RESUMO
AIMS: This study investigated the occurrence and genetic diversity of Enterobacteriaceae with extended-spectrum ß-lactamase (ESBL)-, AmpC- and carbapenemase-mediated resistance in British beef cattle, and related risk factors. METHODS AND RESULTS: Faecal samples (n = 776) were obtained from farms in England and Wales (n = 20) and Scotland (n = 20) in 2015. Isolates from selective agars were identified by MALDI ToF mass spectrometry. Selected isolates were characterized by multiplex PCR (blaCTX -M, blaOXA , blaSHV and blaTEM genes), whole-genome sequencing (WGS), minimum inhibitory concentrations and pulsed-field gel electrophoresis. None of the faecal samples yielded carbapenem-resistant Escherichia coli. Ten (25%) of the farms tested positive for ESBL-producing CTX-M Enterobacteriaceae, 15 (37·5%) of the farms were positive for AmpC phenotype E. coli and none were positive for carbapenem-resistant E. coli. WGS showed a total of 30 different resistance genes associated with E. coli, Citrobacter and Serratia from ESBL agars, and colocation of resistance genes with blaCTX -M1 . Buying bulls and bringing in fattening cattle from another farm were identified as significant risk factors for positive samples harbouring CTX-M Enterobacteriaceae or AmpC phenotype E. coli respectively. CONCLUSIONS: Beef cattle on a proportion of farms in GB carry ESBL-producing Enterobacteriaceae. Factors, such as operating as a closed herd, may have an important role in reducing introduction and transmission of resistant Enterobacteriaceae. The results indicate management factors may play an important role in impacting ESBL prevalence. In particular, further study would be valuable to understand the impact of maintaining a closed herd on reducing the introduction of resistant Enterobacteriaceae. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first study showing the presence of ESBL-producing Enterobacteriaceae in British beef cattle.
Assuntos
Farmacorresistência Bacteriana , Enterobacteriaceae/efeitos dos fármacos , Enterobacteriaceae/isolamento & purificação , Carne Vermelha/microbiologia , beta-Lactamas/farmacologia , Animais , Antibacterianos/farmacologia , Bovinos , Farmacorresistência Bacteriana/genética , Enterobacteriaceae/genética , Enterobacteriaceae/metabolismo , Fazendas/estatística & dados numéricos , Fezes/microbiologia , Microbiologia de Alimentos , Genes Bacterianos/genética , Reino Unido , beta-Lactamases/genéticaRESUMO
AIMS: In 2015, colistin-resistant Escherichia coli and Salmonella with the mcr-1 gene were isolated from a pig farm in Great Britain. Pigs were subsequently monitored over a ~20-month period for the occurrence of mcr-1-mediated colistin resistance and the risk of mcr-1 E. coli entering the food chain was assessed. METHODS AND RESULTS: Pig faeces and slurry were cultured for colistin-resistant E. coli and Salmonella, tested for the mcr-1 gene by PCR and selected isolates were further analysed. Seventy-eight per cent of faecal samples (n = 275) from pigs yielded mcr-1 E. coli after selective culture, but in positive samples only 0·2-1·3% of the total E. coli carried mcr-1. Twenty months after the initial sampling, faecal samples (n = 59) were negative for E. coli carrying mcr-1. CONCLUSIONS: The risk to public health from porcine E. coli carrying mcr-1 was assessed as very low. Twenty months after cessation of colistin use, E. coli carrying mcr-1 was not detected in pig faeces on a farm where it was previously present. SIGNIFICANCE AND IMPACT OF THE STUDY: The results suggest that cessation of colistin use may help over time to reduce or possibly eliminate mcr-1 E. coli on pig farms where it occurs.
Assuntos
Antibacterianos , Colistina , Farmacorresistência Bacteriana , Infecções por Escherichia coli , Proteínas de Escherichia coli/genética , Escherichia coli , Animais , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Colistina/farmacologia , Colistina/uso terapêutico , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , Infecções por Escherichia coli/tratamento farmacológico , Infecções por Escherichia coli/epidemiologia , Infecções por Escherichia coli/microbiologia , Infecções por Escherichia coli/veterinária , Fezes/microbiologia , Estudos Longitudinais , SuínosRESUMO
The aim of this study was to investigate the bacterial strains and farm environment that may contribute to the persistence of ESBL-producing E. coli on a single UK dairy farm. A longitudinal study was conducted comprising 6 visits, between August and October 2010, followed by a further visit at approximately 69weeks after the initial visit. Faecal and environmental samples were collected from different parts of the farm. The persistence and extent of faecal shedding of ESBL E. coli by individual calves was also determined. Twenty two different PFGE types were identified. Four of these were persistent during the study period and were associated with serotypes: O98, O55, O141 and O33. The counts suggest that shedding in calf faeces was an important factor for the persistence of strains, and the data will be useful for parameterising mathematical models of the spread and persistence of ESBL strains within a dairy farm.
Assuntos
Doenças dos Bovinos/epidemiologia , Infecções por Escherichia coli/veterinária , Escherichia coli/fisiologia , Animais , Derrame de Bactérias , Bovinos , Doenças dos Bovinos/genética , Doenças dos Bovinos/microbiologia , Indústria de Laticínios , Meio Ambiente , Escherichia coli/genética , Infecções por Escherichia coli/epidemiologia , Infecções por Escherichia coli/genética , Infecções por Escherichia coli/microbiologia , Proteínas de Escherichia coli/análise , Fazendas , Fezes/microbiologia , Estudos Longitudinais , Modelos Teóricos , Prevalência , Reino Unido/epidemiologia , beta-Lactamases/análiseRESUMO
The aim of this study was to develop a PCR test to detect chromosomal differences between epidemic multidrug resistant (epi-MDR) strains of Salmonella enterica serovar Newport (S. Newport) and non-epi-MDR strains of S. Newport that are endemic to the United Kingdom (UK). Sequence analysis of the biofilm-associated protein A gene (bapA) showed that epi-MDR strains of S. Newport from the United States of America (USA) had a deletion of 309 bp, which was not present in non-epi-MDR strains of S. Newport from the UK. A PCR test was developed using primers designed to target this difference and was applied to a panel of S. Newport isolates comprising of strains from the UK (n=20, non-epi-MDR), from the USA (n=10, epi-MDR) and from Canada (n=7). A second panel of isolates (n=73) was used to assess the test specificity, and these isolates consisted of non-Newport Salmonella serovars (n=25), and other epidemic serovars (n=48). Epi-MDR S. Newport isolates produced a characteristic 505 bp amplicon, whereas non-epi-MDR S. Newport isolates produced an 814 bp amplicon. The bapA PCR has potential to discriminate between these S. Newport strains irrespective of their carrying resistance genes.
Assuntos
Farmacorresistência Bacteriana Múltipla , Reação em Cadeia da Polimerase/veterinária , Salmonella enterica/genética , Antibacterianos/farmacologia , Canadá , Reação em Cadeia da Polimerase/métodos , Salmonella enterica/efeitos dos fármacos , Análise de Sequência de DNA , Reino Unido , Estados UnidosRESUMO
Antimicrobial resistance (AMR) threatens the effective prevention and treatment of bacterial diseases in both humans and animals. Globally, there has been much research done regarding resistant bacteria in the livestock industry, but few published resources collate this information. This report discusses a risk assessment (RA) framework and subsequent analysis of data availability for AMR in bacteria from 4 livestock sectors: dairy cattle, beef cattle, pigs and poultry, with particular reference to ESBL-producing Escherichia coli (ESBL E. coli) prevalence in the dairy cattle sector within the United Kingdom. The aim of this assessment was to identify where quality data exist, for the purpose of parameterising a quantitative RA, and where it would be useful to direct future research to provide quality data to improve the current knowledge base. Such research is necessary to support risk modelling and forecasting capability regarding the relative contributions of factors that maintain the emergence and spread of AMR in bacteria. The review suggested that there are data gaps regarding ESBL E. coli occurrence in the following: beef cattle, bulk tank milk and dairy products, animal-by-products, the farm environment (including after flooding) as well as the effect of animal stress on shedding levels. Filling these data gaps prior to undertaking a full quantitative RA would make the assessment more robust and give greater confidence in the final outcome and consequently inform the targeting and prioritising of interventions to minimise spread of AMR in bacteria in farm animals.
Assuntos
Infecções por Escherichia coli/transmissão , Gado/microbiologia , Zoonoses/transmissão , Resistência beta-Lactâmica , Animais , Indústria de Laticínios , Escherichia coli , Infecções por Escherichia coli/epidemiologia , Infecções por Escherichia coli/microbiologia , Humanos , Medição de Risco , Reino Unido/epidemiologia , Zoonoses/epidemiologia , Zoonoses/microbiologiaRESUMO
AIMS: The aims of this work were to develop a model of dairy farm waste milk and to investigate methods for the bioremediation of milk containing cefquinome residues. METHODS AND RESULTS: Unpasteurized milk and UHT milk that had both been spiked with cefquinome at a concentration of 2 µg ml(-1) were used as a model for waste milk containing cephalosporin residues. Adjustment of the spiked UHT milk to pH 10 or treatment with conditioned medium from bacterial growth producing cefotaximase, were the most effective methods for decreasing the cefquinome concentrations within 24 h. A large-scale experiment (10 l of cefquinome-spiked unpasteurized milk) suggested that fermentation for 22 h at 37°C followed by heating at 60°C for 2 h was sufficient to decrease cefquinome concentrations to below the limit of quantification (<125 µg kg(-1) ) and to kill the majority of the enriched bacterial population. CONCLUSIONS: One or a combination of the bioremediation methods described may have potential as a practical treatment for dairy farm waste milk. SIGNIFICANCE AND IMPACT OF THE STUDY: Treatment of waste milk to decrease cephalosporin residue concentrations and also to kill bacteria prior to feeding to dairy calves could decrease the risk of selection for ESBL bacteria on dairy farms.
Assuntos
Antibacterianos/metabolismo , Cefalosporinas/metabolismo , Indústria de Laticínios , Leite/química , Resíduos , Animais , Biodegradação Ambiental , Bovinos , Escherichia coli/crescimento & desenvolvimento , Fermentação , Temperatura Alta , Modelos Biológicos , beta-Lactamases/metabolismoRESUMO
1. A field study was performed to investigate the presence and characteristics of ciprofloxacin-resistant, extended spectrum ß-lactamase (ESBL) and AmpC Escherichia coli from turkeys in Great Britain. E. coli were isolated from ~9000 boot swab samples from 27 different farms owned by four different companies. Between 1 and 14 visits were made to each farm (mean 3) at between 0 and 15 m intervals (mean ~5 m). 2. CHROMagar ECC with and without ciprofloxacin or cephalosporin antibiotics was used as selective isolation media. Representative isolates with different phenotypes were tested for mutations in gyrA and for: qnrA, B, S, qepA and aac(6')-Ib genes, for ESBL phenotype, the presence of bla genes and plasmid type, and for ampC genes Representative ciprofloxacin-resistant and CTX-M isolates were further tested for serotype and PFGE type. On ciprofloxacin selective media 55% of samples yielded ciprofloxacin resistant E. coli and of those further analysed, most had ciprofloxacin MICs >4 mg/l and mutations in gyrA. 3. For the different companies, the mean number of samples per farm with cefoxitin- or cefotaxime-resistant isolates ranged from 1·0% to 61·9% and 4·7% to 31·7% respectively. Cefotaxime-resistance was most commonly associated with an ESBL phenotype, a CTX-M-1 or CTX-M-14 sequence type and an I1-γ or K plasmid inc type. The mechanism of cefoxitin resistance was not determined for most isolates, but where determined it was bla . 4. PFGE and serotyping showed clonally-related isolates persisting over multiple visits suggesting both more prudent use of antibiotics and improved farm hygiene are needed to address the issue of antimicrobial resistance in isolates from turkeys.
Assuntos
Cefalosporinas/farmacologia , Ciprofloxacina/farmacologia , Farmacorresistência Bacteriana Múltipla/genética , Escherichia coli/efeitos dos fármacos , Perus/microbiologia , Animais , Escherichia coli/isolamento & purificação , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/genética , Genótipo , Testes de Sensibilidade Microbiana , Fenótipo , Sorotipagem , Reino Unido/epidemiologia , beta-Lactamases/química , beta-Lactamases/genéticaRESUMO
The objective of this study was to investigate the hypothesis that some sporadic Salmonella infections in domesticated animals may be associated with Salmonella infections originating from garden birds. Phage type and antimicrobial resistance details of isolates of S. Typhimurium obtained from wild birds were comparable with those from S. Typhimurium infections from domesticated animals or livestock between 2002 and 2010. A small panel of S. Typhimurium isolates (n=37) were characterised by multilocus variable number of tandem repeats analysis (MLVA), pulsed field gel electrophoresis (PFGE) and phage type. The MLVA-PFGE data clustered the strains according to phage type (DT40 or DT56). Within each group there were strains from wild birds and domesticated animals or livestock with MLVA profiles having up to 100% similarity. The results from this study therefore lend support to the hypothesis that Salmonella infections in domesticated animals could be caused by infections carried by wild birds.
Assuntos
Aves/microbiologia , Gado/microbiologia , Salmonelose Animal/microbiologia , Salmonella typhimurium/isolamento & purificação , Animais , Animais Selvagens , Tipagem de Bacteriófagos/veterinária , Análise por Conglomerados , Eletroforese em Gel de Campo Pulsado/veterinária , Salmonelose Animal/genética , Salmonella typhimurium/classificação , Salmonella typhimurium/genética , Sequências de Repetição em TandemRESUMO
Fluoroquinolones are a widely used group of antimicrobials in both human and animal medicine, with ciprofloxacin being a critically important fluoroquinolone for serious human infections. The present study reports on a 1-year survey for the presence of ciprofloxacin-resistant Escherichia coli in turkeys from Great Britain. Boot swabs were taken from 442 turkey flocks comprised of 125 breeding flocks and 317 meat flocks from 337 different farms over a 1-year period (2006 to 2007). CHROMagar ECC containing 1 mg/l ciprofloxacin was used to obtain ciprofloxacin-resistant isolates. Isolates were tested for sensitivity to 16 different antimicrobials. Isolates with ciprofloxacin minimum inhibitory concentrations ≥8 mg/l were tested for mutations in gyrA by polymerase chain reaction and DNA sequencing. Selected isolates were tested by multiplex polymerase chain reaction for qnrA, qnrB and qnrS, qepA and aac(6')-Ib-cr genes. Conjugations were performed to assess the transferability of resistance to quinolones. Ciprofloxacin-resistant E. coli was found in 22.4% of turkey breeding flocks and 60.9% of meat flocks. Two main mutations in gyrA, as well as a range of silent mutations, were identified in resistant isolates. Flocks with transferable resistance genes qnrB, qnrS, and aac(6')-Ib-cr were found at a low flock prevalence of 4.2%, 1.6% and 1.0%, respectively; however, under laboratory conditions only transfer of qnrS genes could be demonstrated. This work has confirmed the occurrence of ciprofloxacin-resistant E. coli strains throughout turkey breeding and meat flocks, with almost one-third of E. coli isolates being resistant to ciprofloxacin. Of those, more than 87% were also resistant to three or more antimicrobial classes.
Assuntos
Ciprofloxacina , Farmacorresistência Bacteriana/genética , Infecções por Escherichia coli/veterinária , Escherichia coli/genética , Doenças das Aves Domésticas/epidemiologia , Doenças das Aves Domésticas/microbiologia , Perus , Animais , DNA Girase/genética , Análise Mutacional de DNA/veterinária , Infecções por Escherichia coli/epidemiologia , Infecções por Escherichia coli/microbiologia , Testes de Sensibilidade Microbiana/veterinária , Reação em Cadeia da Polimerase Multiplex/veterinária , Polimorfismo de Fragmento de Restrição , Prevalência , Reino Unido/epidemiologiaAssuntos
Aborto Animal/epidemiologia , Aborto Animal/microbiologia , Doenças dos Bovinos/epidemiologia , Complicações Infecciosas na Gravidez/veterinária , Salmonelose Animal/complicações , Animais , Bovinos , Indústria de Laticínios , Feminino , Filogenia , Gravidez , Complicações Infecciosas na Gravidez/epidemiologia , Salmonella/classificação , Salmonella/isolamento & purificação , Salmonelose Animal/epidemiologia , Reino Unido/epidemiologiaRESUMO
The number and proportion of CTX-M positive Escherichia coli organisms were determined in feces from cattle, chickens, and pigs in the United Kingdom to provide a better understanding of the risk of the dissemination of extended-spectrum ß-lactamase (ESBL) bacteria to humans from food animal sources. Samples of bovine (n = 35) and swine (n = 20) feces were collected from farms, and chicken cecal contents (n = 32) were collected from abattoirs. There was wide variation in the number of CTX-M-positive E. coli organisms detected; the median (range) CFU/g were 100 (100 × 10(6) to 1 × 10(6)), 5,350 (100 × 10(6) to 3.1 × 10(6)), and 2,800 (100 × 10(5) to 4.7 × 10(5)) for cattle, chickens, and pigs, respectively. The percentages of E. coli isolates that were CTX-M positive also varied widely; median (range) values were 0.013% (0.001 to 1%) for cattle, 0.0197% (0.00001 to 28.18%) for chickens, and 0.121% (0.0002 to 5.88%) for pigs. The proportion of animals designated high-density shedders (≥1 × 10(4) CFU/g) of CTX-M E. coli was 3/35, 15/32, and 8/20 for cattle, chickens, and pigs, respectively. We postulate that high levels of CTX-M E. coli in feces facilitate the dissemination of bla(CTX-M) genes during the rearing of animals for food, and that the absolute numbers of CTX-M bacteria should be given greater consideration in epidemiological studies when assessing the risks of food-borne transmission.
Assuntos
Derrame de Bactérias , Escherichia coli/enzimologia , Escherichia coli/isolamento & purificação , Fezes/microbiologia , beta-Lactamases/biossíntese , Animais , Carga Bacteriana , Bovinos , Galinhas , Microbiologia Ambiental , Microbiologia de Alimentos , Humanos , Suínos , Reino UnidoRESUMO
OBJECTIVES: to determine the prevalence of extended-spectrum ß-lactamases (ESBLs) in Escherichia coli from poultry in Great Britain (GB). METHODS: E. coli was isolated from 388 broiler chicken caecal samples from 22 abattoirs and from boot swabs from 442 turkey flocks over successive 1 year periods. CHROMagar ECC with and without cephalosporin antibiotics was used as isolation medium and the chicken study also used CHROMagar CTX. ESBL phenotype isolates were tested for the presence of bla(CTX-M,) bla(OXA), bla(SHV), bla(TEM) and ampC genes(.) CTX-M isolates were tested for O25 serogroup, replicon, CTX-M sequence, multilocus sequence type (MLST), PFGE type, plasmid transfer and qnrA, qnrB, qnrS, qepA and aac(6')-Ib genes. RESULTS: CTX-M-carrying E. coli were isolated from 54.5% of the broiler abattoirs and from 3.6% of individual broiler caecal samples and were CTX-M sequence types 1 (mainly), 3 and 15 with replicon types I1-γ, A/C and P/F, and I1-γ, respectively. CTX-M-carrying E. coli were isolated from 5.2% of turkey meat production farms and 6.9% of turkey breeder farms and were CTX-M sequence types 1, 14 (mainly), 15 and 55 with mainly replicon types F, FIA, K and I1-γ, respectively. None of the CTX-M isolates was serogroup O25. PFGE/MLST showed the CTX-M isolates to be clonally diverse, although MLST 156 with CTX-M-15 was isolated from both chickens and turkeys and has been previously reported in gulls. CTX-M-negative, ESBL- and bla(TEM)-positive strains were mainly TEM-52C. CONCLUSIONS: poultry-derived CTX-M E. coli in GB are different from major CTX-M sequence types causing disease in humans.
Assuntos
Ceco/microbiologia , Galinhas/microbiologia , Infecções por Escherichia coli/veterinária , Escherichia coli/enzimologia , Perus/microbiologia , beta-Lactamases/biossíntese , Animais , Técnicas de Tipagem Bacteriana , DNA Bacteriano/genética , Eletroforese em Gel de Campo Pulsado , Escherichia coli/isolamento & purificação , Infecções por Escherichia coli/epidemiologia , Infecções por Escherichia coli/microbiologia , Genes Bacterianos , Testes de Sensibilidade Microbiana/métodos , Epidemiologia Molecular , Tipagem Molecular , Tipagem de Sequências Multilocus , Plasmídeos/análise , Prevalência , Reino Unido/epidemiologiaRESUMO
During the clinical phase of bovine spongiform encephalopathy (BSE), a significant decrease was observed in the ratio of muscle glycogen to plasma L-lactic acid concentrations in BSE infected field case and experimentally infected dairy cattle compared with healthy control cattle (P<0.001), this being due to changes in the concentration of both metabolites in the BSE infected cattle compared with the control group. Furthermore, the concentration of plasma alanine was significantly increased (P<0.05) in the infected animals. No significant difference was detected between these two groups in the ratio of hepatic glycogen to plasma lactate. We infer that BSE infected cattle exhibit signs of altered energy metabolism and when applied in conjunction with changes in other metabolite biomarkers these changes may be useful for discriminating BSE infected cattle from healthy cattle or those suffering with other disorders or diseases.
Assuntos
Encefalopatia Espongiforme Bovina/sangue , Glicogênio/metabolismo , Músculo Esquelético/metabolismo , Alanina/sangue , Animais , Bovinos , Indústria de Laticínios , Feminino , Ácido Láctico/sangue , Fígado/metabolismo , MasculinoRESUMO
The alkyl esters of p-hydroxybenzoic acid (parabens) are used widely as preservatives in foods, pharmaceuticals and cosmetics to which the human population is exposed. Recent studies have reported that methylparaben, ethylparaben, n-propylparaben and n-butylparaben all possess oestrogenic activity in several in vitro assays and in animal models in vivo. This study reports on the oestrogenic activity of isobutylparaben in a panel of assays in vitro and in vivo. Isobutylparaben was able to displace [(3)H]oestradiol from cytosolic oestrogen receptor alpha of MCF7 human breast cancer cells by 81% at 100 000-fold molar excess. Using a clonal line of MCF7 cells containing a stably transfected oestrogen-responsive ERE-CAT reporter gene, CAT gene expression could be increased by isobutylparaben such that the magnitude of the response was the same at 10(-5) M isobutylparaben as with 10(-8) M 17beta-oestradiol. Isobutylparaben could also increase expression of the endogenous oestrogen-responsive pS2 gene in MCF7 cells and maximal expression at 10(-5) M isobutylparaben could be inhibited with the anti-oestrogen ICI 182 780. The proliferation of two oestrogen-dependent human breast cancer cell lines MCF7 and ZR-75-1 could be increased with isobutylparaben such that at concentrations of 10(-5) M the proliferation response was of the same magnitude as with 10(-8) M 17beta-oestradiol. Evidence for oestrogen receptor mediation of proliferation effects was provided by the inability of isobutylparaben to influence the growth of oestrogen-unresponsive MDA-MB-231 human breast cancer cells and by the ability of the anti-oestrogen ICI 182 780 to inhibit the isobutylparaben effects on MCF7 cell growth. The proliferation response to 10(-10) M 17beta-oestradiol was not antagonized with isobutylparaben at any concentration from 10(-9) M to 10(-4) M in either MCF7 or ZR-75-1 cells. Finally, subcutaneous administration of isobutylparaben was able to increase the uterine weight in the immature mouse after three daily doses of 1.2 or 12.0 mg per mouse. Previous work using linear-alkyl-chain parabens has shown that oestrogenic activity increases with alkyl chain length from methylparaben to n-butylparaben. The results here show that branching of the alkyl chain to isobutylparaben increases oestrogenic activity beyond that of the equivalent length linear alkyl chain in n-butylparaben.
Assuntos
Estradiol/análogos & derivados , Moduladores de Receptor Estrogênico , Estrogênios não Esteroides/toxicidade , Parabenos/toxicidade , Animais , Neoplasias da Mama/metabolismo , Divisão Celular , Relação Dose-Resposta a Droga , Estradiol/farmacologia , Receptor alfa de Estrogênio , Estrogênios não Esteroides/antagonistas & inibidores , Feminino , Fulvestranto , Regulação Neoplásica da Expressão Gênica , Humanos , Ligantes , Camundongos , Camundongos Endogâmicos , Tamanho do Órgão/efeitos dos fármacos , Ligação Proteica , Proteínas/genética , RNA Mensageiro/genética , Receptores de Estrogênio/metabolismo , Fator Trefoil-1 , Células Tumorais Cultivadas , Proteínas Supressoras de Tumor , Útero/efeitos dos fármacos , Útero/patologiaRESUMO
gamma-Glutamyltranspeptidase (GGT)-deficient mice (GGT(-/-)) display chronic glutathione (GSH) deficiency, growth retardation, and die at a young age (<20 weeks). Using livers from these mice, we investigated the relationship between GSH content, especially mitochondrial, and mitochondrial and cellular function. We found that the GSH content of isolated liver mitochondria was diminished by >/=50% in GGT(-/-) mice when compared with wild-type mice. Respiratory control ratios (RCRs) of GGT(-/-) mice liver mitochondria were =60% those of wild-type mice primarily as a result of impaired state 3 respiration. Mitochondrial adenine nucleotide content was decreased by >/=40% in mitochondria obtained from GGT(-/-) mice. We observed a strong correlation between mitochondrial GSH content and RCRs. Even moderate decreases (<50%) correlated with adverse effects with respect to respiration. Electron microscopy revealed that livers from GGT(-/-) knockout mice were deprived of fat and glycogen, and swollen mitochondria were observed in animals that were severely deprived of GSH. Thus, GGT(-/-) mice exhibit a loss of GSH homeostasis and impaired oxidative phosphorylation, which may be related to the rate of adenosine triphosphate (ATP) formation and subsequently leads to progressive liver injury, which characterizes the diseased state. We also found that supplementation of GGT(-/-) mice with N-acetylcysteine (NAC) partially restored liver GSH, but fully restored mitochondrial GSH and respiratory function. Electron microscopy revealed that the livers of NAC-supplemented GGT(-/-) mice contained fat and glycogen; however, slightly enlarged mitochondria were found in some livers. NAC supplementation did not have any beneficial effect on the parameters examined in wild-type mice.
Assuntos
Glutationa/metabolismo , Mitocôndrias Hepáticas/fisiologia , gama-Glutamiltransferase/deficiência , Difosfato de Adenosina/análise , Trifosfato de Adenosina/análise , Trifosfato de Adenosina/biossíntese , Animais , Glutationa/análise , Fígado/citologia , Fígado/ultraestrutura , Camundongos , Camundongos Knockout , Microscopia Eletrônica , Consumo de OxigênioRESUMO
OBJECTIVE: The purpose of this study was to test the hypothesis that the number of projections influences the accuracy of a simple depth discrimination task when tuned-aperture computed tomography is used. STUDY DESIGN: In each of 4 partially edentulous mandibles, 2 radiopaque steel spheres were attached to the facial and lingual surfaces and 1 ceramic sphere was place in the apical region of an open tooth socket. Errors in estimates of the depth of the apically positioned ceramic sphere relative to the 2 steel spheres were determined from 3-dimensional tuned-aperture computed tomography reconstructions. These data were compared with actual measurements produced independently by means of an optical micrometer. Multiple projections were produced from radially symmetric exposures bearing an angular disparity of 15 degrees. The number of symmetrically dispersed projections per tuned-aperture computed tomography reconstruction was varied systematically (2, 4, 8, 12, and 16 projections). The consequences of this variable, as well as specimen and observer effects, were evaluated in a balanced factorial experimental design. Depth estimates were performed by 10 trained observers. The depth reported was that corresponding to the tuned-aperture computed tomography slice perceived to yield the image of the ceramic sphere in sharpest focus. Resulting data were normalized by logarithmic transformation and analyzed statistically by analysis of variance. RESULTS: No statistically meaningful effects were found for the number of projections (P = .607) or for different observers (P = .093), but a significant specimen effect was demonstrated (P = .006). CONCLUSIONS: Factors other than high contrast limit the perception of image sharpness under these conditions. Depth may be estimated accurately from relatively small numbers of projections.
Assuntos
Intensificação de Imagem Radiográfica/métodos , Radiografia Dentária/métodos , Tomografia Computadorizada por Raios X/métodos , Alvéolo Dental/diagnóstico por imagem , Análise de Variância , Artefatos , Cerâmica , Meios de Contraste , Percepção de Profundidade , Humanos , Arcada Parcialmente Edêntula/diagnóstico por imagem , Reprodutibilidade dos Testes , AçoRESUMO
OBJECTIVE: The purpose of this study was to determine the degree to which the number and angular disparity of component projections influence depth discrimination with tuned-aperture computed tomography. STUDY DESIGN: Groups of three tiny steel spheres served as fiducial references on and in four partially edentulous mandibles. Two spheres were attached to the facial and lingual surfaces of each mandible, and the third was fixed in the apical region of an open tooth socket. Errors in estimates of the depth of the apically positioned sphere relative to the other two spheres were determined from three-dimensional tuned-aperture computed tomography reconstructions. These data were compared with actual measurements produced independently with an optical micrometer. Multiple projections required by the tuned-aperture computed tomography reconstruction algorithm were produced from radially symmetric exposures bearing angular disparities of 5, 15, 30, and 45 degrees. The number of symmetrically dispersed projections per tuned-aperture computed tomography reconstruction likewise was varied systematically (2, 4, 8, 12, and 16 projections). These variables were manipulated through the use of a balanced factorial design. Depth estimates were performed by trained observers; the estimates were based on the determination of tuned-aperture computed tomography slices perceived as imaging the respective apical spheres in sharpest focus. Specimen and observer effects were also considered as independent variables. Resulting data were normalized by logarithmic transformation and analyzed statistically by analysis of variance. RESULTS: Significant differences (p < 0.005) were demonstrated for angular disparity and specimen effects, but the number of projections and the effect of the observer were not found to be statistically significant. CONCLUSIONS: In dentistry, angular disparities of 15 degrees or greater should be used when tuned-aperture computed tomography is being applied to diagnostic tasks requiring maximal depth discrimination accuracy.
Assuntos
Percepção de Profundidade , Processamento de Imagem Assistida por Computador/métodos , Radiografia Dentária/métodos , Tomografia Computadorizada por Raios X/métodos , Algoritmos , Processo Alveolar/diagnóstico por imagem , Análise de Variância , Humanos , Arcada Parcialmente Edêntula/diagnóstico por imagem , Mandíbula/diagnóstico por imagem , Variações Dependentes do Observador , Próteses e Implantes , AçoRESUMO
The mechanism by which alpha-tocopherol (alpha-T) is secreted into the bile is not known; however, we have previously demonstrated that treatment with piperonyl butoxide (PIP, 1 g/kg) results in increased biliary output of both alpha-T and phosphatidylcholine within 3 h of ip injection in rats and that the biliary output of both substances was prevented by chemicals that disrupt microtubules (Toxicol. Appl. Pharmacol. 139, 411-417 (1996)). The P-glycoprotein (Pgp) encoded by the mdr2 gene has been shown to transport phosphatidylcholine into the bile; therefore, in the current study, we utilized the Pgp inhibitor verapamil to investigate the possible involvement of Pgps in the biliary secretion of alpha-T. When rats were iv injected with verapamil (4 mg/kg) 10 min prior to PIP treatment, verapamil prevented the PIP-induced increases in biliary alpha-T and phosphatidylcholine output and resulted in biliary alpha-T outputs that were significantly less than controls. Also, we determined that the biliary alpha-T levels in mdr2 knockout mice were 25% of those in wildtype mice; furthermore, mdr2 liver, lung, and kidney levels of alpha-T and glutathione differed from those of wildtype. To investigate the fate of biliary alpha-T, we injected 14C-labeled alpha-T into the bile duct cannulae of rats and determined that approximately 60% of the radioactivity was reabsorbed within 1 h. Our results indicate that alpha-T undergoes enterohepatic circulation and that the biliary secretion of alpha-T, basally and following chemical treatment, is dependent on the presence of a functioning mdr2 Pgp in rats and mice.
Assuntos
Membro 1 da Subfamília B de Cassetes de Ligação de ATP/fisiologia , Subfamília B de Transportador de Cassetes de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/fisiologia , Bile/química , Vitamina E/metabolismo , Animais , Ácidos e Sais Biliares/análise , Radioisótopos de Carbono/metabolismo , Feminino , Glutationa/análise , Dissulfeto de Glutationa/análise , Fígado/metabolismo , Masculino , Camundongos , Camundongos Knockout , Fosfatidilcolinas/metabolismo , Butóxido de Piperonila/farmacologia , Ratos , Ratos Sprague-Dawley , Verapamil/farmacologia , Vitamina E/análiseRESUMO
OBJECTIVE: Tuned-aperture computed tomography, a new method for creating 3-D radiographic information based on optical aperture theory, was evaluated for diagnostic efficacy in primary caries detection. STUDY DESIGN: Sixty-four extracted teeth with 89 carious lesions were imaged with D-speed film, direct digital, and TACT modalities. A commercially available, 8-bit, charge-coupled device was used in the later two modalities. Six trained observers were asked to identify the presence or absence and depth of interproximal and occlusal lesions for all three modalities. The teeth were sectioned and examined microscopically to determine ground truth. Logistic regression analysis was performed for all three imaging systems for the detection task. Analysis of variance was used for depth determination. Detection of lesion, depth of lesion accuracy, and time for diagnosis were also examined. RESULTS: For caries detection TACT and film were not different (p = 0.2216) with the Wald statistic. Film and TACT were significantly more accurate than the digital system (p = 0.0001). Scheffe's post hoc test revealed that TACT and film were more accurate than the direct digital system for determining lesion depth (p = 0.05) but not statistically different when compared with each other. The detection data were substantiated further by receiver operating characteristic analysis that demonstrated similar statistical relationships. Time required per diagnosis was not shown to be statistically different among the three imaging modalities. CONCLUSIONS: We conclude for caries detection and depth determination that TACT could not be distinguished from film despite the significant relative loss of information capacity in the charge-coupled device receptor. The relatively poorer performance yielded by the digital control images suggests that increased information capacity associated with more modern charge-coupled device detectors may improve diagnostic performance for both direct digital and TACT displays over that demonstrated in this investigation.