Development of a Synthetic Minimal Medium for
            <i>Listeria monocytogenes</i>

Tsai, Jerry; Hodgson, David A.

doi:10.1128/aem.69.11.6943-6945.2003

Cited by 114 publications

(164 citation statements)

References 14 publications

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“…To see if CcpC Lm acts as a repressor of citB Lm and lmo0847 in vivo, a ccpC null mutation was introduced into strains carrying the citB Lm -lacZ or lmo0847-lacZ fusion. Wild-type (HKB214) and ccpC mutant (HKB217) strains were grown at 30°C in HTM defined medium containing glucose as the sole carbon source and supplemented with hemin (0.2 g/ml) and glutamine (1.6 mg/ml) (57). ␤-Galactosidase activity was measured in samples collected during exponential growth phase.…”

Section: Resultsmentioning

confidence: 99%

CcpC-Dependent Regulation ofcitBand lmo0847 inListeria monocytogenes

2006

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In Bacillus subtilis, the catabolite control protein C (CcpC) plays a critical role in regulating the genes encoding the enzymes of the tricarboxylic acid branch of the Krebs citric acid cycle. A gene encoding a potential CcpC homolog and two potential target genes were identified in the Listeria monocytogenes genome. In vitro gel mobility shift assays and DNase I footprinting experiments showed that L. monocytogenes CcpC (CcpC Lm ) interacts with the promoter regions of citB Lm (the gene that is likely to encode aconitase) and lmo0847 (encoding a possible glutamine transporter) and that citrate is a specific inhibitor of this interaction. To study in vivo promoter activity, a new lacZ reporter system was developed. This system allows stable integration into the chromosome of a promoter region transcriptionally fused to a promoterless lacZ gene at a nonessential, ectopic locus. Analysis of strains carrying a citB Lm -lacZ or lmo0847-lacZ fusion revealed that CcpC Lm represses citB Lm and lmo0847 in media containing an excess of glucose and glutamine. In addition, regulation of citB Lm expression in rich medium was growth phase dependent; during exponential growth phase, expression was very low even in the absence of CcpC Lm , but a higher level of citB Lm expression was induced in stationary phase, suggesting the involvement of another, as yet unidentified regulatory factor.Listeria monocytogenes is a gram-positive, facultative, intracellular pathogen responsible for listeriosis, a serious infection with a mortality rate of up to 30%, mainly among pregnant women, their fetuses, and immunocompromised persons. Listeriosis is characterized by gastroenteritis and fetoplacental and central nervous system infection, resulting in spontaneous abortion, neonatal death, septicemia, and meningitis (58). L. monocytogenes is widespread in nature and is predominantly transferred to humans by ingestion of contaminated foods (13). The bacterium can invade and multiply inside a wide range of phagocytic and nonphagocytic mammalian cells and has served as an important model system for studying the mechanisms of both intracellular parasitism and host cell biology (7,40,41).A great deal of effort has been invested in isolating L. monocytogenes virulence genes and understanding mechanisms of pathogenesis (reviewed in reference 58). However, our knowledge of the basic physiology of this important pathogen is very limited, making it difficult to gain a comprehensive understanding of its pathogenesis. There has been increasing evidence that regulation of carbon metabolic pathways plays a critical role in the virulence of pathogenic bacteria. Regulation of the Salmonella enterica virulence operon spv is controlled by cyclic AMP-cyclic AMP receptor protein complex, the major catabolite regulator, as well as by the positive regulator SpvR (38). Similarly, Escherichia coli STb enterotoxin production is repressed by glucose, and repression is relieved by addition of cyclic AMP (4). Leukotoxin production in Actinobacillus actinomycetemcomitan...

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Section: Resultsmentioning

confidence: 99%

CcpC-Dependent Regulation ofcitBand lmo0847 inListeria monocytogenes

2006

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“…monocytogenes grows in the MM used only when suitable carbon sources are added (Premaratne et al, 1991;Tsai & Hodgson, 2003). Growth in MM with glucose, mannose and especially glycerol leads to considerably higher PrfA activity in the wild-type strain than growth in BHI or LB with the same carbon substrates.…”

Section: Discussionmentioning

confidence: 99%

Modulation of PrfA activity in Listeria monocytogenes upon growth in different culture media

et al. 2008

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PrfA is the major transcriptional activator of most virulence genes of Listeria monocytogenes. Its activity is modulated by a variety of culture conditions. Here, we studied the PrfA activity in the L. monocytogenes wild-type strain EGD and an isogenic prfA deletion mutant (EGDDprfA) carrying multiple copies of the wild-type prfA or the mutant prfA* gene (strains EGDDprfApPrfA and EGDDprfApPrfA*) in response to growth in brain heart infusion (BHI), Luria-Bertani broth (LB) or a defined minimal medium (MM) supplemented with one of the three phosphotransferase system (PTS) carbohydrates, glucose, mannose and cellobiose, or the non-PTS carbon source glycerol. Low PrfA activity was observed in the wild-type strain in BHI and LB with all of these carbon sources, while PrfA activity was high in minimal medium in the presence of glycerol. EGDDprfApPrfA*, expressing a large amount of PrfA* protein, showed high PrfA activity under all growth conditions. In contrast, strain EGDDprfApPrfA, expressing an equally high amount of PrfA protein, showed high PrfA activity only when cultured in BHI, and not in LB or MM (in the presence of any of the carbon sources). A ptsH mutant (lacking a functional HPr) was able to grow in BHI but not in LB or MM, regardless of which of the four carbon sources was added, suggesting that in LB and MM the uptake of the used PTS carbohydrates and the catabolism of glycerol are fully dependent on the functional common PTS pathway. The BHI culture medium, in contrast, apparently contains carbon sources (supporting listerial growth) which are taken up and metabolized by L. monocytogenes independently of the common PTS pathway. The growth rates of L. monocytogenes were strongly reduced in the presence of large amounts of PrfA (or PrfA*) protein when growing in MM, but were less reduced in LB and only slightly reduced in BHI. The expression of the genes encoding the PTS permeases of L. monocytogenes was determined in the listerial strains under the applied growth conditions. The data obtained further support the hypothesis that PrfA activity correlates with the expression level and the phosphorylation state of specific PTS permeases.

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“…This strain belongs to serovar 1/2a, a serovar commonly isolated from food preparation sites and one of the two serovars most commonly associated with human listeriosis (15). M35303A is a cysteine auxotroph, hence minimal medium was supplemented with 0.1 mg/ml cysteine (3,27). The wild-type strain 10403S was grown in minimal medium supplemented with both cysteine and methionine at 0.1 mg/mg each.…”

Section: Methodsmentioning

confidence: 99%

Flagellar Motility Is Critical forListeria monocytogenesBiofilm Formation

2007

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The food-borne pathogen Listeria monocytogenes attaches to environmental surfaces and forms biofilms that can be a source of food contamination, yet little is known about the molecular mechanisms of its biofilm development. We observed that nonmotile mutants were defective in biofilm formation. To investigate how flagella might function during biofilm formation, we compared the wild type with flagellum-minus and paralyzed-flagellum mutants. Both nonmotile mutants were defective in biofilm development, presumably at an early stage, as they were also defective in attachment to glass during the first few hours of surface exposure. This attachment defect could be significantly overcome by providing exogenous movement toward the surface via centrifugation. However, this centrifugation did not restore mature biofilm formation. Our results indicate that it is flagellum-mediated motility that is critical for both initial surface attachment and subsequent biofilm formation. Also, any role for L. monocytogenes flagella as adhesins on abiotic surfaces appears to be either minimal or motility dependent under the conditions we examined.

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Development of a Synthetic Minimal Medium for Listeria monocytogenes

Cited by 114 publications

References 14 publications

CcpC-Dependent Regulation ofcitBand lmo0847 inListeria monocytogenes

CcpC-Dependent Regulation ofcitBand lmo0847 inListeria monocytogenes

Modulation of PrfA activity in Listeria monocytogenes upon growth in different culture media

Flagellar Motility Is Critical forListeria monocytogenesBiofilm Formation

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