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Safety evaluation of Lactobacillus delbrueckii subsp. lactis UO 004, a probiotic bacterium.

Lactobacillus delbrueckii subsp. lactis UO 004 was evaluated for its use as a potential probiotic from a safety point of view. The strain did not exhibit mucinolytic or other enzymatic activities that might be detrimental, such as those involving glycosidases (beta-D-glucosaminidase or alpha-D-galactosidase) or arylamidases (factor Xa and quimotrypsin-like activities), frequently present in Lactobacillus strains isolated from patients with endocarditis, although it was able to express protein Ca and kallikrein-like activities. On the other hand, the presence of the strain did not interfere with the growth of certain species of normal intestinal microbiota, such as Enterococcus fecalis, Escherichia coli, Bifidobacterium bifidum or Bacteroides fragilis. Moreover, the potential probiotic strain UO 004 is sensitive to antibiotics with transmissible resistance mechanisms in Lactobacillus such as chloramphenicol, erythromycin, tetracycline and vancomycin. In addition, strain L. delbrueckii UO 004 was not able to translocate towards the intestinal barrier of mice or produce changes in their activity or general health status.

Animals↗

Polypodium leucotomos extract: antioxidant activity and disposition.

The extract of the fern Polypodium leucotomos (PL, Fernblock) is an oral photoprotectant with strong antioxidative properties. Recent studies to determine its chemical composition have shown 4-hydroxycinnamic acid (p-coumaric), 3 methoxy-4-hydroxycinnamic acid (ferulic), 3,4-dihydroxycinnamic acid (caffeic), 3-methoxy-4-hydroxybenzoic acid (vanillic) and 3-caffeoilquinic acid (chlorogenic) to be among its major phenolic components. No conclusive data are available, however, on the H2O2-scavenging capacity of these compounds, or on their absorption and metabolism following their oral intake. In the present work, their antioxidative capacity was assessed by the luminol/H2O2 assay, their absorption studied using Caco-2 cells to resemble the intestinal barrier, and their metabolism investigated using cultured primary rat hepatocytes. The antioxidant capacity of PL components increased in a concentration-dependent manner, with ferulic and caffeic acids the most powerful antioxidants. The apparent permeability results correspond to a human post-oral administration absorption of 70-100% for all tested substances. Coumaric, ferulic and vanillic acids were metabolized by CYP450-dependent mono-oxygenases and partially conjugated to glucuronic acid and sulfate. These phenolic compounds may contribute to the health benefits afforded by this oral photoprotectant.

Animals↗

Detection of specific antibodies against deflagellated Salmonella Enteritidis and S. Enteritidis FliC-specific 9kDa polypeptide.

Specific antibody levels of laying hens and young chickens experimentally infected with Salmonella Enteritidis and vaccinated farm flocks were evaluated by enzyme-linked immunosorbent assays (ELISAs) with two different antigens, deflagellated S. Enteritidis whole cell (DEWC) and S. Enteritidis FliC-specific 9kDa polypeptide (SEP9). Infected laying hens excreted S. Enteritidis throughout the experimental period, and the specific antibody titers in DEWC-ELISA, were significantly higher than the uninfected group. It suggests that this DEWC-specific antibody will serve as an effective indicator of S. Enteritidis infection, especially for non-vaccinated laying flocks. SEP9-specific antibodies were detected in spray-inoculated young chickens but not in oral-inoculated young chickens. Compared with greatly high SEP9-specific antibody levels of vaccinated farm flocks, no response was observed in orally infected hens. These results indicate that S. Enteritidis discontinues expressing SEP9 once it has crossed the intestinal barrier, and that SEP9-ELISA will serve as a valuable monitoring tool for the status of S. Enteritidis vaccination on a flockwide basis, independent of stable S. Enteritidis infections.

Animals↗

On the distribution of genotoxic factors in various organs of mice treated with cycasin.

The distribution of genotoxic factors in various organs of mice treated orally with methylazoxymethanol-beta-D-glycoside (cycasin) was investigated using the DNA-repair host mediated assay. Indicator of genotoxic activity was a pair of streptomycin dependent Escherichia coli strains differing vastly in DNA repair capacity; uvrB/recA vs. uvr+/rec+. The animal-mediated assays were performed by injecting mixtures of the two strains i.v. and orally into mice, which were subsequently treated with the test chemical and from which the differential survival of the indicator bacteria present in several organs was determined. The same strains and selection procedures were also used for assessing the DNA-damaging activity in vitro. In the animal-mediated assays in which cycasin was applied orally, significant effects were observed at doses of 100 and 500 mg/kg body weight. The organ distribution of genotoxic factors in the host animal was as follows: the highest genotoxic activity was observed in the liver, followed by intestine and stomach; a clear effect was also observed in the kidneys and, to a lower extent, in the blood stream and in the lungs at the highest dose administered (500 mg/kg body weight). Under in vitro conditions a marginal genotoxic effect was observed even in the absence of liver homogenate, indicating that the test compound is possible activated (hydrolysed) by the E. coli cells. Therefore the genotoxic activity of cycasin observed in the gastrointestinal tract was not unexpected, since the substance was applied orally, thereby exposing the indicator bacteria in these organs to high levels of unmetabolised compound, especially in the stomach. In the intestine members of the microbial flora probably contribute to the metabolic activation of the test compound. The occurrence of genotoxic factors remote from the gastrointestinal tract shows that the present compound or active metabolites thereof penetrate through the intestinal barrier. The extraordinarily high genotoxic activity observed in the liver suggests that the compound is additionally activated in this organ. In compliance with previous in vitro findings this second activation step might lead to the formation of the highly reactive aldehydic form of methylazoxymethanol (MAMAL) mediated by dehydrogenases. Comparison with carcinogenicity studies indicates a good correlation between the distribution of genotoxic effects as determined in the present studies and the localisation of tumors in various organs of rodents treated with cycasin.

Animals↗

Inhibitors of poly (ADP-ribose) polymerase modulate signal transduction pathways in colitis.

During inflammatory bowel diseases, oxidative and nitrosative stress induces DNA damage and activation of the nuclear enzyme poly (ADP-ribose) polymerase (PARP), resulting in depletion of intracellular energetics, intestinal barrier dysfunction and cellular death. The aim of our study was to evaluate the therapeutic efficacy of in vivo inhibition of PARP in experimental colitis, which was induced by rectal instillation of trinitrobenzene sulfonic acid (TNBS) in rats. In vehicle-treated rats, TNBS treatment resulted in colonic erosion and ulceration. Neutrophil infiltration (indicated by myeloperoxidase activity in the colon) was associated with formation of nitrotyrosine and marked apoptosis. Elevated levels of plasma nitrate/nitrite, metabolites of nitric oxide (NO), were also found. These inflammatory events were associated with the activation of nuclear factor-kappa B (NF-kappa B) and activator protein-1 (AP-1) in the colon; NF-kappa B was maximally activated at 3 and 7 days, whereas AP-1 increased 1 day after TNBS administration and declined thereafter. Treatment of the rats with the PARP inhibitors, 3-aminobenzamide or 1,5-dihydroxyisoquinoline, resolved colonic damage and reduced plasma levels of NO metabolites. Resolution of the damage was associated with reduction of neutrophil infiltration, nitrotyrosine formation and apoptosis. Treatment with PARP inhibitors also reduced DNA binding of NF-kappa B and AP-1 in the colon. These data demonstrate that pharmacological inhibition of PARP ameliorates colitis. Reduction of the inflammatory process is associated with modification of the activation of signal transduction pathways.

Animals↗

The role of dendritic cells during Salmonella infection.

One type of phagocytic antigen-presenting cell (APC) - the dendritic cell (DC) - may have specialized functions during infection with the bacterium Salmonella, including a possible role in transporting Salmonella across the intestinal barrier. In addition, changes in the number, localization and cytokine production of CD8alpha+, CD8alpha-CD4+ and CD8alpha-CD4- DC subsets occur during infection. DCs function in stimulating bacteria-specific T cells by direct presentation of Salmonella antigens and as bystander APCs. Studying the function of DCs during Salmonella infection provides insight into the capacity of these sophisticated APCs, which are a key link between innate and adaptive immunity, to initiate and modulate the immune response to a bacterial infection.

Animals↗

The effect of chitosan and other polycations on tight junction permeability in the human intestinal Caco-2 cell line(1).

Chitosan is a polycationic compound widely employed as dietary supplement and also present in pharmaceutical preparations. Although it has been approved for human consumption, its possible side effects have not been widely investigated and the available data in the literature are still controversial. Several polycationic substances have been shown to affect tight junction permeability in epithelial cell models in vitro. In this study we have compared the effects of chitosan and other polycations (polyethylenimine, poly-L-lysines of different molecular weights) on the integrity of tight junctions and of the actin cytoskeleton in the human intestinal Caco-2 cell line. We have measured trans-epithelial electrical resistance and paracellular passage of the extracellular marker inulin, and we have localized F-actin and tight junctional proteins (ZO1 and occludin) in cell monolayers treated with various concentrations of each polycation. Fluorescent poly-L-lysines were also employed to determine their association with the cell monolayer. Our results indicate that all polycations investigated are able to induce a reversible increase in tight junction permeability. This effect is concentration and energy dependent, affected by the extracellular concentration of divalent cations (calcium, magnesium and manganese) and it is associated with morphological changes in the F-actin cytoskeleton, as well as in the localization of tight junctional proteins. Chitosan, in particular, was the only cationic polymer that displayed an irreversible effect on tight junctions at the highest concentration tested (0.01%). These results indicate that oral ingestion of chitosan may have more widespread health effects by altering intestinal barrier function, thus allowing the entrance into the circulation of potentially toxic and/or allergenic substances.

Journal Article↗

Application of enzymatically stable dipeptides for enhancement of intestinal permeability. Synthesis and in vitro evaluation of dipeptide-coupled compounds.

Transport across the intestinal barrier of compounds with low permeability may be facilitated by targeting the human oligopeptide transporter, hPepT1. A flexible synthetic pathway for attaching compounds to dipeptides through ester or amide bonds was developed. Furthermore, a synthetic approach to functionalize model drugs from one key intermediate was generated and applied to a glucose-6-phosphatase active model drug. The model drug was coupled to D-Glu-Ala through various linkers, and the G-6-Pase activity as well as the aqueous solubility and transport properties of these prodrugs, as compared to those of the parent drugs, were examined. None of the peptide-coupled compounds seemed to be transported by hPepT1, though one of the peptide-coupled compounds had affinity for hPepT1. Interestingly, in one case the parent drug was actively effluxed, while the corresponding peptide-coupled prodrug was not. The low aqueous solubility of the parent compounds was not increased after attachment to a dipeptide. This suggests that only compounds with a certain intrinsic aqueous solubility should be targeted to hPepT1 by attachment to a dipeptide. Important information about the design of peptide-coupled drugs targeted for hPepT1 is presented.

Benzyl Alcohols↗

Markedly high eosinophilia and an elevated serum IL-5 level in an infant with cow milk allergy.

BACKGROUND: Interleukin-5 (IL-5) promotes the production and function of eosinophils, and an increase in the serum soluble CD23 (sCD23) level is suggestive of enhanced type-2 helper T-cell activity. The secretion of a large amount of the proinflammatory cytokine, tumor necrosis factor alpha (TNF-a), has been reported to alter the intestinal barrier capacity. OBJECTIVE: To determine whether or not distinct profiles of cytokine production were involved in the marked peripheral eosinophilia of as high as 20,000/mm3 and the gastrointestinal symptoms seen in an infant with cow milk allergy. METHODS: The levels of IL-5, sCD23, and TNF-alpha in serum and the culture supernatants of mononuclear cells were compared with those in infants with anaphylaxis to cow milk and nonallergic infants. RESULTS: Interleukin-5 was detected in the serum (19 pg/mL) but became undetectable after 2 weeks on a milk-free diet together with clinical remission. A kinetic decrease in the serum sCD23 level was also observed during the administration of a milk-free diet with improvement of the eosinophilia in 2 months. The TNF-alpha produced in vitro after stimulation with cow milk protein was not different from in controls. CONCLUSION: It seems likely that the allergic inflammation due to cow milk can induce marked eosinophilia with an associated increase in IL-5 production.

Animals↗

Anabolic strategies in critical illness.

Critical illness precipitates a marked catabolic response, with protein wasting and loss of lean body mass. Prolongation of this response leads to impaired immunity, poor wound healing, loss of intestinal barrier function and muscle weakness, thereby increasing morbidity and perhaps mortality. Conventional nutritional support only partially ameliorates this process. Disappointingly, specific anabolic and anticatabolic strategies have so far met with only limited success, although recent findings, in particular studies demonstrating the potential value of aggressive insulin therapy and the administration of growth hormone secretagogues, have been encouraging.

Adrenergic Agents↗

Synthesis and in vitro biological evaluation of mannose-containing prodrugs derived from clinically used HIV-protease inhibitors with improved transepithelial transport.

In an approach to improve the pharmacological properties, safety and pharmacokinetic profiles, and their penetration into HIV reservoirs or sanctuaries, and consequently, the therapeutic potential of the current protease inhibitors (PIs) used in clinics, we investigated the synthesis of various mannose-substituted saquinavir, nelfinavir, and indinavir prodrugs, their in vitro stability with respect to hydrolysis, anti-HIV activity, cytotoxicity, and permeation through a monolayer of Caco-2 cells used as a model of the intestinal barrier. Mannose-derived conjugates were prepared in two steps, in good yields, by condensing an acid derivative of a protected mannose with the PIs, followed by deprotection of the sugar protecting group. With respect to hydrolysis, these PI prodrugs are chemically stable with half-life times in the 50-60 h range that are compatible with an in vivo utilization aimed at improving the absorption/penetration or accumulation of the prodrug in specific cells/tissues and liberation of the active free drug inside HIV-infected cells. These stabilities correlate closely with the low in vitro anti-HIV activity measured for those prodrugs wherein the coupling of mannose to the PIs was performed through the peptidomimetic PI's hydroxyl. Importantly, mannose conjugation to the PIs was further found to improve the absorptive transepithelial transport of saquinavir and indinavir but not of nelfinavir across Caco-2 cell monolayers, by contrast to glucose conjugation which had the opposite effect. The mannose-linked prodrugs of saquinavir and indinavir display therefore a most promising therapeutic potential provided that bioavailability, penetration into the HIV infected macrophages, and HIV-reservoirs of these PIs are improved.

Caco-2 Cells↗

Botulinum toxins--cause of botulism and systemic diseases?

Toxins of Clostridium botulinum (types A-G) are known as 'neurotoxins', causing the clinically well-known picture of flaccid muscular paralysis. The molecular biological background is the blocking of acetylcholine secretion in neuromuscular junctions by enzymatic cleavage of molecules forming the machinery of exocytosis. Two 'non-neurotoxins' (types C2, C3) are produced by some strains of C. botulinum types C and D. These affect the cytoskeleton by ribosylating actin filaments. All these toxins are used as cell biological tools for the study of specific actions and effects in different eukaryotic cells. Pharmaceutical and molecular biological research has shown their influence on several crucial organs (or cell cultures thereof) of humans and animals (brain and spinal cord, cerebellum, hippocampus, hypophysis, pancreas, adrenal glands, salivary glands and others). Under natural conditions, botulinum toxins may pass the intestinal barrier and circulate in the bloodstream for a certain time. Carriers occurring naturally in food, such as wheat germ agglutinin, digitonin or saponin, and bacterial toxins such as streptolysin O, perfringolysins, C2 toxin or botulinolysin may also form pores in cell walls. They facilitate the entry of botulinum toxins into cells that may not have natural binding receptors. It is concluded that in vivo actions of different botulinum toxins after their entry into the organism may contribute to the onset of different diseases of hitherto cryptogenic origin. Some examples are given and future problems are discussed.

Animals↗

NO-dependent and NO-independent IL-1 production by a human colonic epithelial cell line under inflammatory stress.

1. The present study was designed to investigate, in an in vitro model of the human intestinal barrier, the ability of epithelial cells to produce interleukin-1 (IL-1), the cellular mechanisms involved in IL-1 release, and the intracellular signalling pathways involved in IL-1 up-regulation during inflammatory stress. 2. This study was based on the human colonic epithelial cell line HT29-Cl.16E, maintained as polarized monolayers on filters mounted in culture chambers and treated with various proinflammatory cytokines (interferon gamma (IFN gamma), tumour necrosis factor alpha (TNF alpha), IL-1 beta) alone or in combination. 3. IL-1 production, restricted to IL-1 alpha, was induced by the combination of IFN gamma/TNF alpha. When IL-1 beta was added to IFN gamma/TNF alpha, it led to an additional production of IL-1 alpha. IL-1 alpha release was associated with cell damage, as shown by the correlation between lactate dehydrogenase (LDH) release and extracellular IL-1 production, and was not accounted for by a secretory mechanism. 4. Both IFN gamma/TNF alpha and IFN gamma/TNF alpha/IL-1 beta induced inducible nitric oxide synthase (iNOS) expression as shown by quantitation of NO2-/NO3- by use of the Griess reagent, quantitation of cells scoring positive with an anti-iNOS antibody and detection of mRNAs coding for iNOS by RT-PCR. The use of NG-monomethyl-L-arginine (L-NMMA), an inhibitor of NOS, led to the demonstration of two distinct signalling pathways in IL-1 production by HT29-Cl.16E cells, one dependent on NO (L-NMMA-sensitive) under treatment with IFN gamma/TNF alpha/IL-1 beta, and the other independent of NO (L-NMMA-insensitive) under treatment with IFN gamma/TNF alpha. 5. Moreover, we examined whether a redox-based mechanism could be responsible for the apparent discrepancy between NO production and NO implication in IL-1 production under IFN gamma/TNF alpha and IFN gamma/TNF alpha/IL-1 beta treatments. Experiments with cysteine, which acts as a powerful reductant, suggest that the nitrosonium character of NO is involved in the NO-dependent pathway in IL-1 production.

Cells, Cultured↗

The role of virulence factors in enterohemorrhagic Escherichia coli (EHEC)--associated hemolytic-uremic syndrome.

Enterohemorrhagic Escherichia coli (EHEC) are the most common cause of postdiarrheal hemolytic-uremic syndrome (HUS). The most important EHEC serotype implicated worldwide is O157:H7. However, several so-called non-O157 EHEC serotypes have emerged. After a mean incubation period of 3 days, patients develop watery diarrhea accompanied by cramping abdominal pain. During the next days, in most patients watery diarrhea changes to bloody diarrhea. One week after the onset of diarrhea, in about 15% of infected patients under 10 years of age EHEC infection results in a systemic complication, HUS. Shiga toxins (Stxs) are considered the major virulence factors of EHEC involved in the pathogenesis of HUS. It is generally believed that after intestinal infection with EHEC, Stxs cross the intestinal barrier and bind to endothelial cells. At this point they presumably injure the host cell by inhibition of protein synthesis, stimulation of prothrombotic messages, or induction of apoptosis. The B subunit of Stx binds to the membrane receptor globotriaosylceramide (Gb3). Gb3 facilitates the endocytosis and intracellular trafficking of the toxin. The Stx A subunit hydrolyzes a specific adenine residue of the 60S ribosomal subunit of mammalian cells. As a consequence, Stx shuts down the protein machinery of the susceptible cell. The HUS is the net effect of a variety of interacting factors, including background risk of acquisition, host factors (such as age), virulence characteristics of the infecting EHEC strain, and exogenous factors. All known EHEC virulence determinants are located on mobile genetic elements, and this has an important impact on the evolution of these pathogens. The evolution of EHEC has a dynamic component that includes different genetic mechanisms. The recent progress in understanding the pathogenesis and epidemiology of EHEC infections forms a basis for the development of future strategies to prevent EHEC infections in humans.

Endothelium, Vascular↗

Skin bioavailability of dietary vitamin E, carotenoids, polyphenols, vitamin C, zinc and selenium.

Dietary bioactive compounds (vitamin E, carotenoids, polyphenols, vitamin C, Se and Zn) have beneficial effects on skin health. The classical route of administration of active compounds is by topical application direct to the skin, and manufacturers have substantial experience of formulating ingredients in this field. However, the use of functional foods and oral supplements for improving skin condition is increasing. For oral consumption, some dietary components could have an indirect effect on the skin via, for example, secondary messengers. However, in the case of the dietary bioactive compounds considered here, we assume that they must pass down the gastrointestinal tract, cross the intestinal barrier, reach the blood circulation, and then be distributed to the different tissues of the body including the skin. The advantages of this route of administration are that the dietary bioactive compounds are metabolized and then presented to the entire tissue, potentially in an active form. Also, the blood continuously replenishes the skin with these bioactive compounds, which can then be distributed to all skin compartments (i.e. epidermis, dermis, subcutaneous fat and also to sebum). Where known, the distribution and mechanisms of transport of dietary bioactive compounds in skin are presented. Even for compounds that have been studied well in other organs, information on skin is relatively sparse. Gaps in knowledge are identified and suggestions made for future research.

Administration, Oral↗

Mixed polymer micelles of amphiphilic and cationic copolymers for delivery of antisense oligonucleotides.

Cationic copolymers were synthesized by conjugation of branched 2 kDa polyethylenimine (PEI) and Pluronic block copolymers (F38, P85, P123). Compositions of these copolymers mixed with corresponding free Pluronics at weight ratio 1:9 were used to complex phosphorothioate oligonucleotides (ODN). As a result stable suspensions of small micelle-like particles (<220 nm) were obtained. Incorporation of ODN in these formulations increased uptake of ODN in KBv cells and increased sequence specific activity of antisense ODN targeted against MDR gene in multidrug resistant cells resulting in inhibition of the functional activity of P-glycoprotein (P-gp) in these cells. Furthermore, these formulations increased transport of ODN across model intestinal barrier, Caco-2 cell monolayers, suggesting that they could be useful for oral delivery of biologically active ODN.

Caco-2 Cells↗

Natural products alleviate exercise-induced fatigue by modulating gut microbiota: a systematic review.

BACKGROUND: Exercise-induced fatigue critically impairs athletic performance and training quality. The gut microbiota, as a key regulator of the "gut-muscle axis," has emerged as a promising anti-fatigue target. Natural products - owing to their diverse sources, structural complexity, and favorable safety profiles - have attracted growing research interest. However, a systematic synthesis comparing their anti-fatigue effects via gut microbiota modulation across different sources is lacking. SCOPE AND APPROACH: We systematically searched PubMed, Web of Science, the Cochrane Library, and CNKI for original studies that administered natural products and concurrently assessed gut microbiota changes and anti-fatigue outcomes. Twenty-six studies (25 animal experiments and 1 human trial) were included and categorized into seven groups by source and chemical characteristics. A descriptive systematic review was conducted to identify common mechanisms and source-specific differentiations. KEY FINDINGS AND CONCLUSIONS: The enrichment of short-chain fatty acid (SCFA)-producing bacteria and the activation of the SCFA-AMPK/PGC-1&#x3b1; axis were shared core events across all product categories. However, source-dependent mechanistic divergences emerged: polysaccharides acted primarily as fermentable substrates with an optimal dose window; polyphenols and saponins exerted dual modulation on both microbiota and host signaling pathways; compound extracts achieved systemic synergy through functional complementation; marine- and animal-derived products exhibited unique targeting profiles and rapid action. Intestinal barrier maintenance and brain-gut axis regulation further extended the anti-fatigue repertoire. Collectively, natural products possess a solid mechanistic basis for alleviating exercise-induced fatigue via gut microbiota remodeling. The differentiated characteristics of these methods in targeting precision and pathway engagement provide a theoretical foundation for designing precision intervention strategies tailored to specific fatigue contexts.

Humans↗

A molecular marker for evaluating the pathogenic potential of foodborne Listeria monocytogenes.

BACKGROUND: Internalin mediates entry of Listeria monocytogenes into some human cultured cell lines and crossing of the intestinal barrier in transgenic mice expressing its receptor, human E-cadherin, in enterocytes. The relevance of these findings for humans is challenged by the observation that some L. monocytogenes isolates express a truncated nonfunctional form of internalin. METHODS: We investigated expression of internalin by use of immunoblot assay in 300 clinical strains obtained in France in a single year and a representative set of 150 strains obtained from food products during the same period. RESULTS: Clinical strains expressed full-length internalin far more frequently (288/300 strains [96%]) than did strains recovered from food products (98/150 strains [65%]; odds ratio, 12.73; 95% confidence interval, 6.27-26.34; P<1 x 10(-7)). All 61 strains (100%) from pregnancy-related cases, 55 (98%) of 56 strains from patients with central nervous system infections, and 151 (93%) of 162 strains from patients with bacteremia expressed full-length internalin. All 110 strains belonging to serovar 4b, the most frequently implicated serovar in human listeriosis, expressed full-length internalin. CONCLUSIONS: This study demonstrates the critical role of internalin in the pathogenesis of human listeriosis. It provides a molecular explanation for the predominance of serovar 4b among clinical strains and supports the usefulness of studying the expression of internalin as a marker of virulence in humans.

Bacteremia↗