RESUMO
Prevalence studies revealed that one-third of the human population is chronically infected with Toxoplasma gondii. Presently, such infections are without medical treatment that effectively eradicates the parasite once it is in its latent form. Moreover, the therapeutics used to treat acute infections are poorly tolerated by patients and also cause the parasite to convert into long-lasting tissue cysts. Hence, there is a dire need for compounds with antiparasitic activity against all forms of T. gondii. This study examines the antiparasitic capacity of nine novel bisphenol Z (BPZ) derivatives to determine whether they possessed any activity that prevented T. gondii replication. To begin assessing the efficacy of the novel derivatives, parasites were treated with increasing concentrations of the compounds, then doubling assays and MitoTracker staining were performed. Three of the nine compounds demonstrated strong inhibitory activity, i.e., parasite replication significantly decreased with higher concentrations. Additionally, many of the treated parasites exhibited decreases in fluorescent signaling and disruption of mitochondrial morphology. These findings suggest that bisphenol Z compounds disrupt mitochondrial function to inhibit parasite replication and may provide a foundation for the development of new and effective treatment modalities against T. gondii.
RESUMO
The continued evolution of antibiotic resistance has increased the urgency for new antibiotic development, leading to exploration of non-traditional sources. In particular, snake venom has garnered attention for its potent antibacterial properties. Numerous studies describing snake venom proteomic composition as well as antibiotic efficacy have created an opportunity to synthesize relationships between venom proteomes and their antibacterial properties. Using literature reported values from peer-reviewed studies, our study generated models to predict efficacy given venom protein family composition, snake taxonomic family, bacterial Gram stain, bacterial morphology, and bacterial respiration strategy. We then applied our predictive models to untested snake species with known venom proteomic compositions. Overall, our results provide potential protein families that serve as accurate predictors of efficacy as well as promising organisms in terms of antibacterial properties of venom. The results from this study suggest potential future research trajectories for antibacterial properties in snake venom by offering hypotheses for a variety of taxa.
Assuntos
Antibacterianos/farmacologia , Antivenenos/farmacologia , Bactérias/crescimento & desenvolvimento , Proteoma/metabolismo , Venenos de Serpentes/farmacologia , Serpentes/metabolismo , Animais , Bactérias/efeitos dos fármacos , Proteoma/análiseRESUMO
With the threat of a post-antibiotic era looming, the search for new and effective antibiotics from novel sources is imperative. Not only has crude snake venom been shown to be effective, but specific components within the venoms, such as Phospholipase A2s and l-amino acid oxidases have been isolated and demonstrated to be effective as well. Despite numerous studies being completed on snake venoms, there is a heavy bias towards utilizing the venoms from the highly toxic Elapidae and Viperidae species. Very few studies have been conducted on the less toxic, but taxonomically more diverse, Colubridae. Furthermore, an extensive review of the literature examining the efficacy and potential specificity of these venoms has not been completed. Therefore, the aims of this study were to elucidate any similarities in snake venoms as well as investigate the efficacy of snake venom antimicrobial properties towards morphologically and metabolically diverse microbial classes and the prevalence of snake species with antimicrobial properties within each snake family. The results indicate that snake venoms and their isolated components are powerful antimicrobial agents but vary in efficacy towards different microbial classes. Furthermore, due to similarities in venom composition, and limited preliminary studies, the less toxic Colubridae family may be a fruitful area of research to find novel antimicrobial agents that are less harmful to humans.
Assuntos
Antibacterianos/farmacologia , Bactérias/efeitos dos fármacos , Venenos de Serpentes/química , Venenos de Serpentes/farmacologia , Serpentes/classificação , Animais , Antibacterianos/química , Análise por Conglomerados , Serpentes/fisiologiaRESUMO
The ionophore monensin displays potent activities against several coccidian parasites of veterinary and medical importance including the opportunistic pathogen of humans, Toxoplasma gondii. While monensin is used widely in animals, toxicity impedes its use in humans. Nonetheless, given its potency, understanding its mode of action would reveal vulnerable aspects of the parasite that can be exploited for drug development. We previously established that monensin induces Toxoplasma to undergo cell cycle arrest and an autophagy-like cell death. Interestingly, these effects are dependent on the mitochondrion-localized TgMSH-1 protein, suggesting that monensin disrupts mitochondrial function. We demonstrate that monensin treatment results in decreased mitochondrial membrane potential and altered morphology. These effects are mitigated by the antioxidant compound N-acetyl-cysteine suggesting that monensin causes an oxidative stress, which was indeed the case based on direct detection of reactive oxygen species. Moreover, over-expression of the antioxidant proteins glutaredoxin and peroxiredoxin 2 protect Toxoplasma from the deleterious effects of monensin. Thus, our studies show that the effects of monensin on Toxoplasma are due to a disruption of mitochondrial function caused by the induction of an oxidative stress and implicate parasite redox biology as a viable target for the development of drugs against Toxoplasma and related pathogenic parasites.
Assuntos
Mitocôndrias/efeitos dos fármacos , Monensin/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Toxoplasma/efeitos dos fármacos , Acetilcisteína/farmacologia , Animais , Células Cultivadas , Fibroblastos/citologia , Fibroblastos/efeitos dos fármacos , Fibroblastos/parasitologia , Prepúcio do Pênis/citologia , Sequestradores de Radicais Livres/farmacologia , Expressão Gênica/efeitos dos fármacos , Glutarredoxinas/genética , Glutarredoxinas/metabolismo , Complexo de Golgi/efeitos dos fármacos , Humanos , Masculino , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Microscopia de Fluorescência , Mitocôndrias/metabolismo , Peroxirredoxinas/genética , Peroxirredoxinas/metabolismo , Ionóforos de Próton/farmacologia , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Toxoplasma/genética , Toxoplasma/metabolismoRESUMO
The UL4 gene is conserved within the genome of defective interfering particles of equine herpesvirus type 1 (EHV-1) that mediate persistent infection. Here, we show that the UL4 protein inhibits EHV-1 reporter gene expression by decreasing the level of transcribed mRNA. The UL4 protein did not bind any gene class of EHV-1 promoters in electromobility or chromatin immunoprecipitation assays, but directly interacted with the TATA box-binding protein (TBP) and the carboxy-terminal domain of RNA polymerase II both in vitro (GST-pulldown assays) and in infected cells (coimmunoprecipitation analyses). Microarray analyses of the expression of the 78 EHV-1 genes revealed that viral late genes important for virion assembly displayed enhanced expression in cells infected with UL4-null virus as compared to wild-type or UL4-restored EHV-1. Quantitative PCR analyses showed that viral DNA replication was not retarded in cells infected with the UL4-null virus as compared to wild-type EHV-1.
Assuntos
Regulação Viral da Expressão Gênica , Infecções por Herpesviridae/metabolismo , Herpesvirus Equídeo 1/genética , Herpesvirus Equídeo 1/metabolismo , RNA Polimerase II/metabolismo , Proteína de Ligação a TATA-Box/metabolismo , Proteínas Virais/metabolismo , Animais , Linhagem Celular , Infecções por Herpesviridae/enzimologia , Infecções por Herpesviridae/genética , Infecções por Herpesviridae/virologia , Interações Hospedeiro-Patógeno , Humanos , Camundongos , Ligação Proteica , RNA Polimerase II/genética , Coelhos , Proteína de Ligação a TATA-Box/genética , Proteínas Virais/genéticaRESUMO
Serial, high multiplicity passage of equine herpesvirus 1 (EHV-1) leads to the generation of defective interfering particles (DIP). EHV-1 DIP inhibit and interfere with the replication of standard EHV-1, establishing a state of persistent infection. These DIP package severely truncated and rearranged forms of the standard viral genome. Contained within the DIP genome are only three genes: UL3, UL4, and a unique hybrid gene (Hyb). The hybrid gene forms through a recombination event that fuses portions of the early regulatory IR4 and UL5 genes and is essential for DIP-mediated interference. The UL4 gene is an early gene dispensable for lytic replication and inhibits viral and cellular gene expression. However, the contribution of the UL4 gene during DIP-mediated persistent infection is unknown. Here, we describe the generation of a completely deleted UL4 virus and its use to investigate the role of the UL4 gene in the generation of the defective genome. Deletion of the UL4 gene resulted in delayed virus growth at late times post-infection. Cells infected with a mutant EHV-1 that lacked expression of the UL4 protein due to an inserted stop codon in the UL4 gene produced defective particles, while cells infected with a mutant EHV-1 that had the complete UL4 gene sequence deleted were unable to produce DIP. These data suggest that the UL4 gene sequence, but not the UL4 protein, is critical for the generation of defective interfering particles.
Assuntos
Vírus Defeituosos/genética , Deleção de Genes , Herpesvirus Equídeo 1/fisiologia , Proteínas Virais/metabolismo , Animais , Linhagem Celular , Genes Virais , Herpesvirus Equídeo 1/genética , Camundongos , Coelhos , Proteínas Virais/genética , Montagem de VírusRESUMO
The UL3 gene of equine herpesvirus-1 (EHV-1) is retained in the genome of defective interfering particles and encodes a ~33kDa myristylated protein. Further characterization showed that the UL3 gene is trans-activated only by the sole immediate early (IE) protein and encodes an early protein that is dispensable for EHV-1 replication and localizes in the tegument of purified virions. UL3-deleted EHV-1 (vL11ΔUL3) exhibits properties of host cell tropism, plaque size, and growth kinetics similar to those of the parental virus. Expression levels of EHV-1 proteins representative of all three gene classes in vL11ΔUL3-infected cells were identical to those in cells infected with parental virus. Mice intranasally infected with vL11ΔUL3 and parental virus showed no significant difference in mortality or virus lung titers. These findings suggest that the UL3 protein does not play a major role in the biology of EHV-1 in cell culture or virulence in the mouse.
Assuntos
Infecções por Herpesviridae/veterinária , Herpesvirus Equídeo 1/fisiologia , Herpesvirus Equídeo 1/patogenicidade , Doenças dos Cavalos/virologia , Proteínas Imediatamente Precoces/metabolismo , Proteínas Virais/metabolismo , Replicação Viral , Animais , Linhagem Celular , Modelos Animais de Doenças , Regulação Viral da Expressão Gênica , Infecções por Herpesviridae/virologia , Herpesvirus Equídeo 1/genética , Cavalos , Humanos , Proteínas Imediatamente Precoces/genética , Camundongos , Camundongos Endogâmicos CBA , Proteínas Virais/genética , VirulênciaRESUMO
Defective interfering particles (DIP) of equine herpesvirus 1 (EHV-1) inhibit standard virus replication and mediate persistent infection. The DIP genome is comprised of only three genes: UL3, UL4, and a hybrid gene composed of portions of the IR4 (EICP22) and UL5 (EICP27) genes. The hybrid gene is important for DIP interference, but the function(s) of the UL3 and UL4 genes are unknown. Here, we show that UL4 is an early gene activated solely by the immediate early protein. The UL4 protein (UL4P) was detected at 4hours post-infection, was localized throughout the nucleus and cytoplasm, and was not present in purified virions. EHV-1 lacking UL4P expression was infectious and displayed cell tropism and pathogenic properties in the mouse model similar to those of parental and revertant viruses. Reporter assays demonstrated that the UL4P has a broad inhibitory function, suggesting a potential role in establishing and/or maintaining DIP-mediated persistent infection.