1 WHO Collaborating Centre for Antimicrobial Resistance in Bacteria from Food animals and Food of Animal Origin, Danish Veterinary Institute, Bülowsvej 27, DK-1790 Copenhagen V, Denmark; 2 Center for Antimicrobial Resistance Monitoring in Food-borne Pathogens. Faculty of Veterinary Science, Chulalongkorn University, Bangkok; 3 WHO International Salmonella and Shigella Centre, National Institute of Health, Department of Medical Sciences, Ministry of Public Health, Bangkok, Thailand
Received 16 June 2003; returned 27 June 2003; revised 17 July 2003; accepted 23 July 2003
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Abstract |
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Methods: A total of 503 isolates were examined for susceptibility to antimicrobial agents, and resistant isolates were examined for the presence of selected resistance genes by PCR.
Results: Only 48 (9.5%) of the isolates were resistant to one or more of the antimicrobial agents tested. A low frequency of resistance was found towards ampicillin (1.8%), chloramphenicol (1.6%), florphenicol (0.4%), nalidixic acid (1.6%), neomycin (0.6%), streptomycin (4.4%), sulfamethoxazole (4.2%), tetracycline (4.0%) and trimethoprim (1.4%), whereas all isolates were susceptible to co-amoxiclav, ceftiofur, ciprofloxacin, colistin and gentamicin. All nine ampicillin-resistant isolates contained a sequence similar to the blaTEM-1b gene, one of the eight chloramphenicol-resistant isolates a sequence similar to the catA1 gene, all three neomycin-resistant isolates a sequence similar to the aphA-2 gene, 16 (73%) of the 22 streptomycin-resistant isolates a sequence similar to the aadA gene, the remaining six (27%) a sequence similar to the strA gene, and all 21 sulfamethoxazole-resistant isolates a sequence similar to the sul2 gene. Thirteen (65%) of the 20 tetracycline-resistant isolates contained the tet(A) gene, four (20%) the tet(B) gene, and one (5%) the tet(C) gene.
Conclusions: This study showed a low frequency of resistance among Salmonella Weltevreden isolated from humans and other reservoirs in South-East Asia and elsewhere. There was no major difference in the occurrence of resistance between source or geographical origin.
Keywords: Salmonella Weltevreden, antimicrobial resistance, genes, South-East Asia
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Introduction |
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In recent years, an increase in the occurrence of antimicrobial resistance among S. enterica has been observed in several countries, but there is only limited information on the occurrence of antimicrobial resistance among Salmonella Weltevreden. Boonmar et al.2 reported a very low frequency of resistance among 111 Salmonella Weltevreden from human infections in 1993 in Thailand, which was in contrast to the frequent occurrence of resistance observed among other serovars. However, in 1994, they reported a much higher occurrence of resistance among 139 isolates. Thong et al.3 examined 95 isolates from different sources in Malaysia and found a low frequency of resistance.
This study was conducted to investigate the occurrence of antimicrobial resistance, and the distribution of resistance genes, among 503 Salmonella Weltevreden isolates from different sources in 10 countries, most of them from South-East Asia.
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Materials and methods |
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A total of 503 isolates were received from laboratories in South-East Asia [Indonesia (45), Laos (6), Malaysia (105), Taiwan (7), Thailand (197), Vietnam (19)], Australia (46), Denmark (16), New Zealand (40) and the USA (22). The isolates were collected during 19952001. Most isolates originated from humans (282), but some were from animals (59), food products (114) and water, the environment or other sources (48).
Susceptibility testing
Susceptibility to antimicrobial agents was performed as MIC determinations. As described in NCCLS guidelines,4 a commercially prepared, dehydrated panel (Sensititre) was used for the following antimicrobial agents: ampicillin, ceftiofur, chloramphenicol, ciprofloxacin, co-amoxiclav, colistin, florphenicol, gentamicin, nalidixic acid, neomycin, streptomycin, sulfamethoxazole, tetracycline and trimethoprim.
Detection of resistance genes
PCR was used to detect the presence of antimicrobial resistance genes. Ampicillin-resistant isolates were examined for blaTEM genes using primer pairs 5'-ATGAGTATTCAACATTTCCG-3' (blaTEM) and 5'-AC-CAATGCTTAATCAGTGAG-3' (blaTEM); chloramphenicol-resistant isolates were examined for catA1 using primers 5'-CGCCTGATGAATGCTCATCCG-3' (catA1) and 5'-CCTGCCACTCATCGCAG TAC-3' (catA1); neomycin-resistant isolates were examined for aphA-2 using primer pairs 5'-GCTATTCGGCTATGACTGGGC-3' (aphA-2) and 5'-CCACCATGATATTCGGCAAGC-3' (aphA-2); streptomycin-resistant isolates were examined for strA and aadA using primer pairs 5'-CCAATCGCAGATAGAAGGC-3' (strA), 5'-CTTGGTGATAACG-GCAATTC-3' (strA), 5'-ATCCTTCGGCGCGATTTTG-3' (aadA) and 5'-GCAGCGCAATGACATTCTTG-3' (aadA); sulfamethoxazole resistant isolates were examined for sul1 and sul2 using primer pairs 5'-CTTCGATGAGAGCCGGCGGC-3' (sul1), 5'-GCAAGGCGGAAACCCGC GCC-3' (sul1), 5'-GCGCTCAAGGCAGATGGCATT-3' (sul2) and 5'-GCGTTTGATACCGGCACCCGT-3' (sul2); and tetracycline-resistant isolates were examined for tet(A), tet(B), tet(C) and tet(G) using primer pairs 5'-GTAATTCTGAGCACTGTCGC-3' [tet(A)], 5'-CTGCCTGGACAACATTGCTT-3' [tet(A)], 5'-CTCAGTATTCCAAGCCTTTG-3' [tet(B)], 5'-ACTCCCCTGAGCTTGAGGGG-3' [tet(B)], 5'-GGTTGAAGGCTCTCAAGGGC-3' [tet(C)], 5'-CCTCTTGCGGGAAT- CGTCC-3' [tet(C)], 5'-GCAGCGAAAGCGTATTTGCG-3' [tet(G)] and 5'-TCCGAAAGCTGTCCAAGCAT-3' [tet(G)].
Using the same primers as in the PCR analysis, DNA sequencing verified the identity of the gene products in selected isolates. The gene product of all blaTEM-positive isolates was sequenced.
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Results |
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Discussion |
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A number of different resistance genes were detected among the 48 resistant isolates. All ampicillin-resistant isolates contained a sequence similar to blaTEM-1b. blaTEM genes have been found previously among Salmonella isolates.5 The catA1 gene has been detected previously in Salmonella isolates,6 but is also widespread among other Gram-negative bacteria. Tetracycline resistance was mainly mediated by tet(A), but a few isolates contained tet(B) and a single isolate contained tet(C). tet(A) is located frequently on transposons such as Tn1721, and the gene has been found widespread among Gram-negative bacteria including salmonella.7 Resistance to sulfamethoxazole was exclusively mediated by sul2. In the past, the sul1 gene has been found mainly in connection with integrons, whereas sul2 is located usually on a variety of plasmids.8 All neomycin-resistant isolates contained aphA-2, which also has been found in several Gram-negative species. Both aadA and strA have been found frequently among streptomycin-resistant isolates, as has Salmonella Typhimurium in Denmark.9 Class I integrons containing the aadA gene have been observed often among clinically important Enterobacteriaceae.10 However, the class I integrons normally contain sul1; this gene was not observed in any strains in this study.
In conclusion, this study showed a low frequency of resistance among Salmonella Weltevreden isolated from humans and other reservoirs in South-East Asia and elsewhere. Resistance was encoded by genes previously widespread in other Enterobacteiaceae. No major differences between country or source could be observed.
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Acknowledgements |
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Footnotes |
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References |
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2 . Boonmar, S., Bangtrakulnonth, A., Pornruangwong, S. et al. (1998). Predominant serovars of Salmonella in humans and foods from Thailand. Journal of Veterinary Medical Sciences 60, 87780.[CrossRef]
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Thong, K. L., Goh, Y. L., Radu, S. et al. (2002). Genetic diversity of clinical and environmental strains of Salmonella enterica serotype Weltevreden isolated in Malaysia. Journal of Clinical Microbiology 40, 24982503.
4 . National Committee for Clinical Laboratory Standards. (2002). Performance Standards for Antimicrobial Disk and Dilution Susceptibility Tests for Bacteria Isolated from AnimalsSecond Edition: Approved Standard M31-A2. NCCLS, Villanova, PA, USA.
5 . Gallardo, F., Ruiz, J., Marco, F. et al. (1999). Increase in incidence of resistance to ampicillin, chloramphenicol and trimethoprim in clinical isolates of Salmonella serotype Typhimurium with investigation of molecular epidemiology and mechanisms of resistance. Journal of Medical Microbiology 48, 36774.[Abstract]
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