Veal Calves Produce Less Antibodies against C. Perfringens Alpha Toxin Compared to Beef Calves.
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ABSTRACT:
SUBMITTER: Valgaeren BR
PROVIDER: S-EPMC4516930 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:BackgroundLinezolid is a critically important oxazolidinone antibiotic used in human medicine. Although linezolid is not licensed for use in food-producing animals, the use of florfenicol in veterinary medicine co-selects for oxazolidinone resistance genes.ObjectiveThis study aimed to assess the occurrence of cfr, optrA, and poxtA in florfenicol-resistant isolates from beef cattle and veal calves from different herds in Switzerland.MethodsA total of 618 cecal samples taken from beef cattle and veal calves at slaughter originating from 199 herds were cultured after an enrichment step on a selective medium containing 10 mg/L florfenicol. Isolates were screened by PCR for cfr, optrA, and poxtA which are genes known to confer resistance to oxazolidinones and phenicols. One isolate per PCR-positive species and herd was selected for antimicrobial susceptibility testing (AST) and whole-genome sequencing (WGS).ResultsOverall, 105 florfenicol-resistant isolates were obtained from 99 (16%) of the samples, corresponding to 4% of the beef cattle herds and 24% of the veal calf herds. Screening by PCR revealed the presence of optrA in 95 (90%) and poxtA in 22 (21%) of the isolates. None of the isolates contained cfr. Isolates included for AST and WGS analysis were Enterococcus (E.) faecalis (n = 14), E. faecium (n = 12), E. dispar (n = 1), E. durans (n = 2), E. gallinarum (n = 1), Vagococcus (V.) lutrae (n = 2), Aerococcus (A.) urinaeequi (n = 1), and Companilactobacillus (C.) farciminis (n = 1). Thirteen isolates exhibited phenotypic linezolid resistance. Three novel OptrA variants were identified. Multilocus sequence typing identified four E. faecium ST18 belonging to hospital-associated clade A1. There was a difference in the replicon profile among optrA- and poxtA-harboring plasmids, with rep9 (RepA_N) plasmids dominating in optrA-harboring E. faecalis and rep2 (Inc18) and rep29 (Rep_3) plasmids in poxtA-carrying E. faecium.ConclusionBeef cattle and veal calves are reservoirs for enterococci with acquired linezolid resistance genes optrA and poxtA. The presence of E. faecium ST18 highlights the zoonotic potential of some bovine isolates. The dispersal of clinically relevant oxazolidinone resistance genes throughout a wide variety of species including Enterococcus spp., V. lutrae, A. urinaeequi, and the probiotic C. farciminis in food-producing animals is a public health concern.
| S-EPMC10301837 | biostudies-literature
Project description:Clostridium perfringens alpha-toxin is a key mediator of gas gangrene, which is a life-threatening infection that manifests as fever, pain, edema, myonecrosis, and gas production. Alpha-toxin possesses phospholipase C and sphingomyelinase activities. The toxin is composed of an N-terminal domain (1-250 aa, N-domain), which is the catalytic site, and a C-terminal domain (251-370 aa, C-domain), which is the membrane-binding site. Immunization of mice with the C-domain of alpha-toxin prevents the gas gangrene caused by C. perfringens, whereas immunization with the N-domain has no effect. The central loop domain (55-93 aa), especially H….SW(84)Y(85)….G, plays an important role in the interaction with ganglioside GM1a. The toxin binds to lipid rafts in the presence of a GM1a/TrkA complex, and metabolites from phosphatidylcholine to diacylglycerol through the enzymatic activity of alpha-toxin itself. These membrane dynamics leads to the activation of endogenous PLC?-1 via TrkA. In addition, treatment with alpha-toxin leads to the formation of diacylglycerol at membrane rafts in ganglioside-deficient DonQ cells; this in turn triggers endocytosis and cell death. This article summarizes the current the membrane-binding mechanism of alpha-toxin in detail.
| S-EPMC4690130 | biostudies-literature
Project description:Clostridium perfringens alpha-toxin is the major virulence factor in the pathogenesis of gas gangrene. Alpha-toxin is a 43-kDa protein with two structural domains; the N-domain contains the catalytic site and coordinates the divalent metal ions, and the C-domain is a membrane-binding site. The role of the exposed loop region (72-93 residues) in the N-domain, however, has been unclear. Here we show that this loop contains a ganglioside binding motif (H … SXWY … G) that is the same motif seen in botulinum neurotoxin and directly binds to a specific conformation of the ganglioside Neu5Acα2-3(Galβ1-3GalNAcβ1-4)Galβ1-4Glcβ1Cer (GM1a) through a carbohydrate moiety. Confocal microscopy analysis using fluorescently labeled BODIPY-GM1a revealed that the toxin colocalized with GM1a and induced clustering of GM1a on the cell membranes. Alpha-toxin was only slightly toxic in β1,4-N-acetylgalactosaminyltransferase knock-out mice, which lack the a-series gangliosides that contain GM1a, but was highly toxic in α2,8-sialyltransferase knock-out mice, which lack both b-series and c-series gangliosides, similar to the control mice. Moreover, experiments with site-directed mutants indicated that Trp-84 and Tyr-85 in the exposed alpha-toxin loop play an important role in the interaction with GM1a and subsequent activation of TrkA. These results suggest that binding of alpha-toxin to GM1a facilitates the activation of the TrkA receptor and induces a signal transduction cascade that promotes the release of chemokines. Therefore, we conclude that GM1a is the primary cellular receptor for alpha-toxin, which can be a potential target for drug developed against this pathogen.
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Project description:Clostridium perfringens mutant strain 121A/91 shows neither enzymatic (phospholipase C) nor hemolytic activity. Nevertheless, the cpa gene and the corresponding alpha-toxin variant are detectable. Vaccination with this genetically constructed alpha-toxin variant, rAT121/91, induces antibodies capable of significantly reducing activities induced by wild-type toxin. Thus, rAT121/91 could be a useful vaccine candidate.
| S-EPMC100128 | biostudies-literature
Project description:All subtypes of Clostridium perfringens (C. perfringens) produce the alpha toxin (CPA), which can cause enteritis or enterotoxemia in lambs, cattle, pigs, and horses, as well as traumatic clostridial myonecrosis in humans and animals. CPA acts on cell membranes, ultimately leading to endocytosis and cell death. Therefore, the neutralization of CPA is crucial for the prevention and treatment of diseases caused by C. perfringens. In this study, utilizing CPA as an antigen, a nanobody (CPA-VHH) with a half-life of 2.9 h, an affinity constant (KD) of 0.9 nmol/L, and good stability below 60 °C was prepared from a natural nanobody library from alpacas. The biological activity analysis of CPA-VHH revealed its ability to effectively neutralize the phospholipase and hemolytic activity of CPA at a 15-fold ratio. In Vero cells, 9.8 μg/mL CPA-VHH neutralized the cytotoxicity of CPA at two times the half-maximal inhibitory concentration (IC50). In a mouse model, 35.7 ng/g body weight (BW) of CPA-VHH neutralized 90% of the lethality caused by a 2× median lethal dose (LD50) of CPA. It was found that CPA-VHH protected 80% of mice within 30 min at 2 × LD50 CPA, but this dropped below 50% after 2 h and to 0% after 4 h. Rescue trials indicated that using CPA-VHH within 30 min post-infection with 2 × LD50 CPA achieved an 80% rescue rate, which decreased to 10% after 2 h. Furthermore, CPA-VHH effectively mitigated the reduction in the expression levels of zonula occludens-1 (ZO-1), Occludin, and Claudin-1, while also attenuating the upregulation of the pro-inflammatory cytokines interleukin-1β (IL-1β), interleukin-6 (IL-6), interleukin-7 (IL-7), interleukin-8 (IL-8), tumor necrosis factor α (TNF-α), and interferon-γ (IFN-γ) induced by CPA infection. Overall, this study has identified a specific nanobody, CPA-VHH, that effectively neutralizes CPA toxins in vitro and in animal models, providing a new tool for inhibiting the pathogenicity resulting from these toxins and laying an important foundation for the development of new anti-C. perfringens toxin-related therapeutic products.
| S-EPMC11360521 | biostudies-literature
Project description:Although granulopoiesis is accelerated to suppress bacteria during infection, some bacteria can still cause life-threatening infections, but the mechanism behind this remains unclear. In this study, we found that mature neutrophils in bone marrow cells (BMCs) were decreased in C. perfringens-infected mice and also after injection of virulence factor α-toxin. C. perfringens infection interfered with the replenishment of mature neutrophils in the peripheral circulation and the accumulation of neutrophils at C. perfringens-infected sites in an α-toxin-dependent manner. Measurements of bacterial colony-forming units in C. perfringens-infected muscle revealed that α-toxin inhibited a reduction in the load of C. perfringens. In vitro treatment of isolated BMCs with α-toxin (phospholipase C) revealed that α-toxin directly decreased mature neutrophils. α-Toxin did not influence the viability of isolated mature neutrophils, while simultaneous treatment of BMCs with granulocyte colony-stimulating factor attenuated the reduction of mature neutrophils by α-toxin. Together, our results illustrate that impairment of the innate immune system by the inhibition of neutrophil differentiation is crucial for the pathogenesis of C. perfringens to promote disease to a life-threatening infection, which provides new insight to understand how pathogenic bacteria evade the host immune system.
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Project description:Heavy veal calves (4-6 months old) are at risk of developing insulin resistance and disturbed glucose homeostasis. Prolonged insulin resistance could lead to metabolic disorders and impaired growth performance. Recently, we discovered that heavy Holstein-Friesian calves raised on a high-lactose or high-fat diet did not differ in insulin sensitivity, that insulin sensitivity was low and 50% of the calves could be considered insulin resistant. Understanding the patho-physiological mechanisms underlying insulin resistance and discovering biomarkers for early diagnosis would be useful for developing prevention strategies. Therefore, we explored plasma metabolic profiling techniques to build models and discover potential biomarkers and pathways that can distinguish between insulin resistant and moderately insulin sensitive veal calves. The calves (n = 14) were classified as insulin resistant (IR) or moderately insulin sensitive (MIS) based on results from a euglycemic-hyperinsulinemic clamp, using a cut-off value (M/I-value <4.4) to identify insulin resistance. Metabolic profiles of fasting plasma samples were analyzed using reversed phase (RP) and hydrophilic interaction (HILIC) liquid chromatography-mass spectrometry (LC-MS). Orthogonal partial least square discriminant analysis was performed to compare metabolic profiles. Insulin sensitivity was on average 2.3x higher (P <0.001) in MIS than IR group. For both RP-LC-MS and HILIC-LC-MS satisfactory models were build (R2Y >90% and Q2Y >66%), which allowed discrimination between MIS and IR calves. A total of 7 and 20 metabolic features (for RP-LC-MS and HILIC-LC-MS respectively) were most responsible for group separation. Of these, 7 metabolites could putatively be identified that differed (P <0.05) between groups (potential biomarkers). Pathway analysis indicated disturbances in glycerophospholipid and sphingolipid metabolism, the glycine, serine and threonine metabolism, and primary bile acid biosynthesis. These results demonstrate that plasma metabolic profiling can be used to identify insulin resistance in veal calves and can lead to underlying mechanisms.
| S-EPMC5472311 | biostudies-literature
Project description:Three vancomycin-resistant veal calf fecal streptococci, identified as Streptococcus gallolyticus (n = 2) and Streptococcus lutetiensis, were shown to harbor vanB2 Tn5382-like elements earlier described in enterococci. One S. gallolyticus strain had a 1,495-bp IS256-related element inserted in vanS(B). The vanB2 Tn5382 element present in the plasmid-free S. lutetiensis strain was transferable to Enterococcus faecium BM4105-RF, Enterococcus faecalis JH2-2, and its recombination-deficient derivative, UV202. The transfer frequencies were comparable between recipient strains (from 1 x 10(-7) to 7 x 10(-6)). All transconjugants acquired a vanB-containing chromosomal insert of approximately 100 kb, apparently by site-specific integration. Secondary transconjugants were not observed in intraspecies retransfer experiments. These observations are consistent with a conjugative, selftransmissible, integrative element that might be involved in the interspecies spread of vanB2 resistance determinants. Two JH2-2-derived transconjugants had also gained additional copies of large vanB-containing chromosomal fragments, a process that involves unexplained mechanisms that seems to require functional host cell-dependent recombination mechanisms.
| S-EPMC166075 | biostudies-literature
Project description:Selenium (Se) is an essential trace mineral important for immune function and overall health of cattle. The nasopharyngeal microbiota in cattle plays an important role in overall respiratory health, especially when stresses associated with weaning, transport, and adaptation to a feedlot affect the normal respiratory defenses. Recent evidence suggests that cattle diagnosed with bovine respiratory disease complex have significantly less bacterial diversity. The objective of this study was to determine whether feeding weaned beef calves Se-enriched alfalfa (Medicago sativa) hay for 9 weeks in a preconditioning program prior to entering the feedlot alters nasal microbiota. Recently weaned beef calves (n = 45) were blocked by sex and body weight, randomly assigned to 3 treatment groups with 3 pens of 5 calves per treatment group, and fed an alfalfa hay based diet for 9 weeks. Alfalfa hay was harvested from fields fertilized with sodium selenate at a rate of 0, 45.0 or 89.9 g Se/ha. Blood samples were collected biweekly and analyzed for whole-blood Se concentrations. Nasal swabs were collected during week 9 from one or two calves from each pen (total n = 16). Calculated Se intake from dietary sources was 3.0, 15.6, and 32.2 mg Se/head/day for calves consuming alfalfa hay with Se concentrations of 0.34 to 2.42 and 5.17 mg Se/kg dry matter, respectively. Whole-blood Se concentrations after 8 weeks of feeding Se-fertilized alfalfa hay were dependent upon Se-application rates (0, 45.0, or 89.9 g Se/ha) and were 155, 345, and 504 ng/mL (PLinear < 0.0001). Microbial DNA was extracted from nasal swabs and amplified and sequenced. Alpha rarefaction curves comparing the species richness (observed OTUs) and overall diversity (Chao1, Observed OTU, and Shannon index) between calves fed selenium-biofortified alfalfa hay compared with control calves showed that Se-supplementation tended to be associated with an enriched nasal microbiota. ANOSIM of unweighted UniFrac distances showed that calves fed high Se-biofortified alfalfa hay clustered separately when compared with control calves in the PCoA plot (R = 0.216, P = 0.04). The bacterial orders Lactobacillales and Flavobacteriales were increased in healthy control calves compared with Clostridiales and Bacteroidales being increased in calves fed Se-biofortified alfalfa hay. Although there were strong trends, no significant differences were noted for any of the bacterial taxa. Based upon these findings, we suggest that weaned beef calves fed Se-biofortified hay tend to have an enriched nasal microbiota. Feeding Se-biofortified alfalfa hay to weaned beef calves prior to entering the feedlot is a strategy for increasing nasopharyngeal microbial diversity.
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Project description:Clostridium perfringens (C. perfringens) type A strains are the main cause of gas gangrene in humans and animals. Treatment of this lethal disease is limited, and the prognosis is not good. Alpha-toxin (CPA) and perfringolysin O (PFO) secreted by C. perfringens play irreplaceable roles in cytotoxicity to host cells, persistence in host tissues, and lethality of gas gangrene pathology. This work determined the influence of amentoflavone, a biflavonoid isolated from Selaginella tamariscina and other plants, on hemolysis and cytotoxicity mediated by CPA and PFO and evaluated the in vivo therapeutic effect on gas gangrene. Our data showed that amentoflavone could block the hemolysis and cytotoxicity induced by CPA and PFO in vitro, thereby mediating significant protection against mortality of infected mice in a mouse gas gangrene model, efficient bacterial clearance in tissues and alleviation of histological damage in vivo. Based on the above results, amentoflavone may be a potential candidate against C. perfringens infection by reducing CPA and PFO-mediated virulence.
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