The prevalence of antimicrobial resistance (AMR) phenotypes and genotypic characteristics (Class 1 integron and AMR gene cassettes) in commensal Escherichia coli (EC) and vancomycin resistant Enterococcus faecium (EF) isolated from humans and swine in a semi-closed, integrated farrow-to-fork population were evaluated in a crosssectional study. Our objective was to establish baseline antimicrobial resistance patterns and to evaluate the stability of isolate recovery phenotype within multiple grab samples per collection day and over multiple biweekly samples collected during a period of several months. This data will serve as a baseline for continuing longitudinal studies within the population. These continuing studies should produce the first comprehensive epidemiological data to document the transmission dynamics of antimicrobial resistance in the farrow-to-fork continuum. Outcome variables assessed included: phenotypic resistance in EC, pan-susceptibility, multi-resistance and genotypic resistance. Potential predictor variables included: 1) host species, 2) unit, 3) unit type, 4) housing cohort by species, and 5) time of day. There were significant differences (p<0.05) between host species with swine at higher odds for both single and multiple resistance. There were also differences in resistance based on unit location, unit-type, and housing cohort within both humans and swine. Our study found no significant differences (p>0.05) in resistance between swine workers and non-swine workers with the sole exception of resistance to cephalothin, with non-swine workers at 1.89 higher odds for resistance (p=0.02). A total of 17 VRE were isolated from human wastewater samples, and to the author’s knowledge these represent the first environmentally isolated VRE in the U.S. Several unique multi-resistance phenotypes were observed and future evaluation of AMR phenotype in continuing longitudinal studies provides a unique opportunity to study phenotypic patterns and dissemination through the study population.

A bstract Few in vitro screening studies on the biological activities of plant extracts that are intended for oral administration consider the effect of the gastrointestinal system. This study investigated this aspect on extracts of Camellia sinensis (green tea) and Salvia officinalis (sage) using antimicrobial activity as amodel for demonstration. Both the crude extracts and their products after exposure to simulated gastric fluid (SGF) as well as simulated intestinal fluid (SIF) were screened for antimicrobial activity. The chromatographic profiles of the crude plant extracts and their SGF as well as SIF products were recorded and compared qualitatively by means of high performance liquid chromatography coupled to mass spectrometry. The effect of epithelial transport on the crude plant extracts was determined by applying them to an in vitro intestinal epithelial model (Caco-2). The crude extracts for both plants exhibited reduced antimicrobial activity after exposure to SGF, while no antimicrobial activity was detected after exposure to SIF. These results suggested chemical modification or degradation of the antimicrobial compounds when exposed to gastrointestinal conditions. This was confirmed by a reduction of the peak areas on the LC–UV–MS chromatograms. From the chromatographic profiles obtained during the transport study, it is evident that some compounds in the crude plant extracts were either not transported across the cell monolayer or they were metabolised during passage through the cells. It can be deduced that the gastrointestinal environment and epithelial transport process can dramatically affect the chromatographic profiles and biological activity of orally ingested natural products.

Abstract Few in vitro screening assays for biological activities of plant extracts consider the potential effect of the gastrointestinal system on orally consumed plant extracts. Crude water and methanol extracts of Tarchonanthus camphoratus (wild camphor) and Agathosma betulina (‘buchu’) were prepared and exposed to simulated gastric fluid and simulated intestinal fluid during dissolution studies to address this aspect. The crude extracts and resulting simulated gastric fluid and simulated intestinal fluid products were screened for antimicrobial activity against Staphylococcus aureus (ATCC 25923), Enterococcus faecalis (ATCC 29212), Escherichia coli (ATCC 25922) and Proteus vulgaris (ATCC 33420). The T. camphoratus crude extract exhibited antimicrobial activity which was reduced after exposure to simulated gastric fluid. After exposure to simulated intestinal fluid no antimicrobial activity was detected, which suggests chemical alteration or degradation of the active compounds. For A. betulina, the crude water extract and simulated gastric fluid product exhibited no antimicrobial activity, while the simulated intestinal fluid product exhibited antimicrobial activity. This suggests activation of antimicrobial constituents during exposure to simulated intestinal fluid. The chemical composition profiles of the crude extracts and products were determined by means of liquid chromatography coupled to an ultraviolet detector (LC-UV) and a mass spectrometer (LC-MS) to qualitatively assess the effect of exposure to simulated gastrointestinal conditions on the chemical composition of the extracts. In many cases, the peak area of compounds decreased after exposure to simulated gastric fluid and simulated intestinal fluid, while the peak area of other compounds increased. Thus, it can be deduced that the antimicrobial activity and chemical composition was altered after exposure to intestinal conditions during dissolution studies.

Aim of the study: In traditional healing, the burning of selected indigenous medicinal plants and the inhalation of the liberated smoke are widely accepted and a practiced route of administration. This study elucidated the rationale behind this commonly practiced treatment by examining the antimicrobial activity for five indigenous South African medicinal plants commonly administered through inhalation (Artemisia afra, Heteropyxis natalensis, Myrothamnus flabellifolius, Pellaea calomelanos and Tarchonanthus camphoratus). Material and Methods: An apparatus was designed to simulate the burning process that occurs in a traditional setting and the smoke fraction was captured for analysis and bioassay. Methanol and acetone extracts as well as the essential oil (for the aromatic species) were prepared and assayed in parallel with the smoke fraction. Results: Antimicrobial data revealed that in most cases, the ‘smoke-extract’ obtained after burning had lower minimum inhibitory concentration (MIC) values than the corresponding solvent extracts and essential oils. The combustion, acetone and methanol extracts produced different chromatographic profiles as demonstrated for Pellaea calomelanos where several compounds noted in the smoke fraction were not present in the other extracts. Conclusion: These results suggest that the combustion process produces an ‘extract’ with superior antimicrobial activity and provides in vitro evidence for inhalation of medicinal smoke as an efficient mode of administration in traditional healing.

Heteropyxis natalensis (Heteropyxidaceae) is traditionally used to treat respiratory disorders, and as a decongestant and antimicrobial agent. The seasonal variation of the hydrodistilled essential oil was investigated. Three trees in the Johannesburg Botanical Garden (Gauteng) indicated similar chemical profiles with fluctuation in the levels of the two major constituents (1,8-cineole and limonene). Little variation between the antimicrobial activity of seasonally collected samples was documented, with standard deviations of ±0.3 to ±3.3 depending on the pathogen studied. Moderate antimicrobial activity (3.0–16.0 mg/ml) was noted for most pathogens tested with Cryptococcus neoformans exhibiting the highest sensitivity (2.0–3.0 mg/ml). The chemogeographical variation of the oil composition from five of the seven distinct localities studied all contains 1,8-cineole and limonene as major constituents. The antimicrobial study of these samples indicated little variability between localities (standard deviation of ±0.5 to ±3.8). As observed in the seasonal variation study, C. neoformans displayed the highest sensitivity (0.5–2.0 mg/ml). One oil sample (Lagalametse), was distinctly different both chemically and microbiologically.

No description available.

Bacterial resistance to antibiotics has increased rapidly within recent years, and it has become a serious threat to public health. Infections caused by multi-drug resistant bacteria entail higher morbidity, mortality, and a burden to health care systems. The use of biocides, including silver compounds, may affect the resistance to both biocides and antibiotics and, thereby, can be a driving factor in this development. The aim of the following thesis was to investigate the frequency of silver resistance and the effects of silver exposure on bacterial populations being of clinical significance and from geographically different parts of the world. Furthermore, it explored the genetic background of silver resistance, and if silver could select directly or indirectly for antibiotic resistance. By a range of methods, from culture in broth to whole genome sequencing, bacterial populations from humans, birds and from the environment were characterized. The studies showed that sil genes, encoding silver resistance, occurred at a high frequency. Sil genes were found in 48 % of Enterobacter spp., in 41 % of Klebsiella spp. and in 21 % of all human Escherichia coli isolates with production of certain types of extended-spectrum beta-lactamases (CTX-M-14 and CTX-M-15). In contrast, silver resistance was not found in bird isolates or in bacterial species, such as Pseudomonas aeruginosa and Legionella spp., with wet environments as their natural habitat. One silver-resistant Enterobacter cloacae strain was isolated from a chronic leg ulcer after only three weeks of treatment with silver-based dressings. The in-vivo effects of these dressings were limited, and they failed to eradicate both Gram-positive and Gram-negative bacteria. The activity of silver nitrate in vitro was bacteriostatic on Gram-positive species such as S. aureus and bactericidal on Gram-negative species. In Enterobacteriaceae, sil genes were associated with silver resistance phenotypes in all but one case. Using whole genome sequencing, single nucleotide polymorphisms in the silS gene were discovered after silver exposure in isolates with expressed silver resistance. This resistance could co-select for resistance to beta-lactams, co-trimoxazole and gentamicin. The findings of this thesis indicate that silver exposure may cause phenotypic silver resistance, and it may reduce the susceptibility to mainly beta-lactams and select for bacteria with resistance to clinically important antibiotics.

No description available.

Experiments detailed in this thesis investigated the bacterial response towards electrophiles and how this could be exploited as a novel target for antimicrobial chemotherapy. The principal conclusions are: (1) The root colonising bacterium <I>P. putida</I> does not appear to have a potassium efflux system similar to KefB and KefC. The exposure of log-phase <I>P. putida</I> cultures to sublethal concentrations of NEM resulted in a temporary viable non-culturable state, which could be suppressed by the presence of specific peptides in the recovery medium. Stationary phase cultures were more resistant to NEM, partially due to a faster metabolism, and did not undergo the temporary viable but non-culturable state. (2) Micotitre assays suitable for automated screening were developed to screen the Zeneca chemical library for activators of KefB and KefC and for activators and inhibitors of methylglyoxal synthase (MGS). (3) MGS was partially purified for <I>E. coli</I> and the N-terminal amino acid sequence was determined. This led to the identification of the gene encoding MGS, <I>mgsA</I>. An MGS over-expression vector and a chromosomal knock-out strain were constructed. (4) The over-expression of MGS resulted in an accumulation of MGS to approximately 25% of the total cell protein, corresponding to up to 900-fold enhanced MGS activity in crude extract. High level MGS expression had little effect on growth and viability of cells growth in LB, K₁₀ glucose or medium limited in phosphate. Severe growth inhibition was observed in cells over-expressing MGS in K₁₀ with either glycerol or xylose as the sole carbon source. (5) An MGS-deficient strain grew normally in LB, on glucose, glycerol, xylose, and in medium limited in phosphate or nitrogen. The MGS knock-out strain was severely inhibited during elevated xylose metabolism in the absence of methylglyoxal (MG) production. This suggests a prominent role for MGS in adaptation from "famine" (limited carbon) to "feast" (elevated carbon metabolism) conditions.

No description available.