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1.
The emergence of bacterial resistance to antibiotics is a major health problem and, therefore, it is critical to develop new antibiotics with novel modes of action. FtsZ, a tubulin-like GTPase, plays an essential role in bacterial cell division, and its homologs are present in almost all eubacteria and archaea. During cell division, FtsZ forms polymers in the presence of GTP that recruit other division proteins to make the cell division apparatus. Therefore, inhibition of FtsZ polymerization will prevent cells from dividing, leading to cell death. Using a fluorescent FtsZ polymerization assay, the screening of >100,000 extracts of microbial fermentation broths and plants followed by fractionation led to the identification of viriditoxin, which blocked FtsZ polymerization with an IC50 of 8.2 microg/ml and concomitant GTPase inhibition with an IC50 of 7.0 microg/ml. That the mode of antibacterial action of viriditoxin is via inhibition of FtsZ was confirmed by the observation of its effects on cell morphology, macromolecular synthesis, DNA-damage response, and increased minimum inhibitory concentration as a result of an increase in the expression of the FtsZ protein. Viriditoxin exhibited broad-spectrum antibacterial activity against clinically relevant Gram-positive pathogens, including methicillin-resistant Staphylococcus aureus and vancomycin-resistant Enterococci, without affecting the viability of eukaryotic cells.  相似文献   

2.
A multiplex PCR-based method was designed for the simultaneous detection of the main pathogens involved in warm-water streptococcosis in fish (Streptococcus iniae, Streptococcus difficilis, Streptococcus parauberis, and Lactococcus garvieae). Each of the four pairs of oligonucleotide primers exclusively amplified the targeted gene of the specific microorganism. The sensitivity of the multiplex PCR using purified DNA was 25 pg for S. iniae, 12.5 pg for S. difficilis, 50 pg for S. parauberis, and 30 pg for L. garvieae. The multiplex PCR assay was useful for the specific detection of the four species of bacteria not only in pure culture but also in inoculated fish tissue homogenates and naturally infected fish. Therefore, this method could be a useful alternative to the culture-based method for the routine diagnosis of warm-water streptococcal infections in fish.  相似文献   

3.
Viriditoxin activates ATP hydrolysis (ATPase) and swelling in rat liver mitochondria. The monocarboxylic ionophore of divalent cations, A23187, inhibits both activities at low concentrations of viriditoxin, but does not inhibit the ATPase induced by viriditoxin at concentrations above 2.5 × 10?5M. However, the monocarboxylic ionophore of monovalent cations, monensin, has no effect on the viriditoxin induced ATPase, but inhibits the valinomycin induced activity. Viriditoxin may facilitate the active transport of membrane bound calcium into the matrix of mitochondria  相似文献   

4.
Streptococcus iniae was isolated from diseased wild fish collected near a mariculture facility where gilthead sea bream and European sea bass exhibited a similar infection. Species-specific PCR and ribotyping confirmed that wild and cultured fish were infected by a single S. iniae clone. Wild fish are therefore potential amplifiers of pathogenic S. iniae strains.  相似文献   

5.
Streptococcus iniae is a Gram-positive bacterium and a severe pathogen to a wide range of economically important fish species. In addition, S. iniae is also a zoonotic pathogen and can cause serious infections in humans. In this study, we identified from a pathogenic S. iniae strain a putative bacteriocin, Sil, and examined its biological activity. Sil is composed of 101 amino acid residues and shares 35.6% overall sequence identity with the lactococcin 972 of Lactococcus lactis. Immunoblot analysis showed that Sil was secreted by S. iniae into the extracellular milieu. Purified recombinant Sil (rSil) exhibited a dose-dependent inhibitory effect on the growth of Bacillus subtilis but had no impact on the growths of other 16 Gram-positive bacteria and 10 Gram-negative bacteria representing 23 different bacterial species. Treatment of rSil by heating at 50°C abolished the activity of rSil. rSil bound to the surface of B. subtilis but induced no killing of the target cells. Cellular study revealed that rSil interacted with turbot (Scophthalmus maximus) head kidney monocytes and inhibited the innate immune response of the cells, which led to enhanced cellular infection of S. iniae. Antibody blocking of the extracellular Sil produced by S. iniae significantly attenuated the infectivity of S. iniae. Consistent with these in vitro observations, in vivo study showed that administration of turbot with rSil prior to S. iniae infection significantly increased bacterial dissemination and colonization in fish tissues. Taken together, these results indicate that Sil is a novel virulence-associated bacteriostatic and an immunoregulator that promotes S. iniae infection by impairing the immune defense of host fish.  相似文献   

6.
Two bacterial etiological agents of the disease, Streptococcus iniae and Streptococcus parauberis has been associated with fish mortalities and heavy economic loss in all over the world. Bacterial identification based on 16S rRNA sequencing is very fast, accurate and reliable in comparison to other traditional phenotype methodologies. In this study, we investigate the usefulness of this method for diagnosis and identification of Streptococcus species. We have selected 61 phylogeographic strains of Streptococcus (34 strains of S. iniae and 27 strains of S. parauberis) and designed the universal primer against the identified most hypervariable region of the 16S rRNA gene. Our universal primer able to identify any geographical strains and offers a useful and fast alternative in a clinical laboratory under routine conditions. Based on our studies, we have developed an algorithm for appropriate control of S. iniae and S. parauberis disease. We suggested the phenotype observation along with universal primer combination to detect any kind of infection or carriers at early stages.  相似文献   

7.
Streptococcus iniae has become one of the most serious aquatic pathogens in the last decade, causing large losses in wild and farmed fish worldwide. There is clear evidence that this pathogen is capable not only of causing serious disease in fish but also of being transferred to and infecting humans. In this study, we investigate the interaction of S. iniae with two murine macrophage cell lines, J774-A1 and RAW 264.7. Cytotoxicity assay demonstrated significant differences between live and UV-light killed IUSA-1 strains. The burst respiratory activity decreased to baseline after 1 and 4 h of exposure for J774-A1 and RAW 264.7, respectively. Immunofluorescent and ultrastructural study of infected cells confirmed the intracellular localization of bacteria at 1 h and 24 h post-infection. Using qRT-PCR arrays, we investigated the changes in the gene expression of immune relevant genes associated with macrophage activation. In this screening, we identified 11 of 84 genes up-regulated, we observed over-expression of pro-inflammatory response as IL-1α, IL-1β, and TNF-α, without a good anti-inflammatory response. Present findings suggest a capacity of S. iniae to modulate a mammalian macrophages cell lines to their survival and replication intracellular, which makes this cell type as a reservoir for continued infection.  相似文献   

8.
Inhibition of the functional activity of Filamenting temperature-sensitive mutant Z (FtsZ) protein, an essential and highly conserved bacterial cytokinesis protein, is a promising approach for the development of a new class of antibacterial agents. Berberine, a benzylisoquinoline alkaloid widely used in traditional Chinese and native American medicines for its antimicrobial properties, has been recently reported to inhibit FtsZ. Using a combination of in silico structure-based design and in vitro biological assays, 9-phenoxyalkyl berberine derivatives were identified as potent FtsZ inhibitors. Compared to the parent compound berberine, the derivatives showed a significant enhancement of antibacterial activity against clinically relevant bacteria, and an improved potency against the GTPase activity and polymerization of FtsZ. The most potent compound 2 strongly inhibited the proliferation of Gram-positive bacteria, including methicillin-resistant S. aureus and vancomycin-resistant E. faecium, with MIC values between 2 and 4 µg/mL, and was active against the Gram-negative E. coli and K. pneumoniae, with MIC values of 32 and 64 µg/mL respectively. The compound perturbed the formation of cytokinetic Z-ring in E. coli. Also, the compound interfered with in vitro polymerization of S. aureus FtsZ. Taken together, the chemical modification of berberine with 9-phenoxyalkyl substituent groups greatly improved the antibacterial activity via targeting FtsZ.  相似文献   

9.
Seaweeds producing natural products with in vitro efficacy against bacterial pathogens offer the opportunity for therapeutic services in fish aquaculture, for example inhibiting the growth of pathogens through the direct release of bioactive metabolites into the culture environment. The red alga Asparagopsis taxiformis was used to test this hypothesis because of its effective natural products and mechanisms to release these metabolites into the surrounding environment and because it can be cultivated intensively in land-based systems. Here we quantify the release, accumulation and residence time of the major halogenated metabolites from A. taxiformis in the cultivation medium and subsequently test the in vitro bioactivity of these water-soluble metabolites against multiple strains of the pathogenic bacterium Streptococcus iniae. The two major halogenated metabolites in A. taxiformis (bromoform and dibromoacetic acid) were released into the water and reaching steady-state concentrations of 27.7?±?1.5 μg L?1 and 4.8?±?1.3 μg L?1, respectively, under standard cultivation conditions. This water delayed the in vitro growth of the pathogenic bacteria S. iniae but did not prevent it. Increasing the levels of A. taxiformis metabolites in the water by three orders of magnitude with aqueous extracts of biomass inhibited the growth of S. iniae, but it was also toxic to fish. In an immersion challenge experiment, intermediate non-ichthyotoxic levels of A. taxiformis metabolites were ineffective as a treatment for barramundi infected by S. iniae. The evidence suggests that there is little potential for A. taxiformis bioactive metabolites to provide therapy services to fish infected with S. iniae.  相似文献   

10.
Streptococcus iniae causes severe septicemia and meningitis in farmed fish and is also occasionally zoonotic. Vaccination against S. iniae is problematic, with frequent breakdown of protection in vaccinated fish. The major protective antigens in S. iniae are the polysaccharides of the capsule, which are essential for virulence. Capsular biosynthesis is driven and regulated by a 21-kb operon comprising up to 20 genes. In a long-term study, we have sequenced the capsular operon of strains that have been used in autogenous vaccines across Australia and compared it with the capsular operon sequences of strains subsequently isolated from infected vaccinated fish. Intriguingly, strains isolated from vaccinated fish that subsequently become infected have coding mutations that are confined to a limited number of genes in the cps operon, with the remainder of the genes in the operon remaining stable. Mutations in strains in diseased vaccinated fish occur in key genes in the capsular operon that are associated with polysaccharide configuration (cpsG) and with regulation of biosynthesis (cpsD and cpsE). This, along with high ratios of nonsynonymous to synonymous mutations within the cps genes, suggests that immune response directed predominantly against capsular polysaccharide may be driving evolution in a very specific set of genes in the operon. From these data, it may be possible to design a simple polyvalent vaccine with a greater operational life span than the current monovalent killed bacterins.  相似文献   

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