Underlying mechanisms involved in the decrease of milk secretion during Escherichia coli endotoxin induced mastitis in lactating mice.
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ABSTRACT:
SUBMITTER: Kobayashi K
PROVIDER: S-EPMC4028753 | biostudies-literature | 2013
REPOSITORIES: biostudies-literature
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Project description:Changes in the milk microbiota during the course of mastitis are due to the nature of a sporadic occurring disease difficult to study. In this study we experimentally induced mastitis by infusion of Escherichia coli endotoxins in one udder quarter each of nine healthy lactating dairy cows and assessed the bacteriological dynamics and the milk microbiota at four time points before and eight time points after infusion. As control, saline was infused in one udder quarter each of additionally nine healthy cows that followed the same sampling protocol. The milk microbiota was assessed by sequencing of the 16 S rRNA gene and a range of positive and negative controls were included for methodological evaluation. Two different data filtration models were used to identify and cure data from contaminating taxa. Endotoxin infused quarters responded with transient clinical signs of inflammation and increased SCC while no response was observed in the control cows. In the milk microbiota data no response to inflammation was identified. The data analysis of the milk microbiota was largely hampered by laboratory and reagent contamination. Application of the filtration models caused a marked reduction in data but did not reveal any associations with the inflammatory reaction. Our results indicate that the microbiota in milk from healthy cows is unaffected by inflammation.
| S-EPMC10251687 | biostudies-literature
Project description:Escherichia coli, one of the main causative agents of bovine mastitis, is responsible for significant losses on dairy farms. In order to better understand the pathogenicity of E. coli mastitis, an accurate characterization of E. coli strains isolated from mastitis cases is required. By using phylogenetic analyses and whole genome comparison of 5 currently available mastitis E. coli genome sequences, we searched for genotypic traits specific for mastitis isolates. Our data confirm that there is a bias in the distribution of mastitis isolates in the different phylogenetic groups of the E. coli species, with the majority of strains belonging to phylogenetic groups A and B1. An interesting feature is that clustering of strains based on their accessory genome is very similar to that obtained using the core genome. This finding illustrates the fact that phenotypic properties of strains from different phylogroups are likely to be different. As a consequence, it is possible that different strategies could be used by mastitis isolates of different phylogroups to trigger mastitis. Our results indicate that mastitis E. coli isolates analyzed in this study carry very few of the virulence genes described in other pathogenic E. coli strains. A more detailed analysis of the presence/absence of genes involved in LPS synthesis, iron acquisition and type 6 secretion systems did not uncover specific properties of mastitis isolates. Altogether, these results indicate that mastitis E. coli isolates are rather characterized by a lack of bona fide currently described virulence genes.
| S-EPMC4725725 | biostudies-literature
Project description:This work investigates the pharmacodynamic effectiveness of cefquinome against environmental Escherichia coli mastitis infection, following an intramammary administration. We established the pharmacokinetic and pharmacodynamic (PK/PD) model in lactating mice. The PK/PD parameters were identified to achieve an antibacterial efficacy as indicated by PD activity, cytokine expression and PK/PD simulation. From our findings, given an 200 μg/gland dose once daily can achieve a considerable therapeutic effectiveness in experimental circumstance.
| S-EPMC5539083 | biostudies-other
Project description:BackgroundEscherichia coli bovine mastitis is a disease of significant economic importance in the dairy industry. Molecular characterization of mastitis-associated E. coli (MAEC) did not result in the identification of common traits. Nevertheless, a mammary pathogenic E. coli (MPEC) pathotype has been proposed suggesting virulence traits that differentiate MAEC from commensal E. coli. The present study was designed to investigate the MPEC pathotype hypothesis by comparing the genomes of MAEC and commensal bovine E. coli.ResultsWe sequenced the genomes of eight E. coli isolated from bovine mastitis cases and six fecal commensal isolates from udder-healthy cows. We analyzed the phylogenetic history of bovine E. coli genomes by supplementing this strain panel with eleven bovine-associated E. coli from public databases. The majority of the isolates originate from phylogroups A and B1, but neither MAEC nor commensal strains could be unambiguously distinguished by phylogenetic lineage. The gene content of both MAEC and commensal strains is highly diverse and dominated by their phylogenetic background. Although individual strains carry some typical E. coli virulence-associated genes, no traits important for pathogenicity could be specifically attributed to MAEC. Instead, both commensal strains and MAEC have very few gene families enriched in either pathotype. Only the aerobactin siderophore gene cluster was enriched in commensal E. coli within our strain panel.ConclusionsThis is the first characterization of a phylogenetically diverse strain panel including several MAEC and commensal isolates. With our comparative genomics approach we could not confirm previous studies that argue for a positive selection of specific traits enabling MAEC to elicit bovine mastitis. Instead, MAEC are facultative and opportunistic pathogens recruited from the highly diverse bovine gastrointestinal microbiota. Virulence-associated genes implicated in mastitis are a by-product of commensalism with the primary function to enhance fitness in the bovine gastrointestinal tract. Therefore, we put the definition of the MPEC pathotype into question and suggest to designate corresponding isolates as MAEC.
| S-EPMC5422975 | biostudies-literature
Project description:BackgroundLipopolysaccharide (LPS), also referred to as endotoxin, is the major constituent of the outer leaflet of the outer membrane of virtually all Gram-negative bacteria. The lipid A moiety, which anchors the LPS molecule to the outer membrane, acts as a potent agonist for Toll-like receptor 4/myeloid differentiation factor 2-mediated pro-inflammatory activity in mammals and, thus, represents the endotoxic principle of LPS. Recombinant proteins, commonly manufactured in Escherichia coli, are generally contaminated with endotoxin. Removal of bacterial endotoxin from recombinant therapeutic proteins is a challenging and expensive process that has been necessary to ensure the safety of the final product.ResultsAs an alternative strategy for common endotoxin removal methods, we have developed a series of E. coli strains that are able to grow and express recombinant proteins with the endotoxin precursor lipid IVA as the only LPS-related molecule in their outer membranes. Lipid IVA does not trigger an endotoxic response in humans typical of bacterial LPS chemotypes. Hence the engineered cells themselves, and the purified proteins expressed within these cells display extremely low endotoxin levels.ConclusionsThis paper describes the preparation and characterization of endotoxin-free E. coli strains, and demonstrates the direct production of recombinant proteins with negligible endotoxin contamination.
| S-EPMC4404585 | biostudies-literature
Project description:BackgroundMastitis is one of the most common infectious diseases in dairy cattle and causes significant financial losses in the dairy industry worldwide. Antibiotic therapy has been used as the most effective strategy for clinical mastitis treatment. However, due to the extensive use of antibacterial agents, antimicrobial resistance (AMR) is considered to be one of the reasons for low cure rates in bovine mastitis. In addition, biofilms could protect bacteria by restricting antibiotic access and shielding the bacterial pathogen from mammary gland immune defences. The functional mechanisms of quorum sensing E. coli regulators B an d C (QseBC) have been well studied in E. coli model strains; however, whether QseBC regulates antibiotic susceptibility and biofilm formation in clinical E. coli strain has not been reported.MethodsIn this study, we performed construction of the qseBC gene mutant, complementation of the qseBC mutant, antimicrobial susceptibility testing, antibacterial activity assays, biofilm formation assays, real-time reverse transcription PCR (RT-PCR) experiments and electrophoretic mobility shift assays (EMSAs) to investigate the role of qseBC in regulating biofilm formation and antibiotic susceptibility in the clinical E. coli strain ECDCM2.ResultsWe reported that inactivation of QseBC led to a decrease in biofilm formation capacity and an increase in antibiotic susceptibility of an E. coli strain isolated from a dairy cow that suffered from mastitis. In addition, this study indicated that QseBC increased biofilm formation by upregulating the transcription of the biofilm-associated genes bcsA, csgA, fliC, motA, wcaF and fimA and decreased antibiotic susceptibility by upregulating the transcription of the efflux-pump-associated genes marA, acrA, acrB, acrD, emrD and mdtH. We also performed EMSA assays, and the results showed that QseB can directly bind to the marA promoter.ConclusionsThe QseBC two-component system affects antibiotic sensitivity by regulating the transcription of efflux-pump-associated genes. Further, biofilm-formation-associated genes were also regulated by QseBC TCS in E. coli ECDCM2. Hence, this study might provide new clues to the prevention and treatment of infections caused by the clinical E. coli strains.
| S-EPMC7102498 | biostudies-literature
Project description:Bovine mastitis is a widespread disease in dairy cows, and is often caused by bacterial mammary gland infection. Mastitis causes reduced milk production and leads to excessive use of antibiotics. We present meta-analysis of transcriptional profiles of bovine mastitis from 10 studies and 307 microarrays, allowing identification of much larger sets of affected genes than any individual study. Combining multiple studies provides insight into the molecular effects of Escherichia coli infection in vivo and uncovers differences between the consequences of E. coli vs. Staphylococcus aureus infection of primary mammary epithelial cells (PMECs). In udders, live E. coli elicits inflammatory and immune defenses through numerous cytokines and chemokines. Importantly, E. coli infection causes downregulation of genes encoding lipid biosynthesis enzymes that are involved in milk production. Additionally, host metabolism is generally suppressed. Finally, defensins and bacteria-recognition genes are upregulated, while the expression of the extracellular matrix protein transcripts is silenced. In PMECs, heat-inactivated E. coli elicits expression of ribosomal, cytoskeletal and angiogenic signaling genes, and causes suppression of the cell cycle and energy production genes. We hypothesize that heat-inactivated E. coli may have prophylactic effects against mastitis. Heat-inactivated S. aureus promotes stronger inflammatory and immune defenses than E. coli. Lipopolysaccharide by itself induces MHC antigen presentation components, an effect not seen in response to E. coli bacteria. These results provide the basis for strategies to prevent and treat mastitis and may lead to the reduction in the use of antibiotics.
| S-EPMC4775050 | biostudies-literature
Project description:Mastitis remains a major infection of dairy cows and an important issue for the dairy farmers, and Escherichia coli (E. coli) bovine mastitis is a disease of significant economic importance in the dairy industry. Our study identified six isolates belong to phylogroup B2 from 69 bovine mastitis E. coli strains. Except for one serotype O1 strain, all group B2 isolates were identified into serotype O2 and showed significantly higher mortality in the mouse infection than other phylogroups' strains. Genomic analyses and further tests were performed to examine the role of secretion systems, fimbriae, and toxins during the systemic infection of O2:K1 strain BCE049. Two integral T6SS loci and three predicted effectors clusters were found to assemble the functional T6SS complex and deliver diverse toxic effectors to modulate bacterial virulence in the mouse infection model. A total of four T4SS loci were harbored in the BCE049 genome, three of them are encoded in different plasmids, respectively, whereas the last one locates within the bacterial chromosome at FQU84_16715 to FQU84_16760, and was significantly involved in the bacterial pathogenicity. Numerous predicted pilus biosynthesis gene loci were found in the BCE049 genome, whereas most of them lost long fragments encoding key genes for the pili assembly. Unexpectedly, a type IV pilus gene locus locating at FQU84_01405 to FQU84_01335 in the plasmid 2, was found to be required for the full virulence of mastitis strain BCE049. It should be noted that a genetic neighborhood inserted with diverse genes is encoded by the plasmid 1, which harbors three prominent toxins including β-hemolysin, cytotoxic necrotizing factor 2 and cytolethal distending toxin type III. Consequent studies verified that these toxins significantly contributed to the bacterial pathogenicity. These findings provide a molecular blueprint for understanding the underlying mechanisms employed by the bovine mastitis E. coli to colonize in host and cause systemic infection.
| S-EPMC7904677 | biostudies-literature
Project description:BackgroundBovine mastitis is one of the most costly and prevalent diseases affecting dairy cows worldwide. In order to develop new strategies to prevent Escherichia coli-induced mastitis, a detailed understanding of the molecular mechanisms underlying the host immune response to an E. coli infection is necessary. To this end, we performed a global gene-expression analysis of mammary gland tissue collected from dairy cows that had been exposed to a controlled E. coli infection. Biopsy samples of healthy and infected utter tissue were collected at T = 24 h post-infection (p.i.) and at T = 192 h p.i. to represent the acute phase response (APR) and chronic stage, respectively. Differentially expressed (DE) genes for each stage were analyzed and the DE genes detected at T = 24 h were also compared to data collected from two previous E. coli mastitis studies that were carried out on post mortem tissue.ResultsNine-hundred-eighty-two transcripts were found to be differentially expressed in infected tissue at T = 24 (P < 0.05). Up-regulated transcripts (699) were largely associated with immune response functions, while the down-regulated transcripts (229) were principally involved in fat metabolism. At T = 192 h, all of the up-regulated transcripts were associated with tissue healing processes. Comparison of T = 24 h DE genes detected in the three E. coli mastitis studies revealed 248 were common and mainly involved immune response functions. KEGG pathway analysis indicated that these genes were involved in 12 pathways related to the pro-inflammatory response and APR, but also identified significant representation of two unexpected pathways: natural killer cell-mediated cytotoxicity pathway (KEGG04650) and the Rig-I-like receptor signalling pathway (KEGG04622).ConclusionsIn E. coli-induced mastitis, infected mammary gland tissue was found to significantly up-regulate expression of genes related to the immune response and down-regulate genes related to fat metabolism. Up to 25% of the DE immune response genes common to the three E. coli mastitis studies at T = 24 h were independent of E. coli strain and dose, cow lactation stage and number, tissue collection method and gene analysis method used. Hence, these DE genes likely represent important mediators of the local APR against E. coli in the mammary gland.
| S-EPMC3053262 | biostudies-literature
Project description:Virulence factors of mammary pathogenic Escherichia coli (MPEC) have not been identified, and it is not known how bacterial gene content influences the severity of mastitis. Here, we report a genome-wide identification of genes that contribute to fitness of MPEC under conditions relevant to the natural history of the disease. A highly virulent clinical isolate (M12) was identified that killed Galleria mellonella at low infectious doses and that replicated to high numbers in mouse mammary glands and spread to spleens. Genome sequencing was combined with transposon insertion site sequencing to identify MPEC genes that contribute to growth in unpasteurized whole milk, as well as during G. mellonella and mouse mastitis infections. These analyses show that strain M12 possesses a unique genomic island encoding a group III polysaccharide capsule that greatly enhances virulence in G. mellonella Several genes appear critical for MPEC survival in both G. mellonella and in mice, including those for nutrient-scavenging systems and resistance to cellular stress. Insertions in the ferric dicitrate receptor gene fecA caused significant fitness defects under all conditions (in milk, G. mellonella, and mice). This gene was highly expressed during growth in milk. Targeted deletion of fecA from strain M12 caused attenuation in G. mellonella larvae and reduced growth in unpasteurized cow's milk and lactating mouse mammary glands. Our results confirm that iron scavenging by the ferric dicitrate receptor, which is strongly associated with MPEC strains, is required for MPEC growth and may influence disease severity in mastitis infections.IMPORTANCE Mastitis caused by E. coli inflicts substantial burdens on the health and productivity of dairy animals. Strains causing mastitis may express genes that distinguish them from other E. coli strains and promote infection of mammary glands, but these have not been identified. Using a highly virulent strain, we employed genome-wide mutagenesis and sequencing to discover genes that contribute to mastitis. This extensive data set represents a screen for mastitis-associated E. coli fitness factors and provides the following contributions to the field: (i) global comparison of genes required for different aspects of mastitis infection, (ii) discovery of a unique capsule that contributes to virulence, and (iii) conclusive evidence for the crucial role of iron-scavenging systems in mastitis, particularly the ferric dicitrate transport system. Similar approaches applied to other mastitis-associated strains will uncover conserved targets for prevention or treatment and provide a better understanding of their relationship to other E. coli pathogens.
| S-EPMC5752858 | biostudies-literature