Conventional Susceptibility Testing Methods Research Articles

Direct antibiotic susceptibility testing of blood cultures of gram-negative bacilli using the Drug Susceptibility Testing Microfluidic (DSTM) device

Proper treatment of bloodstream infections requires rapid, early determination of appropriate antibiotic agents, emphasizing the need for more rapid drug susceptibility testing. The Drug Susceptibility Testing Microfluidic (DSTM) device represents a novel method in which a small amount of bacterial suspension is injected into the microchip-like device and cultured for 3 h. However, it remains unknown whether the DSTM method can directly determine antibiotic susceptibilities from positive blood cultures. Here, we developed a new approach to directly assess drug susceptibility, using the DSTM method for positive blood cultures. We compare the utility and accuracy of DSTM with those of conventional susceptibility testing methods. Fifty positive blood cultures identified as gram-negative bacilli were used herein. The outcomes of drug susceptibility and resistance assays for positive blood cultures were compared to those of conventional susceptibility testing methods to evaluate their utility and accuracy. Method agreement rates between DSTM and standard methods often exceed 90%, suggesting a high positive correlation with conventional methods. Furthermore, our results show that a combination of multiple drugs in the DSTM device helps identify extended-spectrum β-lactamase (ESBL)- and AmpC-β-lactamase (AmpC-)-producing microorganisms. In conclusion, DSTM method enables effective drug susceptibility and resistance screening within 3 h from positive blood cultures and is suitable for the rapid and personalized determination of the antimicrobial regimen.

Antibacterial Effects of Methanol Extract of Bryophyllum pinnatum L on Methicillin Resistant Staphylococcus aureus (MRSA) Isolated from Urine

There are major concerns about rising levels of methicillin resistant Staphylococcus aureus (MRSA). This is due to the difficulties in treating the infections which they cause and .the ease with which they spread in hospitals. This has necessitated the continuous search for alternative anti-MRSA agents. Efforts in this study was therefore directed at isolation of MRSA from the urine of patients and its susceptibility to the methanol extract and aqueous fraction of Bryophyllum pinnatum . Urine samples from the urine of patients were screened for the presence of Staphylococcus aureus using conventional microbiological methods. Confirmed isolates were screened for methicillin resistance by confirming their susceptibility or otherwise to 30µg cefoxitin. Detection of Mec A gene by Polymerase chain reaction (PCR) was further used to confirm some MRSA isolates. Conventional susceptibility testing methods were used to compare the activity of both methanol extract of Brophyllum pinnatum and its aqueous fraction on the MRSA isolates. Results obtained confirmed the susceptibility of the MRSA isolates to the extracts and that their activity was time dependent. It also showed that the extract was only moderately toxic with an LD50 of 866.03mg/kg body weight and that at the MIC and 2xMIC their activity was only bacteriostatic. Results obtained are intended to be used to prove that in the search for alternative anti- MRSA agents from natural sources, Bryophyllum pinnatum will be a possible candidate for further investigation.

Detection of plasmid-mediated AmpC in Escherichia coli, Klebsiella pneumoniae and Proteus mirabilis

Aims:This study investigated the prevalence of plasmid-mediated AmpC production in selected clinical isolates of Escherichia coli, Klebsiella species and Proteus mirabilis, and compared the results of boronic acid disc screening...

Antimycobacterial susceptibility testing: Present practices and future trends

Along with the worldwide renewed interest in tuberculosis, the high incidence of non-tuberculous disease in HIV-infected patients and the continuing problem of disease caused by rapidly growing mycobacteria, there has been a renewed interest in mycobacterial susceptibility testing. This renewed interest stems from the needs both to provide accurate, reliable and timely susceptibility test information for the management of patients infected with these mycobacteria and to identify new and more effective antimycobacterial agents. Indeed, many new agents have already been identified and are currently used to treat these infections, but the conventional susceptibility testing methods, in many instances, have not been adequately evaluated for these new agents. Perhaps the time has come to give consideration to abandoning such time-honored practices and criteria as the proportion method, the 99% threshold, and "critical concentrations". New methods of susceptibility testing are in various stages of development and many of these methods have advantages and improvements over the existing methods. There is an increased understanding of the pathobiology of mycobacterial infections and an increased understanding of the pharmacokinetics of antimycobacterial agents and the mechanisms of action and resistance. This article gives an overview of the present practices and an assessment of the current needs and potential for the near future of antimycobacterial susceptibility testing.

Multipoint microbiological assay for detecting beta-lactamase.

A multipoint microbiological assay for determining beta-lactamase production by clinical isolates of bacteria was evaluated. With strains of Haemophilus influenzae, Neisseria gonorrhoeae, and Branhamella catarrhalis there was excellent correlation between results obtained using this method and those obtained using the chromogenic cephalosporin reference method. The multipoint method is an inexpensive yet reliable adjunct to conventional susceptibility testing methods.

Activity of amphotericin B and itraconazole against intraphagocyticCandida albicans

The activity of amphotericin B and itraconazole against intracellularCandida albicans was determined in vitro using murine resident peritoneal macrophages. Amphotericin B at concentrations of 0.5 and 2 µg/ml produced significantly less rapid killing of intracellular than of extracellularCandida albicans as measured in macrophage-free medium. Amphotericin B at concentrations of 0.1 µg/ml or itraconazole concentrations of up to 3 µg/ml produced only fungistatic or limited fungicidal activity against both intracellular and extracellular organisms. Against intracellularCandida albicans amphotericin B acted by direct antifungal action rather than through stimulation of macrophage function, as demonstrated by the fact that (i) activity persisted when macrophages were successively exposed to amphotericin B, washed and disrupted by sonication, and (ii) no activity was seen when amphotericin B was tested against intracellular amphotericin B-resistantCandida tropicalis orSalmonella typhimurium. Pre-exposure of macrophages to itraconazole (0.4 µg/ml) inhibited subsequent killing activity of amphotericin B (2 µg/ml) against intracellular susceptibleCandida albicans. These experiments validate the conventional methods of susceptibility testing for determining the fungistatic activity of antifungal agents.

Emergence of resistance during beta-lactam therapy of gram-negative infections. Bacterial mechanisms and medical responses.

Some Gram-negative, non-fastidious bacilli, although classified as susceptible by conventional susceptibility testing methods, become resistant during therapy with the newer beta-lactam compounds. Emergence of resistance results primarily from the selection of resistant clones pre-existing within the susceptible bacterial populations. Most of the resistant clones produce large amounts of beta-lactamases which inhibit the beta-lactam antibiotics by hydrolysis, rather than by binding. In addition, resistant clones can limit the penetration of beta-lactam molecules through the outer membrane by a decreased expression of their porins. Less commonly, when beta-lactamase activity together with alteration of the permeability barrier does not prevent the access of the antibiotic molecules to their target, altered penicillin-binding proteins (PBPs) can produce resistance. However, the risk of resistance emerging during therapy varies with the beta-lactam drug administered. Some compounds such as cefpirome, BMY 28142, SCH 34343, or imipenem appear to be associated with a low risk. In addition, emergence of resistance can be reduced by using higher dosages of beta-lactam agents, or by combining them with other drugs such as aminoglycosides or quinolones.