Browsing by Author "Dicks, Leon M. T."

Now showing 1 - 5 of 5
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    Development of a novel selection / counter‑selection system for chromosomal gene integrations and deletions in lactic acid bacteria
    (BMC (part of Springer Nature), 2019-03-29) Van Zyl, Winschau F.; Dicks, Leon M. T.; Deane, Shelly M.
    Background: The underlying mechanisms by which probiotic lactic acid bacteria (LAB) enhance the health of the consumer have not been fully elucidated. Verification of probiotic modes of action can be achieved by using singleor multiple-gene knockout analyses of bacterial mutants in in vitro or in vivo models. We developed a novel system based on an inducible toxin counter-selection system, allowing for rapid and efficient isolation of LAB integration or deletion mutants. The Lactococcus lactis nisin A inducible promoter was used for expression of the Escherichia coli mazF toxin gene as counter-selectable marker. Results: The flippase (FLP)/flippase recognition target (FRT) recombination system and an antisense RNA transcript were used to create markerless chromosomal gene integrations/deletions in LAB. Expression of NisR and NisK signalling proteins generated stable DNA integrations and deletions. Large sequences could be inserted or deleted in a series of steps, as demonstrated by insertion of the firefly bioluminescence gene and erythromycin resistance marker into the bacteriocin operons or adhesion genes of Lactobacillus plantarum 423 and Enterococcus mundtii ST4SA. Conclusions: The system was useful in the construction of L. plantarum 423 and E. mundtii ST4SA bacteriocin and adhesion gene mutants. This provides the unique opportunity to study the role of specific probiotic LAB genes in complex environments using reverse genetics analysis. Although this work focuses on two probiotic LAB strains, L. plantarum 423 and E. mundtii ST4SA, the system developed could be adapted to most, if not all, LAB species.
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    Gut bacteria and neuropsychiatric disorders
    (MDPI, 2021-12-14) Dicks, Leon M. T.; Hurn, Diron; Hermanus, Demi; Indrio, Flavia
    Bacteria in the gut microbiome plays an intrinsic part in immune activation, intestinal permeability, enteric reflex, and entero-endocrine signaling. Apart from physiological and structural changes brought about by gut bacteria on entero-epithelial cells and mucus layers, a vast number of signals generated in the gastro-intestinal tract (GIT) reaches the brain via the vagus nerve. Research on the gut–brain axis (GBA) has mostly been devoted to digestive functions and satiety. Less papers have been published on the role gut microbiota play in mood, cognitive behavior and neuropsychiatric disorders such as autism, depression and schizophrenia. Whether we will be able to fully decipher the connection between gut microbiota and mental health is debatable, especially since the gut microbiome is diverse, everchanging and highly responsive to external stimuli. Nevertheless, the more we discover about the gut microbiome and the more we learn about the GBA, the greater the chance of developing novel therapeutics, probiotics and psychobiotics to treat gastro-intestinal disorders such as inflammatory bowel disease (IBD) and irritable bowel syndrome (IBS), but also improve cognitive functions and prevent or treat mental disorders. In this review we focus on the influence gut bacteria and their metabolites have on neuropsychiatric disorders.
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    Molecular insights into probiotic mechanisms of action employed against intestinal pathogenic bacteria
    (Taylor & Francis, 2020-10) van Zyl, Winschau F.; Deane, Shelly M.; Dicks, Leon M. T.
    Gastrointestinal (GI) diseases, and in particular those caused by bacterial infections, are a major cause of morbidity and mortality worldwide. Treatment is becoming increasingly difficult due to the increase in number of species that have developed resistance to antibiotics. Probiotic lactic acid bacteria (LAB) have considerable potential as alternatives to antibiotics, both in prophylactic and therapeutic applications. Several studies have documented a reduction, or prevention, of GI diseases by probiotic bacteria. Since the activities of probiotic bacteria are closely linked with conditions in the host’s GI-tract (GIT) and changes in the population of enteric microorganisms, a deeper understanding of gut-microbial interactions is required in the selection of the most suitable probiotic. This necessitates a deeper understanding of the molecular capabilities of probiotic bacteria. In this review, we explore how probiotic microorganisms interact with enteric pathogens in the GIT. The significance of probiotic colonization and persistence in the GIT is also addressed.
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    Profiling the production of antimicrobial secondary metabolites by xenorhabdus khoisanae J194 under different culturing conditions
    (Frontiers Media, 2020-03-30) Booysen, Elzaan; Rautenbach, Marina; Stander, Marietjie A.; Dicks, Leon M. T.
    Species from the genus Xenorhabdus, endosymbiotic bacteria of Steinernema nematodes, produce several antibacterial and antifungal compounds, some of which are anti-parasitic. In this study, we report on the effect growth conditions have on the production of antimicrobial compounds produced by Xenorhabdus khoisanae J194. The strain was cultured in aerated and non-aerated broth, respectively, and on solid media. Production of antimicrobial compounds was detected after 24 h of growth in liquid media, with highest levels recorded after 96 h. Highest antimicrobial activity was obtained from cells cultured on solid media. By using ultraperformance liquid chromatography linked to mass spectrometry and HPLC, a plethora of known Xenorhabdus compounds were identified. These compounds are the PAX lipopeptides (PAX 1′, PAX 3′, PAX 5, and PAX 7E), xenocoumacins and xenoamicins. Differences observed in the MS-MS fractionation patterns collected in this study, when compared to previous studies indicated that this strain produces novel xenoamicins. Three novel antimicrobial compounds, khoicin, xenopep and rhabdin, were identified and structurally characterized based on MS-MS fractionation patterns, amino acid analysis and whole genome analysis. The various compounds produced under the three different conditions indicates that the secondary metabolism of X. khoisanae J194 may be regulated by oxygen, water activity or both. Based on these findings X. khoisanae J194 produce a variety of antimicrobial compounds that may have application in disease control.
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    Soymilk bio-enrichment by indigenously isolated riboflavin-producing strains of Lactobacillus plantarum
    (Elsevier, 2019-11) Bhushan, Bharat; Kumkum, C. R.; Kumari, Mamta; Ahire, Jayesh J.; Dicks, Leon M. T.; Mishra, Vijendra
    Lactobacilli (n = 68) isolated from human feces and fermented milk products were screened for the production of riboflavin (vitamin B2) by culturing into riboflavin assay medium (RAM). Cell-free culture supernatants from positive isolates (BBC32A, BBC32B, BBC33 and BIF43) were transferred onto RAM agar pre-seeded with Enterococcus faecalis MTCC2729 (a riboflavin auxotroph). The enhanced growth of B2-auxotroph confirmed the bioavailability of produced vitamin. Isolate BBC32B produced the highest riboflavin (319 ± 36 μg/l), followed by BBC33 (304 ± 91 μg/l), BBC32A (276 ± 8 μg/l) and BIF43 (257 ± 91 μg/l). All four isolates contained riboflavin genes ribG, ribB, ribA and ribH. Sequencing of DNA fragments amplified from the 16S–23S rRNA intergenic spacer region and areas flanking the 23S rRNA gene grouped these isolates within the species Lactobacillus plantarum. Identifications were confirmed by sequencing 1300-bp of amplified 16S rDNA fragments. Fermentation of soymilk by single cultures of L. plantarum BBC32B, BBC33 and BIF43 yielded 49.05%, 38.97% and 35.94% riboflavin enrichment respectively, which is more than 18.75% recorded in literature for Lactobacillus fermentum MTCC8711. Maximum levels of riboflavin were obtained within 12 h of fermentation in soymilk. Lactobacillus plantarum BBC32B may be used as starter culture for developing of riboflavin-enriched soymilk.