Molecular characterization of antibiotic resistance in clinical Salmonella typhi isolated in Ghana

Abstract

Fifty-eight clinical Salmonella typhi strains isolated from patients suspected of suffering from typhoid fever were obtained at the KorleBu Teaching Hospital and the Noguchi Memorial Institute for Medical Research, both located in Ghana, Africa. Each isolate was examined for susceptibility to ampicillin, chloramphenicol, streptomycin, tetracycline, and trimethoprim/sulfamethoxazole by the disk diffusion assay. Five of the isolates were resistant to all five antibiotics while 10 isolates were resistant to ampicillin, chloramphenicol, and trimethoprim/sulfamethoxazole, which are considered 'first line' antibiotics in the treatment of typhoid fever. Thirty-four isolates were resistant to at least one of the antibiotics tested and 62% of these isolates possessed conjugable plasmids belonging to incompatibility group IncHI. Ninety percent of the conjugable plasmids conferred a multiple drug-resistant phenotype on the strains harboring them. Additionally, 14 strains contained plasmids that were transformable and six of them encoded multiple drug resistance. Our findings indicate that multiple drug resistance to the 'first line' antibiotics in S. typhi may be more prevalent in Africa than previously thought.

Characterization of Multidrug-Resistant Typhoid Outbreaks in Kenya

“…Plasmids isolated from both the donor serovar Typhi, and their transconjugants were subjected to PCR to determine whether they belonged to the IncHI1 incompatibility group. The RepHI1A replicon, present in IncHI plasmids, was amplified by using PCR with the primers 5Ј-GGTCCAACCCATTGCTTTAC-3Ј and 5Ј-CACGGAAAGAA ATCACAAC-3Ј as previously reported (8,14) on a Techgene MiniCycler (Techne, Inc., Princeton, N.J.). Reaction conditions consisted of 50 ng of plasmid DNA and 100 nM concentrations of each primer in a buffer composed of 10 mM Tris-HCl (pH 8.3), 50 mM KCl, 1.5 mM MgCl 2 , a 200 M concentration of deoxynucleoside triphosphate mixture, and 1 U of Taq polymerase in a final volume of 100 l. Amplification conditions were 30 cycles of 94°C for 30 s, 55°C for 30 s, and 72°C for 30 s, with a final extension step of 72°C for 10 min.…”

“…Reaction conditions consisted of 50 ng of plasmid DNA and 100 nM concentrations of each primer in a buffer composed of 10 mM Tris-HCl (pH 8.3), 50 mM KCl, 1.5 mM MgCl 2 , a 200 M concentration of deoxynucleoside triphosphate mixture, and 1 U of Taq polymerase in a final volume of 100 l. Amplification conditions were 30 cycles of 94°C for 30 s, 55°C for 30 s, and 72°C for 30 s, with a final extension step of 72°C for 10 min. PCR products were resolved by electrophoresis on 1.2% gels at 120 V run for 1 h. Amplicons of 365 bp were considered positive for the RepHI1A replicon, as previously reported (14,25).…”

“…Typhoid fever, caused by Salmonella enterica serovar Typhi is endemic in most parts of Central America (7,20), Southeast Asia (12,15,16), and the Indian subcontinent (23,25), and recently increasing numbers of cases have been reported in Africa (13,14). Globally, it is estimated that typhoid causes over 16 million cases of illness each year, resulting in over 600,000 deaths (31).…”

“…Thereafter, several cases of MDR serovar Typhi also resistant to nalidixic acid and the fluoroquinolones have been reported in Bangladesh (2), India (18) Thailand, Vietnam (21), and Tajikistan (17), raising concerns about further spread to other regions where typhoid fever is endemic. In addition, molecular characterization of the serovar Typhi outbreak strains revealed that resistance to commonly used antimicrobial agents, including chloramphenicol, ampicillin, and trimethoprim, was encoded by plasmids of the HI incompatibility group (14,16,24).…”

Antibiogram and characterization of resistance markers among Escherichia coli Isolates from urinary tract infections

“…Transfer of resistance was assayed using a standard rifampin-resistant E. coli recipient, CGSC 6576 (F -, λ -, recA1, IN(rrnD-rrnE)1, rpoB331(rifR), hsdR19), kindly supplied by Dr. John Wertz from Coli Genetic Stock Centre, Yale, USA), according to the method described by Robertson and colleagues [14]. Plasmid DNA was isolated as described by Engebrecht et al [15] and transformation was performed using a HMS174 (F -recA1 hsdR(r K12 -m K12 + ) (Rif R ) competent cell kit (Novagen, Darmstadt, Germany) per the manufacturer's instructions.…”