RESUMO
The global poultry trend toward the more responsible use of antibiotics is becoming recurrent and has demanded the need to generate new natural alternatives. Probiotics have gained importance as an option to use as growth promoters. This study aimed to evaluate Bacillus subtillis QST713 as a substitute for an antibiotic growth promoter (BMD). A total of 150 male broilers were assigned to three dietary treatments: 1) control diet (CO), 2) control diet + 500 g/t of BMD (AGP), and 3) control diet + 100 g/t of B. subtilis QST713 (PB), respectively. Each treatment was monitored for 5 wk for the productive variables: body weight, accumulated feed consumption, food conversion, and European efficiency factor. At the end of each week, fresh fecal samples were cultured and quantified for E. coli, Enterococcus spp., and Lactobacillus spp. At the end of the trial, blood samples were analyzed for hemogram and intestinal samples (anterior portion) for histomorphometry. The data were statistically analyzed with an analysis of variance and subjected to a least significant difference test (Tukey). The zootechnical yields were similar in the AGP and PB groups (P Ë 0.05); both superior to the control group. In the hematological profiles, no difference was observed between the experimental groups. E. coli and Enterococcus counts were significantly lower (P Ë 0.05), and Lactobacillus counts were significantly (P Ë 0.05) higher in the PB group, relative to CO and AGP groups. No differences (P Ë 0.05) were found in bacterial counts between the CO and AGP groups. The intestinal mucosa and villi in the PB group were significantly (P Ë 0.05) longer and with less deeper crypts than CO and AGP groups. We conclude that B. subtillis QST713, used at the suggested commercial dose (100 g/ton), is an effective growth-promoting alternative to BMD that modulates the microbiota and intestinal architecture, thus producing zootechnical yields consistent with BMD.
Assuntos
Bacitracina , Probióticos , Ração Animal/análise , Animais , Bacillus subtilis , Bacitracina/farmacologia , Galinhas , Dieta/veterinária , Escherichia coli , MasculinoRESUMO
Membrane alanyl and glutamyl aminopeptidases (APN and APA, respectively) are established targets for the development of biomedical tools in human pathologies. APN overexpression correlates with the progression of tumours, including melanoma. Bacitracin, widely used as a topical antibiotic, inhibits subtilisin-like serine peptidases and disulphide isomerases. In the present contribution, we demonstrate that bacitracin is a non-competitive α = 1 and α < 1 inhibitor of porcine kidney APN and APA, respectively, with Ki values in the micromolar range. To test a potential application of this result, we assayed the effect of bacitracin on murine melanoma MB16F10 cell line viability. We demonstrated the cell line expresses an APN-like activity inhibited by bacitracin and bestatin. Additionally, we identified a cytotoxic effect of bacitracin. Further experiments are required to understand in depth the mechanisms of action of bacitracin on melanoma cells. They will clarify the therapeutic potential of bacitracin for melanoma treatment.
Assuntos
Bacitracina , Antígenos CD13 , Glutamil Aminopeptidase/antagonistas & inibidores , Animais , Bacitracina/farmacologia , Antígenos CD13/antagonistas & inibidores , Linhagem Celular Tumoral , Rim , Camundongos , SuínosRESUMO
Bestatin and bacitracin are inhibitors of metallo aminopeptidases and bacterial proteases. However, their effects on other human peptidases, like dipeptidyl peptidase IV (DPP-IV, EC 3.4.14.5) are not established. Inhibitors of DPP-IV activity are used for treating type 2 diabetes mellitus, cancers and immune system diseases. Bacitracin and bestatin inhibited porcine membrane-bound DPP-IV (pDPP-IV) activity. Mechanisms were different, i.e. non-competitive with αâ¯>â¯1 (αâ¯=â¯3.9) and Ki value of 75⯵M for bestatin, and competitive with Ki value of 630⯵M for bacitracin. The binding mode in the tertiary complex enzyme:substrate:bestatin suggested the structural basis of the inhibitory effect and that bestatin is potentially selective for DPP-IV, ineffective vs. S9 family members dipeptidyl peptidase 8/9 and fibroblast activation protein. In the human melanoma MeWo cell line, bestatin and bacitracin inhibited aminopeptidase N (APN) and DPP-IV activities, reduced cell viability and increased DNA fragmentation, suggesting induction of apoptosis. Since bacitracin and bestatin are already marketed drugs, studying in depth the molecular mechanisms underlying their effects on melanoma cells is warranted. Additionally, bestatin emerges as a new lead compound for the development of DPP-IV inhibitors, and a promising dual APN/DPP-IV inhibitor for the treatment of pathologies in which both enzymes are upregulated.
Assuntos
Antineoplásicos/farmacologia , Bacitracina/farmacologia , Dipeptidil Peptidase 4/metabolismo , Inibidores da Dipeptidil Peptidase IV/farmacologia , Leucina/análogos & derivados , Melanoma/enzimologia , Animais , Linhagem Celular Tumoral , Membrana Celular/efeitos dos fármacos , Membrana Celular/enzimologia , Sobrevivência Celular/efeitos dos fármacos , Dipeptidil Peptidase 4/química , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Rim/enzimologia , Leucina/farmacologia , Melanoma/tratamento farmacológico , Modelos Moleculares , Relação Estrutura-Atividade , SuínosRESUMO
The purpose of this study was to verify the ability of a probiotic in the feed to maintain the stability of the gut microbiota in chickens after antibiotic therapy and its association with growth performance. One thousand six hundred twenty 1-day-old Cobb male were housed in floor pens (36 pens, 45 birds/pen) and were fed corn-/soya bean meal-based diets supplemented with or without probiotic (Bacillus subtilis) during the entire rearing phase. From 21 to 24 days of age (three consecutive days), the chickens were submitted to antibiotic therapy via drinking water (bacitracin and neomycin) in order to mimic a field treatment and induce dysbiosis. Growth performance was monitored until 42 days of age. At 2, 4 and 6 days after antibiotic therapy, three chickens from each pen were euthanized and the contents of the small intestine and caeca were collected and pooled. The trial was conducted with four treatments and nine replicates in a 2 × 2 factorial arrangement for performance characteristics (with and without probiotic × with and without antibiotic therapy); for the intestinal microbiota, it was in a 2 × 2 × 3 factorial arrangement (with and without probiotic × with and without antibiotic therapy × 2, 4 and 6 days after the antibiotic therapy) with three replicates per treatment. Terminal restriction length polymorphism (T-RFLP) analysis showed that the structure of gut bacterial community was shaped by the intestinal segment and by the time after the antibiotic therapy. The number of 16S rDNAs copies in caecum contents decreased with time after the therapeutic treatment. The antibiotic therapy and dietary probiotic supplementation decreased richness and diversity indexes in the caecal contents. The improved performance observed in birds supplemented with probiotic may be related to changes promoted by the feed additive in the structure of the intestinal bacterial communities and phylogenetic groups. Antibiotic therapy modified the bacterial structure, but did not cause loss of broiler performance.
Assuntos
Ração Animal/análise , Bacitracina/farmacologia , Galinhas/crescimento & desenvolvimento , Microbioma Gastrointestinal/efeitos dos fármacos , Neomicina/farmacologia , Probióticos/farmacologia , Fenômenos Fisiológicos da Nutrição Animal , Animais , Antibacterianos/administração & dosagem , Antibacterianos/farmacologia , Bacitracina/administração & dosagem , Galinhas/microbiologia , Dieta/veterinária , Neomicina/administração & dosagem , Distribuição AleatóriaRESUMO
Antibiotic growth promoters have been used for decades in poultry farming as a tool to maintain bird health and improve growth performance. Global concern about the recurrent emergence and spreading of antimicrobial resistance is challenging the livestock producers to search for alternatives to feed added antibiotics. The use of phytogenic compounds appears as a feasible option due to their ability to emulate the bioactive properties of antibiotics. However, detailed description about the effects of in-feed antibiotics and alternative natural products on chicken intestinal microbiota is lacking. High-throughput sequencing of 16S rRNA gene was used to study composition of cecal microbiota in broiler chickens supplemented with either bacitracin or a blend of chestnut and quebracho tannins over a 30-day grow-out period. Both tannins and bacitracin had a significant impact on diversity of cecal microbiota. Bacitracin consistently decreased Bifidobacterium while other bacterial groups were affected only at certain times. Tannins-fed chickens showed a drastic decrease in genus Bacteroides while certain members of order Clostridiales mainly belonging to the families Ruminococcaceae and Lachnospiraceae were increased. Different members of these groups have been associated with an improvement of intestinal health and feed efficiency in poultry, suggesting that these bacteria could be associated with productive performance of birds.
Assuntos
Bacitracina/farmacologia , Galinhas/microbiologia , Microbioma Gastrointestinal/efeitos dos fármacos , Microbiota/efeitos dos fármacos , Taninos/farmacologia , Ração Animal , Animais , Antibacterianos/farmacologia , Bacteroides/efeitos dos fármacos , Bacteroides/genética , Bifidobacterium/efeitos dos fármacos , Bifidobacterium/genética , Clostridiales/efeitos dos fármacos , Clostridiales/genética , Intestinos/microbiologia , Microbiota/genética , RNA Ribossômico 16S/genéticaRESUMO
Antimicrobials are sometimes given to food animals at low doses in order to promote faster growth. However, the mechanisms by which those drugs improve performance are not fully understood. This study aimed to investigate the impact of zinc bacitracin (55g/ton), enramycin (10g/ton); halquinol® (30g/ton); virginiamycin (16,5g/ton) and avilamycin (10g/ton) on the cecal microbiota of broiler chicken, compared to a control group. Six hundred and twenty four chicks (Cobb 500) arriving to an experimental unit were randomly assigned into each treatment with four repetitions per treatment. The cecal content of 16 animals per treatment (n = 96) was used for DNA extraction and sequencing of the V4 region of the 16S rRNA gene using Illumina technology. The use of antimicrobials induced significant changes in membership but not in structure of the cecal microbiota compared to the control group, suggesting a greater impact on the less abundant species of bacteria present in that environment. Halquinol was the only drug that did not affect microbial membership. Firmicutes comprised the major bacterial phylum present in the cecum of all groups. There was no statistical difference in relative abundances of the main phyla between treated animals and the control group (all P>0.05). Treatment with enramycin was associated with decreased richness and with lower relative abundance of unclassified Firmicutes, Clostridium XI, unclassified Peptostreptococcaceae (all P<0.001) and greater abundance of Clostridium XIVb (P = 0.004) and Anaerosporobacter spp. (P = 0.015), and treatment with bacitracin with greater relative abundance of Bilophila spp. (P = 0.004). Several bacterial genera were identified as representative of usage of each drug. This study used high throughput sequencing to characterize the impact of several antimicrobials in broiler chicken under controlled conditions and add new insights to the current knowledge on how AGPs affect the cecal microbiota of chicken.
Assuntos
Ração Animal , Antibacterianos/farmacologia , Ceco/microbiologia , Galinhas/microbiologia , Aditivos Alimentares/farmacologia , Microbioma Gastrointestinal/efeitos dos fármacos , Animais , Antibacterianos/administração & dosagem , Bacitracina/farmacologia , Bactérias/classificação , Bactérias/efeitos dos fármacos , Galinhas/crescimento & desenvolvimento , Cloroquinolinóis/farmacologia , Variação Genética , Oligossacarídeos/farmacologia , Peptídeos/farmacologia , Análise de Componente Principal , Distribuição Aleatória , Ribotipagem , Virginiamicina/farmacologiaRESUMO
BACKGROUND:: Topical antimicrobial drugs are indicated for limited superficial pyodermitis treatment, although they are largely used as self-prescribed medication for a variety of inflammatory dermatoses, including atopic dermatitis. Monitoring bacterial susceptibility to these drugs is difficult, given the paucity of laboratory standardization. OBJECTIVE:: To evaluate the prevalence of Staphylococcus aureus topical antimicrobial drug resistance in atopic dermatitis patients. METHODS:: We conducted a cross-sectional study of children and adults diagnosed with atopic dermatitis and S. aureus colonization. We used miscellaneous literature reported breakpoints to define S. aureus resistance to mupirocin, fusidic acid, gentamicin, neomycin and bacitracin. RESULTS:: A total of 91 patients were included and 100 S. aureus isolates were analyzed. All strains were methicillin-susceptible S. aureus. We found a low prevalence of mupirocin and fusidic acid resistance (1.1% and 5.9%, respectively), but high levels of neomycin and bacitracin resistance (42.6% and 100%, respectively). Fusidic acid resistance was associated with more severe atopic dermatitis, demonstrated by higher EASI scores (median 17.8 vs 5.7, p=.009). Our results also corroborate the literature on the absence of cross-resistance between the aminoglycosides neomycin and gentamicin. CONCLUSIONS:: Our data, in a southern Brazilian sample of AD patients, revealed a low prevalence of mupirocin and fusidic acid resistance of S. aureus atopic eczema colonizer strains. However, for neomycin and bacitracin, which are commonly used topical antimicrobial drugs in Brazil, high levels of resistance were identified. Further restrictions on the use of these antimicrobials seem necessary to keep resistance as low as possible.
Assuntos
Antibacterianos/farmacologia , Dermatite Atópica/microbiologia , Farmacorresistência Bacteriana , Staphylococcus aureus/efeitos dos fármacos , Adolescente , Adulto , Bacitracina/farmacologia , Criança , Pré-Escolar , Estudos Transversais , Testes de Sensibilidade a Antimicrobianos por Disco-Difusão/métodos , Feminino , Ácido Fusídico/farmacologia , Gentamicinas/farmacologia , Humanos , Lactente , Masculino , Mupirocina/farmacologia , Neomicina/farmacologia , Adulto JovemRESUMO
Abstract: Background: Topical antimicrobial drugs are indicated for limited superficial pyodermitis treatment, although they are largely used as self-prescribed medication for a variety of inflammatory dermatoses, including atopic dermatitis. Monitoring bacterial susceptibility to these drugs is difficult, given the paucity of laboratory standardization. Objective: To evaluate the prevalence of Staphylococcus aureus topical antimicrobial drug resistance in atopic dermatitis patients. Methods: We conducted a cross-sectional study of children and adults diagnosed with atopic dermatitis and S. aureus colonization. We used miscellaneous literature reported breakpoints to define S. aureus resistance to mupirocin, fusidic acid, gentamicin, neomycin and bacitracin. Results: A total of 91 patients were included and 100 S. aureus isolates were analyzed. All strains were methicillin-susceptible S. aureus. We found a low prevalence of mupirocin and fusidic acid resistance (1.1% and 5.9%, respectively), but high levels of neomycin and bacitracin resistance (42.6% and 100%, respectively). Fusidic acid resistance was associated with more severe atopic dermatitis, demonstrated by higher EASI scores (median 17.8 vs 5.7, p=.009). Our results also corroborate the literature on the absence of cross-resistance between the aminoglycosides neomycin and gentamicin. Conclusions: Our data, in a southern Brazilian sample of AD patients, revealed a low prevalence of mupirocin and fusidic acid resistance of S. aureus atopic eczema colonizer strains. However, for neomycin and bacitracin, which are commonly used topical antimicrobial drugs in Brazil, high levels of resistance were identified. Further restrictions on the use of these antimicrobials seem necessary to keep resistance as low as possible.
Assuntos
Humanos , Masculino , Feminino , Lactente , Pré-Escolar , Criança , Adolescente , Adulto , Adulto Jovem , Staphylococcus aureus/efeitos dos fármacos , Farmacorresistência Bacteriana , Dermatite Atópica/microbiologia , Antibacterianos/farmacologia , Bacitracina/farmacologia , Gentamicinas/farmacologia , Neomicina/farmacologia , Estudos Transversais , Mupirocina/farmacologia , Testes de Sensibilidade a Antimicrobianos por Disco-Difusão/métodos , Ácido Fusídico/farmacologiaRESUMO
Food handlers carrying enterotoxin-producing Staphylococcus are a potential source of food poisoning. The aim of this study was to analyze genes encoding enterotoxins in coagulase-positive Staphylococcus (CoPS) and coagulase-negative Staphylococcus (CoNS) isolated from the anterior nostrils and hands of food handlers at a university restaurant in the city of Natal, Northeast Brazil. Thirty food handlers were screened for the study. The isolates were subjected to Gram staining, a bacitracin sensitivity test, mannitol fermentation, and catalase and coagulase tests. CoNS and CoPS strains were subsequently identified by a Vitek 2 System (BioMerieux, France) and various biochemical tests. Polymerase chain reaction was used to detect genes for enterotoxins A, B, C, D, E, G, H, and I (sea, seb, sec, sed, see, seg, seh, and sei) and a disc-diffusion method was used to determine susceptibility to several classes of antimicrobials. All food handlers presented staphylococci on their hands and/or noses. The study found 58 Staphylococcus spp., of which 20.7% were CoPS and 79.3% were CoNS. S. epidermidis was the most prevalent species. Twenty-nine staphylococci (50%) were positive for one or more enterotoxin genes, and the most prevalent genes were seg and sei, each with a frequency of 29.3%. Indeed, CoNS encoded a high percentage of enterotoxin genes (43.5%). However, S. aureus encoded even more enterotoxin genes (75%). Most isolates showed sensitivity to the antibiotics used for testing, except for penicillin (only 35% sensitive). The results from this study reinforce that coagulase-negative as well as coagulase-positive staphylococci isolated from food handlers are capable of genotypic enterotoxigenicity.
Assuntos
Enterotoxinas/genética , Manipulação de Alimentos , Genes Bacterianos/genética , Staphylococcus/isolamento & purificação , Antibacterianos/farmacologia , Bacitracina/farmacologia , Brasil , Enterotoxinas/isolamento & purificação , Mãos/microbiologia , Humanos , Testes de Sensibilidade Microbiana , Nariz/microbiologia , Penicilinas/farmacologia , Restaurantes , Staphylococcus/efeitos dos fármacos , Staphylococcus/enzimologia , Staphylococcus/metabolismo , UniversidadesRESUMO
Guedes-Pinto paste is the filling material most employed in Brazil for endodontic treatment of deciduous teeth; however, the Rifocort® ointment has been removed. Thus, the aim of this study was to investigate the antimicrobial potential of filling pastes, by proposing three new pharmacological associations to replace Rifocort® ointment with drugs of already established antimicrobial power: Nebacetin® ointment, 2% Chlorhexidine Gluconate gel, and Maxitrol® ointment. A paste composed of Iodoform, Rifocort® ointment and Camphorated Paramonochlorophenol (CPC) was employed as the gold standard (G1). The other associations were: Iodoform, Nebacetin® ointment and CPC (G2); Iodoform, 2% Chlorhexidine Digluconate gel and CPC (G3); Iodoform, Maxitrol® ointment and CPC (G4). The associations were tested for Staphylococcus aureus (S. aureus), Streptococcus mutans (S. mutans), Streptococcus oralis (S. oralis), Enterococcus faecalis (E. faecalis), Escherichia coli (E. coli), and Bacillus subtilis (B. subtilis), using the methods of dilution on solid medium - orifice agar - and broth dilution. The results were tested using statistical analysis ANOVA and Kruskal-Wallis. They showed that all the pastes had a bacteriostatic effect on all the microorganisms, without any statistically significant difference, compared with G1. S. aureus was statistically significant (multiple comparison test of Tukey), insofar as G2 and G3 presented the worst and the best performance, respectively. All associations were bactericidal for E. coli, S. aureus, S. mutans and S. oralis. Only G3 and G4 were bactericidal for E. faecalis, whereas no product was bactericidal for B. subtilis. Thus, the tested pastes have antimicrobial potential and have proved acceptable for endodontic treatment of primary teeth.
Assuntos
Antibacterianos/farmacologia , Bactérias/efeitos dos fármacos , Materiais Restauradores do Canal Radicular/farmacologia , Dente Decíduo/efeitos dos fármacos , Análise de Variância , Bacitracina/farmacologia , Bactérias/crescimento & desenvolvimento , Clorexidina/análogos & derivados , Clorexidina/farmacologia , Combinação de Medicamentos , Fluprednisolona/farmacologia , Testes de Sensibilidade Microbiana , Neomicina/farmacologia , Pomadas , Polimixina B/farmacologia , Prednisolona/análogos & derivados , Prednisolona/farmacologia , Reprodutibilidade dos Testes , Rifamicinas/farmacologia , Estatísticas não Paramétricas , Fatores de TempoRESUMO
Guedes-Pinto paste is the filling material most employed in Brazil for endodontic treatment of deciduous teeth; however, the Rifocort® ointment has been removed. Thus, the aim of this study was to investigate the antimicrobial potential of filling pastes, by proposing three new pharmacological associations to replace Rifocort® ointment with drugs of already established antimicrobial power: Nebacetin® ointment, 2% Chlorhexidine Gluconate gel, and Maxitrol® ointment. A paste composed of Iodoform, Rifocort® ointment and Camphorated Paramonochlorophenol (CPC) was employed as the gold standard (G1). The other associations were: Iodoform, Nebacetin® ointment and CPC (G2); Iodoform, 2% Chlorhexidine Digluconate gel and CPC (G3); Iodoform, Maxitrol® ointment and CPC (G4). The associations were tested for Staphylococcus aureus (S. aureus), Streptococcus mutans (S. mutans), Streptococcus oralis (S. oralis), Enterococcus faecalis (E. faecalis), Escherichia coli (E. coli), and Bacillus subtilis (B. subtilis), using the methods of dilution on solid medium – orifice agar – and broth dilution. The results were tested using statistical analysis ANOVA and Kruskal-Wallis. They showed that all the pastes had a bacteriostatic effect on all the microorganisms, without any statistically significant difference, compared with G1. S. aureus was statistically significant (multiple comparison test of Tukey), insofar as G2 and G3 presented the worst and the best performance, respectively. All associations were bactericidal for E. coli, S. aureus, S. mutans and S. oralis. Only G3 and G4 were bactericidal for E. faecalis, whereas no product was bactericidal for B. subtilis. Thus, the tested pastes have antimicrobial potential and have proved acceptable for endodontic treatment of primary teeth.
Assuntos
Antibacterianos/farmacologia , Bactérias/efeitos dos fármacos , Materiais Restauradores do Canal Radicular/farmacologia , Dente Decíduo/efeitos dos fármacos , Análise de Variância , Bacitracina/farmacologia , Bactérias/crescimento & desenvolvimento , Clorexidina/análogos & derivados , Clorexidina/farmacologia , Combinação de Medicamentos , Fluprednisolona/farmacologia , Testes de Sensibilidade Microbiana , Neomicina/farmacologia , Pomadas , Polimixina B/farmacologia , Prednisolona/análogos & derivados , Prednisolona/farmacologia , Reprodutibilidade dos Testes , Rifamicinas/farmacologia , Estatísticas não Paramétricas , Fatores de TempoRESUMO
Although homologs of the ATP-dependent Lon protease exist in all domains of life, the relevance of this protease in archaeal physiology remains a mystery. In this study, we have constructed and phenotypically characterized deletion and conditional lon mutants in the model haloarchaeon Haloferax volcanii to elucidate the role of the unusual membrane-bound LonB protease in archaea. Hvlon could be deleted from the chromosome only when a copy of the wild type gene was provided in trans suggesting that Lon is essential for survival in this archaeon. Successful complementation of the lethal phenotype of ΔHvlon was attained by expression of the heterologous protease gene Nmlon from the haloalkaliphilic archaeon Natrialba magadii, meaning that the biological function of Lon is conserved in these organisms. Suboptimal cellular levels of Lon protein affected growth rate, cell shape, cell pigmentation, lipid composition and sensitivity to various antibiotics. The contents of bacterioruberins and some polar lipids were increased in the lon mutants suggesting that Lon is linked to maintenance of membrane lipid balance which likely affects cell viability in this archaeon. The phenotypes associated to a membrane-bound LonB protease mutant were examined for the first time providing insight on the relevance of this protease in archaeal physiology.
Assuntos
Proteínas Arqueais/genética , Haloferax volcanii/enzimologia , Lipídeos de Membrana/metabolismo , Peptídeo Hidrolases/genética , Antibacterianos/farmacologia , Proteínas Arqueais/metabolismo , Bacitracina/farmacologia , Sequência de Bases , Expressão Gênica , Regulação da Expressão Gênica em Archaea , Haloferax volcanii/efeitos dos fármacos , Lovastatina/farmacologia , Testes de Sensibilidade Microbiana , Viabilidade Microbiana/efeitos dos fármacos , Novobiocina/farmacologia , Peptídeo Hidrolases/metabolismo , Pigmentação , Ligação Proteica , Puromicina/farmacologiaRESUMO
UNLABELLED: The aim of this study was to evaluate the antibiotic susceptibility of Listeria innocua (L. innocua) and Listeria monocytogenes (L. monocytogenes) cells in the presence of citral and carvacrol at sublethal concentrations in an agar medium. The presence of terpenes in the L. monocytogenes and L. innocua culture medium provided a reduction in the minimal inhibitory concentration (MIC) of all the antibiotics tested. These effects were dependent on the concentration of terpenes present in the culture medium. The combination of citral and carvacrol potentiated antibiotic activity by reducing the MIC values of bacitracin and colistin from 32.0 and 128.0 µg ml⻹ to 1.0 and 2.0 µg ml⻹, respectively. Thus, both Listeria species became more susceptible to these drugs. In this way, the colistin and bacitracin resistance of L. monocytogenes and L. innocua was reversed in the presence of terpenes. Results obtained in this study show that the phytochemicals citral and carvacrol potentiate antibiotic activity, reducing the MIC values of cultured L. monocytogenes and L. innocua. SIGNIFICANCE AND IMPACT OF THE STUDY: Phytochemicals citral and carvacrol potentiate antibiotic activity of erythromycin, bacitracin and colistin by reducing the MIC values of cultured Listeria monocytogenes and Listeria innocua. This effect in reducing the MIC values of the antibiotics tested in both micro-organisms was increased when natural antimicrobials were combined. This finding indicated that the combination among terpenes and antibiotic may contribute in reducing the required dosage of antibiotics due to the possible effect of terpenes on permeation barrier of the micro-organism cell membrane.
Assuntos
Antibacterianos/farmacologia , Bacitracina/farmacologia , Colistina/farmacologia , Farmacorresistência Bacteriana/efeitos dos fármacos , Listeria monocytogenes/efeitos dos fármacos , Listeria/efeitos dos fármacos , Monoterpenos/farmacologia , Monoterpenos Acíclicos , Ágar/metabolismo , Cimenos , Listeria/crescimento & desenvolvimento , Listeria/fisiologia , Listeria monocytogenes/crescimento & desenvolvimento , Listeria monocytogenes/fisiologia , Testes de Sensibilidade MicrobianaRESUMO
OBJECTIVE: The recovery of mutans streptococci in saliva and dental biofilm samples depends, in part, on the culture medium used. In this study, we compared (i) the culture media Sucrose-Bacitracin agar (SB-20), Modified SB-20 (SB-20M) and Mitis Salivarius Bacitracin agar (MSB) in the count of colony forming units (cfu) of mutans streptococci and (ii) in the morphological and biochemical differentiation between Streptococcus mutans and Streptococcus sobrinus. DESIGN: Samples of non-stimulated saliva from 20 children were plated on SB-20, SB-20M and MSB, and incubated in microaerophilia at 37°C for 72h. Identification of microorganisms was based on analysis of colony morphology under stereomicroscopy. The biochemical identification of colonies was done by biochemical tests using sugar fermentation, resistance to bacitracin and hydrogen peroxide production. RESULTS: There was no significant difference (p>0.05) in the number of cfu of mutans streptococci recovered on SB-20 and SB-20M agar. Comparing the media, SB-20 and SB-20M yielded a larger number of mutans streptococci colonies (p<0.05) and were more effective than MSB in the identification of S. sobrinus (p<0.05), but not of S. mutans (p>0.05). CONCLUSION: There was no significant difference between SB-20 and SB-20M culture media in the count of mutans streptococci, demonstrating that the replacement of sucrose by coarse granular cane sugar did not alter the efficacy of the medium. Compared with MSB, SB-20 and SB-20M allowed counting a larger number of mutans streptococci colonies and a more effective morphological identification of S. sobrinus.
Assuntos
Ágar/classificação , Técnicas Bacteriológicas , Meios de Cultura/classificação , Streptococcus mutans/isolamento & purificação , Streptococcus sobrinus/isolamento & purificação , Antibacterianos/farmacologia , Bacitracina/farmacologia , Carga Bacteriana , Técnicas de Tipagem Bacteriana , Biofilmes , Criança , Pré-Escolar , Farmacorresistência Bacteriana , Fermentação , Humanos , Saliva/microbiologia , Sacarose/metabolismoRESUMO
OBJECTIVES: Determining the effect of membrane-impermeant thiol/disulfide exchange inhibitors on rhesus rotavirus infectivity in MA104 cells and investigating protein disulfide isomerase (PDI) as a potential target for these inhibitors. METHODS: Cells were treated with DTNB [5,5-dithio-bis-(2-nitrobenzoic acid)], bacitracin or anti-PDI antibodies and then infected with virus. Triple-layered particles (TLPs) were also pretreated with inhibitors before inoculation. The effects of these inhibitors on α-sarcin co-entry, virus binding to cells and PDI-TLP interaction were also examined. FACS analysis, cell-surface protein biotin-labeling, lipid-raft isolation and ELISA were performed to determine cell-surface PDI expression. RESULTS: Infectivity became reduced by 50% when cells or TLPs were treated with 1 or 6 mM DTNB, respectively; infectivity became reduced by 50% by 20 mM bacitracin treatment of cells whereas TLPs were insensitive to bacitracin treatment; anti-PDI antibodies decreased viral infectivity by about 45%. The presence of DTNB (2.5 mM) or bacitracin (20 mM) was unable to prevent virus binding to cells and rotavirus-induced α-sarcin co-entry. CONCLUSIONS: It was concluded that thiol/disulfide exchange was involved in rotavirus entry process and that cell-surface PDI was at least a potential target for DTNB and bacitracin-induced infectivity inhibition.
Assuntos
Isomerases de Dissulfetos de Proteínas/antagonistas & inibidores , Infecções por Rotavirus/tratamento farmacológico , Rotavirus/efeitos dos fármacos , Rotavirus/fisiologia , Ligação Viral/efeitos dos fármacos , Internalização do Vírus/efeitos dos fármacos , Animais , Antibacterianos/farmacologia , Anticorpos/imunologia , Bacitracina/farmacologia , Linhagem Celular , Dissulfetos/metabolismo , Ácido Ditionitrobenzoico/farmacologia , Endorribonucleases/metabolismo , Proteínas Fúngicas/metabolismo , Macaca mulatta , Isomerases de Dissulfetos de Proteínas/imunologia , Isomerases de Dissulfetos de Proteínas/metabolismo , Infecções por Rotavirus/metabolismo , Compostos de Sulfidrila/metabolismo , Reagentes de Sulfidrila/farmacologiaRESUMO
Mechanisms of leukocyte NADPH oxidase regulation remain actively investigated. We showed previously that vascular and macrophage oxidase complexes are regulated by the associated redox chaperone PDI. Here, we investigated the occurrence and possible underlying mechanisms of PDI-mediated regulation of neutrophil NADPH oxidase. In a semirecombinant cell-free system, PDI inhibitors scrRNase (100 µg/mL) or bacitracin (1 mM) near totally suppressed superoxide generation. Exogenously incubated, oxidized PDI increased (by ~40%), whereas PDIred diminished (by ~60%) superoxide generation. No change occurred after incubation with PDI serine-mutated in all four redox cysteines. Moreover, a mimetic CxxC PDI inhibited superoxide production by ~70%. Thus, oxidized PDI supports, whereas reduced PDI down-regulates, intrinsic membrane NADPH oxidase complex activity. In whole neutrophils, immunoprecipitation and colocalization experiments demonstrated PDI association with membrane complex subunits and prominent thiol-mediated interaction with p47(phox) in the cytosol fraction. Upon PMA stimulation, PDI was mobilized from azurophilic granules to cytosol but did not further accumulate in membranes, contrarily to p47(phox). PDI-p47(phox) association in cytosol increased concomitantly to opposite redox switches of both proteins; there was marked reductive shift of cytosol PDI and maintainance of predominantly oxidized PDI in the membrane. Pulldown assays further indicated predominant association between PDIred and p47(phox) in cytosol. Incubation of purified PDI (>80% reduced) and p47(phox) in vitro promoted their arachidonate-dependent association. Such PDI behavior is consistent with a novel cytosolic regulatory loop for oxidase complex (re)cycling. Altogether, PDI seems to exhibit a supportive effect on NADPH oxidase activity by acting as a redox-dependent enzyme complex organizer.
Assuntos
Membrana Celular/enzimologia , Citosol/enzimologia , NADPH Oxidases/metabolismo , Neutrófilos/enzimologia , Isomerases de Dissulfetos de Proteínas/metabolismo , Superóxidos/metabolismo , Substituição de Aminoácidos , Antibacterianos/farmacologia , Bacitracina/farmacologia , Membrana Celular/genética , Ativação Enzimática/efeitos dos fármacos , Ativação Enzimática/fisiologia , Inibidores Enzimáticos/farmacologia , Humanos , Mutação de Sentido Incorreto , NADPH Oxidases/genética , Oxirredução/efeitos dos fármacos , Isomerases de Dissulfetos de Proteínas/genética , Transporte Proteico/efeitos dos fármacos , Transporte Proteico/fisiologiaRESUMO
Bacitracin and the membrane-impermeant thiol reagent 5,5'-dithiobis-2-nitrobenzoic acid (DTNB) are agents known to inhibit protein disulfide isomerase (PDI), a cell-surface protein critical in HIV-1 entry therefore they are fusion inhibitors (FI). Here we investigated the possibility that Bacitracin and or DTNB might have other antiviral activities besides FI. By means of residual activity assays, we found that both compounds showed antiviral activity only to viruses T-tropic HIV-1 strain. Cell-based fusion assays showed inhibition on HeLa-CD4-LTR-ß-gal (CD4) and HL2/3 cells treated with Bacitracin, and DTNB with the latest compound we observed fusion inhibition on both cells but strikingly in HL2/3 cells (expressing Env) indicating a possible activity on both, the cell membrane and the viral envelope. A time-of-addition experiment showed that both compounds act on HIV entry inhibition but DTNB also acts at late stages of the viral cycle. Lastly, we also found evidence of long-lasting host cell protection in vitro by DTNB, an important pharmacodynamic parameter for a topical microbicide against virus infection, hours after the extracellular drug was removed; this protection was not rendered by Bacitracin. These drugs proved to be leading compounds for further studies against HIV showing antiviral characteristics of interest.
Assuntos
Fármacos Anti-HIV/farmacologia , Bacitracina/farmacologia , Ácido Ditionitrobenzoico/farmacologia , Infecções por HIV/tratamento farmacológico , HIV-1/efeitos dos fármacos , Tropismo Viral , Linhagem Celular Tumoral , Infecções por HIV/virologia , HIV-1/fisiologia , Humanos , Linfócitos T/virologia , Tropismo Viral/efeitos dos fármacos , Internalização do Vírus/efeitos dos fármacosRESUMO
Protein disulfide isomerase (PDI) enzymes are eukaryotic oxidoreductases that catalyze oxidation, reduction and isomerization of disulfide bonds in polypeptide substrates. Here, we report the biochemical characterization of a PDI enzyme from the protozoan parasite Entamoeba histolytica (EhPDI). Our results show that EhPDI behaves mainly as an oxidase/isomerase and can be inhibited by bacitracin, a known PDI inhibitor; moreover, it exhibits chaperone-like activity. Albeit its physiological role in the life style of the parasite (including virulence and survival) remains to be studied, EhPDI could represent a potential drug target for anti-amebic therapy.
Assuntos
Entamoeba histolytica/enzimologia , Isomerases de Dissulfetos de Proteínas/metabolismo , Antibacterianos/farmacologia , Bacitracina/farmacologia , Entamoeba histolytica/efeitos dos fármacos , Concentração de Íons de Hidrogênio , Concentração Inibidora 50 , Insulina/metabolismo , Chaperonas Moleculares/metabolismo , Muramidase/química , Muramidase/metabolismo , Oxirredutases/metabolismo , Isomerases de Dissulfetos de Proteínas/antagonistas & inibidores , Isomerases de Dissulfetos de Proteínas/química , Dobramento de Proteína , Ribonuclease Pancreático/química , Ribonuclease Pancreático/metabolismoRESUMO
The study was focused on the role of the penicillin binding protein PBP4* of Bacillus subtilis during growth in high salinity rich media. Using pbpE-lacZ fusion, we found that transcription of the pbpE gene is induced in stationary phase and by increased salinity. This increase was also corroborated at the translation level for PBP4* by western blot. Furthermore, we showed that a strain harboring gene disruption in the structural gene (pbpE) for the PBP4* endopeptidase resulted in a salt-sensitive phenotype and increased sensitivity to cell envelope active antibiotics (vancomycin, penicillin and bacitracin). Since the pbpE gene seems to be part of a two-gene operon with racX, a racX::pRV300 mutant was obtained. This mutant behaved like the wild-type strain with respect to high salt. Electron microscopy showed that high salt and mutation of pbpE resulted in cell wall defects. Whole cells or purified peptidoglycan from WT cultures grown in high salt medium showed increased autolysis and susceptibility to mutanolysin. We demonstrate through zymogram analysis that PBP4* has murein hydrolyze activity. All these results support the hypothesis that peptidoglycan is modified in response to high salt and that PBP4* contributes to this modification.
Assuntos
Bacillus subtilis/metabolismo , N-Acetil-Muramil-L-Alanina Amidase/fisiologia , Proteínas de Ligação às Penicilinas/fisiologia , Salinidade , D-Ala-D-Ala Carboxipeptidase Tipo Serina/fisiologia , Antibacterianos/farmacologia , Bacillus subtilis/crescimento & desenvolvimento , Bacillus subtilis/ultraestrutura , Bacitracina/farmacologia , Bacteriólise , Parede Celular/efeitos dos fármacos , Parede Celular/ultraestrutura , Microscopia Eletrônica de Transmissão , N-Acetil-Muramil-L-Alanina Amidase/deficiência , Penicilina G/farmacologia , Proteínas de Ligação às Penicilinas/deficiência , Peptidoglicano/metabolismo , D-Ala-D-Ala Carboxipeptidase Tipo Serina/deficiência , Transcrição Gênica , Vancomicina/farmacologiaRESUMO
A survey of the changes in populations of heterotrophic bacteria, coliform microorganisms and S. meliloti was conducted in samples taken from the water irrigation channels of the Neuquén River (Argentina). Fifty-six water samples were collected during the spring-summer seasons of 1997-1999 years. Both the heterotrophic plate count bacterial and the number of coliforms oscillated between 110-5050 CFU/ml and 8-1400 CFU/100 ml, respectively, during the period this study was carried out. Fecal coliforms were detected in 91.1% of the water samples investigated. Moreover, the results showed that S. meliloti capable of nodulating alfalfa (Medicago sativa L.) Cuf 101 were present in 68% of the water samples and in effectiveness studies, no isolate out of 25 evaluated could be classified as superior N fixers. That is, they did not produce plants equal in weight to nitrate-grown plants (KNO3 0.05%). All the S. meliloti strains were resistant to novobiocin and bacitracin, while 72% of the microsymbionts demonstrated resistance to between seven and ten antibiotics. Results presented in this study showed that irrigation waters of the Neuquén river could act as dispersal agents of both ineffective S. meliloti strains and thermotolerant coliform bacteria.