., Toledo, M.R.F. and Trabulsi, L.R. (1983). Natureza genética da fermentação de lactose em amostras de Salmonella typhimurium. Rev. Microbiol. 8: 110-116.

Ambler, R.P. and Scott, G.K. (1978). Partial amino acid sequence of penicillinase coded by Escherichia coli plasmid R6K. Proc. Natl. Acad. Sci. (USA) 75: 3732-3736.

Anderson, E.S. (1968). The ecology of transferable drug resistance in the enterobacteria. Ann. Rev. Microbiol: 22: 131-180.

Boissinot, M., Mercier, J. and Levesque, R.G. (1987). Development of natural and synthetic DNA probes for OXA-2 and TEM-1 ß-lactamases. Antimicrob. Agents Chemother. 31: 728-734.

Burton, N.F., Day, M.J. and Bull, A.T. (1982). Distribution of bacterial plasmids in clean and polluted sites in South Wales River. AppL Environ. Microbiol. 44: 1026-1029.

Can, A., Amborski, R.L., Culley Jr., D.D. and Amborski, G.F. (1976). Aerobic bacteria in the intestinal tracts of Bullfrogs (Rana catesbiana) maintained at low temperature. Herpetologica 32: 239-244.

Chang, B.J. and Bolton, S.M. (1987). Plasmids and resistance to antimicrobial agents in Aeromonas sobria and Aeromonas hydrophila clinical isolates. Antimicrob. Agents Chemother. 31: 1281-1282.

Christiansen, C., Christiansen, G., Bak, A.L. and Stendemp, A. (1973). Extrachromosomal deoxyribonucleic acid in different entero-bacteria. J. Bacteriol. 144: 367-477.

Davis, C.E. and Nandan, J. (1970). The evolution of R factor. A study of preantibiotic community in Borneo. New England J. Med. 283: 117-122.

Eckhardt, T. (1978). A rapid method for the identification of plasmid desoxyribonucleic acid in bacteria. Plasmid 1: 584-588.

Edwards, P.R. and Ewing, W.H. (1986). Identification of Enterobacteriaceae. Elsevier Science Publishing Co. Inc. (4.ed.). New York, USA.

Foster, T.J. (1983). Plasmid determined resistance to antimicrobial drugs and toxic metal ions in bacteria. Microbiol. Rev. 47: 361-409.

Gardner, P., Smith, D.H. and Beer, H. (1969). Recovery of resistance (R) factors from a drug-free community. Lancet 2: 774-776.

Glassman, D.L. and MacNicol, L.A. (1984). Plasmid frequency in natural populations of estuarine microorganism. Appt Environ. Microbiol 48: 518.

Gossling, J., Loesche, W.L. and Nace, G.W. (1982). Large intestine bacterial flora of nonhibemating and hibernating Leopard frogs (Rana pipiens). Appl. Environ. Microbiol. 44: 59-66.

Graves, S.R., Rawlinson, P.A., Kennelly-Merrit, S.A., MacLaren, D.A., Harvey, K.J. and Thorton, W.B. (1988). Enteric bacteria of reptiles on Java and the Krakatau Islands. Phil. Trans. R. Soc. Lond. B 322: 355-361.

Gross, D.C., Vidaver, A.K. and Keralis, M.B. (1979). Indigenous plasmids from phytopathogenic Corynebacterium species. J. Gen. MicrobioL 115: 479-489.

Hada, H.S. and Sizemore, R.K. (1981). Incidence of plasmids in marine Vibrio spp. isolated from an oil field in the northwestern gulf of Mexico. Appl. Environ. Microbiol. 41: 199-202.

Hughes, V. and Datta, N. (1983). Conjugative plasmids in bacteria of the pre-antibiotic era. Nature 302:725-727.

Iveson, J.B., Mackay-Scollay, E.M. and Bamford, V.W. (1969). Salmonella and Arizona in reptiles and in man in Western Australia. J. Hyg. Camb. 67: 135-145.

Jamieson, A.F. and Bremner, D.A. (1979). Characterization of plasmids from antibiotic resistant Shigella isolates by agarose gel electrophoresis. J. Gen. Microbiol. 117: 73-81.

Kado, C.I. and Liu, S.T. (1981). Rapid procedure for detection and isolation of large and small plasmids. J. BacterioL 145: 1365-1373.

Kobori, H., Sullivan, C.W. and Shizuya, H. (1984). Bacterial plasmids in Antartic natural microbial assemblages. AppL Environ. Microbiol. 48: 515-518.

Kron, M.A., Currie-McCumber, D.M.C. and Medeiros, A.A. (1987). Molecular epidemiology of OHIO-1 b-lactamase. Antimicrob. Agents Chemother 31: 2007-2009.

Levy, S.B. (1984). Resistance to the tetracyclines. In: Antimicrobial drug resistance (Bryan, L., ed.). Academic Press, Orlando, Fl.

Lins, Z.C. (1971). Studies on enteric bacterias in the lower Amazon region: II Salmonella types isolated from wild reptiles in Pará state-Brazil. Rev. Microbiol. 2: 165-169.

Maniatis, T., Fritsch, E.F. and Sambrook, J. (1982). Molecular cloning. A laboratory manual. Cold Springer Harbor Laboratory, Cold Spring Harbor, New York - USA.

Marshall, B., Tachibana, C. and Levy, S.B. (1983). Frequency of tetracycline resistance determinant classes among lactose-fermenting conforms. Antimicrob. Agents Chemoter 24: 835-840.

Marshall, B., Morrissey, S., Tlynn, P. and Levy, S.B. (1986). A new tetracycline-resistance determinant, class E, isolated from Enterobacteriaceae. Gene 50: 111-117.

Mattew, M., Hedges, R.W. and Smith, J.T. (1979). Types of b-Lactamase determined by plasmids in gram negative bacteria. J. Bacteriol. 138: 657-662.

Mattew, M. (1979). Plasmid-mediated b-lactamase of gram negative bacteria: properties and distribution. J. Antimicrob. Chemother. 5: 349-358.

Mekalanos, J.J. (1983). Duplication and amplification oftoxin genes in Vi brio cholerae. Cell 35: 253-263.

Mendez, B., Tachibana, C. and Levy, S.B. (1980). Heterogeneity of tetracycline resistance determinants. Plasmid 3: 99-108.

Milch, H. and Nguyen, T.K. (1981). Klebsiella and Enterobacter strains derived from hospital infections. Acta Microbiol. Acad. Sci. Hung. 28: 171-196.

Miller, J.H. (1972). Curing of episomes from Escherichia coli strains with acridine orange. p. 104-106. In: Experiments in molecular genetics (Cold Spring Harbor Lab., ed). Cold Spring Harbor, New York.

Moller, R.K., Jorgensen, N.H.F., Christiansen, C., Christiansen, G., Bak, A.L. and Stenderup. A. (1978). Characterization of plasmids from wild type Enterobacteriaceae, p. 257-261. In: Microbiology (D.Schiessinger, ed.). American Society for Microbiology, Washington, DC.

Moreno, G., Lopez, C.A.M., Belluomini, H.E., Pessoa, G.V.A., Biasi, P. and Andrade, J.C.R. (1973). Enterobactérias isoladas de anfíbios e repteis. Rev. Inst. Med. Trop. 15: 122-126.

Petit, A., Sirot, D.L., Chanal, C.M., Sirot, J.L., Labia, R., Gerbaud, G. and Cluzel, R.A. (1988). Novel plasmid-mediated b-Lactamases in clinical isolates of Klebsiella pneumoniae more resistant to ceftazidine than to other broad-spectrum cephalosporins. Antimicrob. Agents Chemother 32: 626-630.

Roggendorf, M. and Muller, H.E. (1976). Enterobacteria of reptiles. Zbl. Bakt. I. Abt. orig. A. 236: 22-35. Southern, E.M. (1975). Detection of specific sequences among DNA fragments separated by gel electrophoresis. J. Mol. Biol. 98: 503-517.

van der Waaij, D., Cohen, B.J. and Nace, G.W. (1974). Colonization pattern of aerobic gram-negative bacteria in the cloacae of Rana pipiens. Lab. Anim. Sci. 24: 307-317.

von Wright, A., Suominen, M. and Sivela, S. (1986). Identification of lactose fermentation plasmids of streptococcal dairy starter strains by Southern hybridization. Letters in Appl. Microbiol. 2: 73-76.

Willets, N. and Wilkins, B. (1984). Processing of plasmid DNA during bacterial conjugation. Microbiol. Rev. 48: 24-41.

Zalmer, H. and Maas, W.K. (1972). Biology of antibiotics. (Springer Verlag Inc., ed.). New York.