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We report a design methodology for creating high-performance photonic crystals with arbitrary geometric shapes. This design approach enables the inclusion of subwavelength shapes into the photonic crystal unit cell, synergistically combining metamaterials concepts with on-chip guided-wave photonics. Accordingly, we use the term "photonic metacrystal" to describe this class of photonic structures. Photonic metacrystals exploiting three different design freedoms are demonstrated experimentally. With these additional degrees of freedom in the design space, photonic metacrystals enable added control of light-matter interactions and hold the promise of significantly increasing temporal confinement in all-dielectric metamaterials.
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Early radical cystectomy (RC) is a therapeutic option for non-muscle invasive bladder cancer (NMIBC). The 15-year overall survival after early RC in NMIBC patients is about 70%. Nevertheless, RC is associated with significant morbidity and mortality and therefore requires careful patient selection. The aim of the following review is to assess the selection process for early RC in NMIBC. Especially, the new European Association of Urology (EAU) risk calculator identifying NMIBC patients with very high risk for disease progression is described in detail. Furthermore, the technical aspects of the procedure are evaluated. A review of the current literature (PubMed) and national and international guideline recommendations was also conducted.
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Cistectomía , Neoplasias de la Vejiga Urinaria , Humanos , Invasividad Neoplásica , Selección de Paciente , Neoplasias de la Vejiga Urinaria/cirugíaRESUMEN
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
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Infection and inflammation of the mammary gland, and especially prevention of mastitis, are still major challenges for the dairy industry. Different approaches have been tried to reduce the incidence of mastitis. Genetic selection of cows with lower susceptibility to mastitis promises sustainable success in this regard. Bos taurus autosome (BTA) 18, particularly the region between 43 and 59 Mb, harbors quantitative trait loci (QTL) for somatic cell score, a surrogate trait for mastitis susceptibility. Scrutinizing the molecular bases hereof, we challenged udders from half-sib heifers having inherited either favorable paternal haplotypes for somatic cell score (Q) or unfavorable haplotypes (q) with the Staphylococcus aureus pathogen. RNA sequencing was used for an in-depth analysis of challenge-related alterations in the hepatic transcriptome. Liver exerts highly relevant immune functions aside from being the key metabolic organ. Hence, a holistic approach focusing on the liver enabled us to identify challenge-related and genotype-dependent differentially expressed genes and underlying regulatory networks. In response to the S. aureus challenge, we found that heifers with Q haplotypes displayed more activated immune genes and pathways after S. aureus challenge compared with their q half-sibs. Furthermore, we found a significant enrichment of differentially expressed loci in the genomic target region on BTA18, suggesting the existence of a regionally acting regulatory element with effects on a variety of genes in this region.
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Predisposición Genética a la Enfermedad , Hígado/metabolismo , Mastitis Bovina/inmunología , Infecciones Estafilocócicas/veterinaria , Staphylococcus aureus , Transcriptoma , Animales , Bovinos , Industria Lechera , Susceptibilidad a Enfermedades/veterinaria , Femenino , Haplotipos , Glándulas Mamarias Animales/metabolismo , Mastitis Bovina/genética , Fenotipo , Sitios de Carácter Cuantitativo , Selección Genética , Análisis de Secuencia de ARN , Infecciones Estafilocócicas/genética , Staphylococcus aureus/genética , Vacunación/veterinariaRESUMEN
We developed a time-efficient semi-automated axon quantification method using freeware in human cranial nerve sections stained with paraphenylenediamine (PPD). It was used to analyze a total of 1238 facial and masseteric nerve biopsies. The technique was validated by comparing manual and semi-automated quantification of 129 (10.4%) randomly selected biopsies. The software-based method demonstrated a sensitivity of 94% and a specificity of 87%. Semi-automatic axon counting was significantly faster (p < 0.001) than manual counting. It took 1 hour and 47 minutes for all 129 biopsies (averaging 50 sec per biopsy, 0.04 seconds per axon). The counting process is automatic and does not need to be supervised. Manual counting took 21 hours and 6 minutes in total (average 9 minutes and 49 seconds per biopsy, 0.52 seconds per axon). Our method showed a linear correlation to the manual counts (R = 0.944 Spearman rho). Attempts have been made by several research groups to automate axonal load quantification. These methods often require specific hard- and software and are therefore only accessible to a few specialized laboratories. Our semi-automated axon quantification is precise, reliable and time-sparing using publicly available software and should be useful for an effective axon quantification in various human peripheral nerves.
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BACKGROUND: In dairy herds, mastitis causes detrimental economic losses. Genetic selection offers a sustainable tool to select animals with reduced susceptibility towards postpartum diseases. Studying underlying mechanisms is important to assess the physiological processes that cause differences between selected haplotypes. Therefore, the objective of this study was to establish an in vivo infection model to study the impact of selecting for alternative paternal haplotypes in a particular genomic region on cattle chromosome 18 for mastitis susceptibility under defined conditions in uniparous dairy cows. RESULTS: At the start of pathogen challenge, no significant differences between the favorable (Q) and unfavorable (q) haplotypes were detected. Intramammary infection (IMI) with Staphylococcus aureus 1027 (S. aureus, n = 24, 96 h) or Escherichia coli 1303 (E. coli, n = 12, 24 h) was successfully induced in all uniparous cows. This finding was confirmed by clinical signs of mastitis and repeated recovery of the respective pathogen from milk samples of challenged quarters in each animal. After S. aureus challenge, Q-uniparous cows showed lower somatic cell counts 24 h and 36 h after challenge (P < 0.05), lower bacterial shedding in milk 12 h after challenge (P < 0.01) and a minor decrease in total milk yield 12 h and 24 h after challenge (P < 0.01) compared to q-uniparous cows. CONCLUSION: An in vivo infection model to study the impact of genetic selection for mastitis susceptibility under defined conditions in uniparous dairy cows was successfully established and revealed significant differences between the two genetically selected haplotype groups. This result might explain their differences in susceptibility towards IMI. These clinical findings form the basis for further in-depth molecular analysis to clarify the underlying genetic mechanisms for mastitis resistance.
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Mastitis Bovina/genética , Mastitis Bovina/microbiología , Herencia Paterna , Animales , Bovinos , Industria Lechera , Escherichia coli , Infecciones por Escherichia coli/veterinaria , Femenino , Haplotipos , Masculino , Leche/microbiología , Infecciones Estafilocócicas/veterinaria , Staphylococcus aureusRESUMEN
The susceptibility of animals to periparturient diseases has a great effect on the economic efficiency of dairy industries, on the frequency of antibiotic treatment, and on animal welfare. The use of selection for breeding cows with reduced susceptibility to diseases offers a sustainable tool to improve dairy cattle farming. Several studies have focused on the association of distinct bovine chromosome 18 genotypes or haplotypes with performance traits. The aim of this study was to test whether selection of Holstein Friesian heifers via SNP genotyping for alternative paternal chromosome 18 haplotypes associated with favorable (Q) or unfavorable (q) somatic cell scores influences postpartum reproductive and metabolic diseases. Thirty-six heifers (18 Q and 18 q) were monitored from 3 wk before calving until necropsy on d 39 (± 4 d) after calving. Health status and rectal temperature were measured daily, and body condition score and body weight were assessed once per week. Blood samples were drawn twice weekly, and levels of insulin, nonesterified fatty acids, insulin-like growth factor-I, growth hormone, and ß-hydroxybutyrate were measured. Comparisons between the groups were performed using Fisher's exact test, chi-squared test, and the GLIMMIX procedure in SAS. Results showed that Q-heifers had reduced incidence of metritis compared with q-heifers and were less likely to develop fever. Serum concentrations of ß-hydroxybutyrate were lower and insulin-like growth factor-I plasma concentrations were higher in Q- compared with q-heifers. However, the body condition score and withers height were comparable between haplotypes, but weight loss tended to be lower in Q-heifers compared with q-heifers. No differences between the groups were detected concerning retained fetal membranes, uterine involution, or onset of cyclicity. In conclusion, selection of chromosome 18 haplotypes associated with a reduced somatic cell score resulted in a decreased incidence of postpartum reproductive and metabolic diseases in this study. The presented data add to the existing knowledge aimed at avoiding negative consequences of genetic selection strategies in dairy cattle farming. The underlying causal mechanisms modulated by haplotypes in the targeted genomic region and immune competence necessitate further investigation.
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Bovinos/genética , Cromosomas de los Mamíferos , Haplotipos , Periodo Posparto , Reproducción , Selección Genética , Ácido 3-Hidroxibutírico/sangre , Animales , Peso Corporal , Bovinos/metabolismo , Enfermedades de los Bovinos/genética , Industria Lechera , Ácidos Grasos no Esterificados/sangre , Femenino , Hormona del Crecimiento/sangre , Insulina/sangre , Lactancia , Retención de la Placenta/veterinaria , Polimorfismo de Nucleótido Simple , EmbarazoRESUMEN
The original article [1] contained an error whereby the captions to Figs 2 and 3 were mistakenly inverted; this has now been corrected.
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BACKGROUND: A major challenge in modern medicine and animal husbandry is the issue of antimicrobial resistance. One approach to solving this potential medical hazard is the selection of farm animals with less susceptibility to infectious diseases. Recent advances in functional genome analysis and quantitative genetics have opened the horizon to apply genetic marker information for efficiently identifying animals with preferential predisposition regarding health traits. The current study characterizes functional traits with a focus on udder health in dairy heifers. The animals were selected for having inherited alternative paternal haplotypes for a genomic region on Bos taurus chromosome (BTA) 18 genetically associated with divergent susceptibility to longevity and animal health, particularly mastitis. RESULTS: In the first weeks of lactation, the q heifers which had inherited the unfavorable (q) paternal haplotype displayed a significantly higher number of udder quarters with very low somatic cell count (< 10,000 cells / ml) compared to their paternal half-sib sisters with the favorable (Q) paternal haplotype. This might result in impaired mammary gland sentinel function towards invading pathogens. Furthermore, across the course of the first lactation, there was indication that q half-sib heifers showed higher somatic cell counts, a surrogate trait for udder health, in whole milkings compared to their paternal half-sib sisters with the favorable (Q) paternal haplotype. Moreover, heifers with the haplotype Q had a higher feed intake and higher milk yield compared to those with the q haplotype. Results of this study indicate that differences in milk production and calculated energy balance per se are not the main drivers of the genetically determined differences between the BTA18 Q and q groups of heifers. CONCLUSIONS: The paternally inherited haplotype from a targeted BTA18 genomic region affect somatic cell count in udder quarters during the early postpartum period and might also contribute to further aspects of animal's health and performance traits due to indirect effects on feed intake and metabolism.
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Glándulas Mamarias Animales/fisiología , Herencia Paterna , Animales , Bovinos , Mapeo Cromosómico , Ingestión de Alimentos/genética , Metabolismo Energético/genética , Femenino , Haplotipos/genética , Lactancia/genética , Masculino , Glándulas Mamarias Animales/microbiología , Mastitis Bovina/genética , Periodo PospartoRESUMEN
BACKGROUND AND OBJECTIVES: Early persistent facial paralysis is characterized by intact muscles of facial expression through maintained perfusion but lacking nerve supply. In facial reanimation procedures aiming at restoration of facial tone and dynamics, neurotization through a donor nerve is performed. Critical for reanimating target muscles is axonal capacity of both donor and recipient nerves. In cases of complete paralysis, the proximal stump of the extratemporal facial nerve trunk may be selected as a recipient site for coaptation. To further clarify the histological basis of this facial reanimation procedure we conducted a human cadaver study examining macro and micro anatomical features of the facial nerve trunk including its axonal capacity in human cadavers. Axonal loads, morphology and morbidity of different donor nerves are discussed reviewing literature in context of nerve transfers. METHODS: From 6/2015 to 9/2016 in a group of 53 fresh frozen cadavers a total of 106 facial halves were dissected. Biopsies of the extratemporal facial nerve trunk (FN) were obtained at 1âcm distal to the stylomastoid foramen. After histological processing and digitalization of 99 specimens available, 97 were selected eligible for fascicle counts and 87 fulfilled quality criteria for a semi-automated computer-based axon quantification software using ImageJ/Fiji. RESULTS: An average of 3.82 fascicles (range, 1 to 9) were noted (nâ=â97). 6684±1884 axons (range, 2655- 12457) were counted for the entire group (nâ=â87). Right facial halves showed 6364±1904 axons (nâ=â43). Left facial halves demonstrated 6996±1833 axons (nâ=â44) with no significant difference (pâ=â0.73). Female cadavers featured 6247±2230 (nâ=â22), male showed 6769±1809 axons (nâ=â40). No statistical difference was seen between genders (pâ=â0.59). A comparison with different studies in literature is made. The nerve diameter in 82 of our specimens could be measured at 1933±424 µm (range, 975 to 3012). CONCLUSIONS: No donor nerve has been described to match axonal load or fascicle number of the extratemporal facial nerve main trunk. However, the masseteric nerve may be coapted for neurotization of facial muscles with a low complication rate and good clinical outcomes. Nerve transfer is indicated from 6 months after onset of facial paralysis if no recovery of facial nerve function is seen.
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Músculos Faciales/anatomía & histología , Nervio Facial/anatomía & histología , Parálisis Facial/cirugía , Transferencia de Nervios/métodos , Axones , Músculos Faciales/patología , Nervio Facial/patología , Parálisis Facial/patología , Femenino , Humanos , MasculinoRESUMEN
The zygomaticus major (ZM) is important for the human smile. There are conflicting data about whether the zygomatic or buccal branches of the facial nerve are responsible for its motor innervation. The literature provides no precise distinction of the transition zone between these two branch systems. In this study, a definition to distinguish the facial nerve branches at the level of the body of the zygoma is proposed. In the light of this definition, we conducted an anatomical study to determine how the source of innervation of the ZM was distributed. A total of 96 fresh-frozen cadaveric facial halves were dissected under loupe magnification. A hemiparotidectomy was followed by antegrade microsurgical dissection. Any branch topographically lying superficial to the zygoma or touching it was classed as zygomatic, and any neighboring inferior branch was considered buccal. The arborization of the facial nerve was diffuse in all cases. In 64 out of 96 specimens (67%, 95% CI: 56% to 76%), zygomatic branches innervated the ZM. Buccal branches innervated ZM in the other 32 facial halves (33%, 95% CI: 24% to 44%). There were no differences in respect of sex or facial side. All facial halves displayed additional branches, which crossed the muscle on its inner surface without supplying it. In 31 specimens, a nerve branch ran superficial to ZM in its cranial third. According to our classification, the zygomaticus major is innervated by zygomatic branches in 67% of cases and by buccal branches in 33%. Clin. Anat. 31:560-565, 2018. © 2018 Wiley Periodicals, Inc.
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Músculos Faciales/inervación , Nervio Facial/anatomía & histología , Variación Anatómica , Femenino , Humanos , Masculino , Sonrisa/fisiologíaRESUMEN
OBJECTIVES: Anti-Müllenria hormone (AMH) is an established biomarker for assessing ovarian reserve and predicting response to controlled ovarian stimulation. Its routine clinical use is hampered by the variability and low-throughput of available enzyme-linked immunosorbent assays (ELISA). The presented study examined if a fully automated AMH electrochemiluminescence assay (ECLIA; Elecsys® AMH assay, Roche Diagnostics) was suitable for measuring AMH levels in healthy women and in those diagnosed with polycystic ovary syndrome (PCOS). DESIGN AND METHODS: Five European laboratories evaluated the Elecsys® AMH assay independently under routine conditions over eight months. Within-run imprecision, repeatability, intermediate precision, linearity and functional sensitivity were assessed. The Elecsys® AMH assay was compared to a manual ELISA microtiter plate format test (AMH Gen II ELISA, modified version; Beckman Coulter Inc.) using 1729 routine serum samples. AMH reference intervals were determined in 887 healthy women with regular menstrual cycle aged 2050 years, and 149 women diagnosed with PCOS. RESULTS: The fully automated Elecsys® AMH assay showed excellent precision, linearity, and functional sensitivity. The coefficient of variation was 1.8% for repeatability and 4.4% for intermediate precision. Values measured with the Elecsys® AMH assay were highly correlated with the manual ELISA method (modified version) but 2428% lower. Reference intervals showed the expected AMH decline with age in healthy women and increased AMH levels in women with PCOS. CONCLUSIONS: The Elecsys® AMH assay demonstrated good precision under routine conditions, and is suitable for determining AMH levels in serum and lithium-heparin plasma.
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Hormona Antimülleriana/análisis , Ensayo de Inmunoadsorción Enzimática/métodos , Mediciones Luminiscentes/métodos , Adolescente , Adulto , Hormona Antimülleriana/sangre , Automatización de Laboratorios/instrumentación , Automatización de Laboratorios/métodos , Biomarcadores/sangre , Ensayo de Inmunoadsorción Enzimática/normas , Femenino , Humanos , Laboratorios , Mediciones Luminiscentes/normas , Ciclo Menstrual/metabolismo , Persona de Mediana Edad , Síndrome del Ovario Poliquístico/sangre , Valores de Referencia , Reproducibilidad de los Resultados , Sensibilidad y EspecificidadRESUMEN
INTRODUCTION: The correct therapeutic management of acute sigmoid diverticulitis (SD) is still controversially discussed. Essential to the success of therapy is primarily the long-term resolution of Patient symptoms after surgical or conservative therapy. The aim of this study was to compare the long-term outcome after conservative and surgical treatment of Patients with acute SD. PATIENTS AND METHODS: Consecutive admissions of all Patients with acute SD were prospectively recruited from January 2005 to June 2008 with the exception of a free perforation. The following data were recorded: age, sex, first or recurrent episode of SD, computed tomography (CT) stage, white blood cell count, C-reactive protein, persistent symptoms and recurrence after conservative and surgical therapy. Furthermore, information on the rates of postoperative sexual and bladder dysfunction was collected. The long-term outcome was evaluated by a standardized questionnaire. In June 2008 all Patients were contacted using a standardized questionnaire. RESULTS: A total of 153 Patients were included in the study of whom 70 (45.8 %) presented with the first episode, 83 (54.2 %) had a prior history of SD and 40 Patients were treated conservatively whereas 113 Patients were surgically treated by sigmoid resection. Uncomplicated SD was seen in 16 Patients (conservative 4, surgical 12, p = 0.961), phlegmonous SD was seen in 88 cases (conservative 29, surgical 59, p = 0.026) and covered perforated SD in 49 cases (conservative 7, surgical 42, p = 0.022). The median follow-up was 32 months (range 12-52 months). At follow-up 25 % of conservative and 8.8 % of Patients treated surgically complained about persistent symptoms (p = 0.009). The following symptoms occurred (conservative vs. surgery): painful defecation (22.5 % versus 8.8 %, p = 0.024.), constipation (25 % versus 8.8 %, p = 0.009), abdominal cramp (22.5 % versus 4.4 %, p = 0.001) and painful flatulence (25 % versus 8.8 %, p = 0.009). Sexual or bladder dysfunction occurred postoperatively in 7 % and 9 %, respectively. Of the conservatively treated Patients 32.5 % had a recurrence of SD during follow-up compared to 3.5 % of surgically treated Patients (p < 0.001). CONCLUSIONS: Surgical treatment of acute SD is more effective than conservative therapy for the prophylaxis of recurrent SD and avoidance of persistent symptoms.
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Diverticulitis del Colon/cirugía , Laparoscopía/métodos , Complicaciones Posoperatorias/etiología , Enfermedades del Sigmoide/cirugía , Enfermedad Aguda , Anciano , Antibacterianos/administración & dosificación , Enfermedad Crónica , Medios de Contraste/administración & dosificación , Diverticulitis del Colon/diagnóstico por imagen , Quimioterapia Combinada , Servicio de Urgencia en Hospital , Femenino , Estudios de Seguimiento , Humanos , Aumento de la Imagen/métodos , Enfermedades Intestinales/etiología , Perforación Intestinal/diagnóstico por imagen , Perforación Intestinal/cirugía , Masculino , Persona de Mediana Edad , Complicaciones Posoperatorias/diagnóstico por imagen , Recurrencia , Medición de Riesgo , Enfermedades del Sigmoide/diagnóstico por imagen , Tomografía Computarizada por Rayos XRESUMEN
Based on analyses in Arabidopsis thaliana ecotype C24, the AtSUC1 protein was previously characterised as a male gametophyte-specific H(+)/sucrose symporter. Later, expression analyses in ecotype Columbia-0 (Col-0) identified AtSUC1 expression also in trichomes (not detected in trichome-less C24 plants) and roots, suggesting ecotype-specific differences in AtSUC1 expression. Here, we present data on additional ecotype-specific differences in AtSUC1 expression in other tissues. Using different AtSUC1 promoter-reporter gene lines, we performed comparative analyses of AtSUC1 expression in floral tissues of C24 and Col-0 plants, and using an AtSUC1-specific antiserum, we performed immunohistochemical analyses on tissue sections from C24, Col-0, Landsberg erecta (Ler) and Wassilewskaija (Ws) ecotypes. We show that AtSUC1 expression occurs in the funicular epidermis of C24, Ler and Ws, but not in Col-0. In contrast, we observed high levels of AtSUC1 protein in pollen grains of Col-0, lower levels in pollen of C24 and Ler, and no AtSUC1 protein in Ws pollen. Moreover, our reporter gene analyses identified a previously undetected expression of AtSUC1 in the female gametophyte, and revealed that AtSUC1 expression in the funicular epidermis is absent from unpollinated siliques and is induced upon successful pollination. The impact of these findings on the potential physiological role of AtSUC1 is discussed.
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Proteínas de Arabidopsis/metabolismo , Arabidopsis/genética , Proteínas de Transporte de Membrana/metabolismo , Óvulo Vegetal/metabolismo , Proteínas de Plantas/metabolismo , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Regulación de la Expresión Génica de las Plantas , Genes Reporteros , Proteínas de Transporte de Membrana/genética , Mutagénesis Insercional , Epidermis de la Planta/metabolismo , Proteínas de Plantas/genética , Plantas Modificadas Genéticamente/genética , Plantas Modificadas Genéticamente/metabolismo , Polen/metabolismo , Regiones Promotoras Genéticas , Transformación GenéticaRESUMEN
The pathogenicity of Staphylococcus aureus is determined by its ability to express multiple virulence factors. Thus far the virulence potential of S. aureus isolates has been described by the virulence gene repertoire, which, in part, varies considerably among the different isolates. Extracellular proteins constitute a reservoir of virulence factors and have been shown to play an important role in the pathogenicity of bacteria. Analyses of the expression of these virulence factors and elucidation of regulatory networks involved in S. aureus virulence by using gel based proteomics can yield information important for our understanding of the virulence potential of this pathogen and its interaction with the host. In addition, these approaches are critical for a comprehensive understanding of secretion and modification of virulence factors.
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Proteómica , Staphylococcus aureus , Proteínas Bacterianas , Infecciones Estafilocócicas/microbiología , Staphylococcus aureus/genética , Factores de Virulencia/genéticaRESUMEN
The gram-positive bacterium Staphylococcus aureus is a frequent component of the human microbial flora that can turn into a dangerous pathogen. As such, this organism is capable of infecting almost every tissue and organ system in the human body. It does so by actively exporting a variety of virulence factors to the cell surface and extracellular milieu. Upon reaching their respective destinations, these virulence factors have pivotal roles in the colonization and subversion of the human host. It is therefore of major importance to obtain a clear understanding of the protein transport pathways that are active in S. aureus. The present review aims to provide a state-of-the-art roadmap of staphylococcal secretomes, which include both protein transport pathways and the extracytoplasmic proteins of these organisms. Specifically, an overview is presented of the exported virulence factors, pathways for protein transport, signals for cellular protein retention or secretion, and the exoproteomes of different S. aureus isolates. The focus is on S. aureus, but comparisons with Staphylococcus epidermidis and other gram-positive bacteria, such as Bacillus subtilis, are included where appropriate. Importantly, the results of genomic and proteomic studies on S. aureus secretomes are integrated through a comparative "secretomics" approach, resulting in the first definition of the core and variant secretomes of this bacterium. While the core secretome seems to be largely employed for general housekeeping functions which are necessary to thrive in particular niches provided by the human host, the variant secretome seems to contain the "gadgets" that S. aureus needs to conquer these well-protected niches.
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Proteínas Bacterianas/metabolismo , Staphylococcus/metabolismo , Proteínas Bacterianas/fisiología , Microscopía Electrónica , Transporte de Proteínas/fisiología , Proteómica/métodos , Transducción de Señal/fisiología , Staphylococcus/patogenicidad , Staphylococcus/ultraestructura , Staphylococcus aureus/metabolismo , Staphylococcus aureus/patogenicidad , Staphylococcus aureus/ultraestructura , VirulenciaRESUMEN
Phosphoenolpyruvate carboxylase (PEPCase, EC 4.1.1.3) is a key enzyme of C(4) photosynthesis. It has evolved from ancestral non-photosynthetic (C(3)) isoforms and thereby changed its kinetic and regulatory properties. We are interested in understanding the molecular changes, as the C(4) PEPCases were adapted to their new function in C(4) photosynthesis and have therefore analysed the PEPCase genes of various Alternanthera species. We isolated PEPCase cDNAs from the C(4) plant Alternanthera pungens H.B.K., the C(3)/C(4) intermediate plant A. tenella Colla, and the C(3) plant A. sessilis (L.) R.Br. and investigated the kinetic properties of the corresponding recombinant PEPCase proteins and their phylogenetic relationships. The three PEPCases are most likely derived from orthologous gene classes named ppcA. The affinity constant for the substrate phosphoenolpyruvate (K (0.5) PEP) and the degree of activation by glucose-6-phosphate classified the enzyme from A. pungens (C(4)) as a C(4) PEPCase isoform. In contrast, both the PEPCases from A. sessilis (C(3)) and A. tenella (C(3)/C(4)) were found to be typical C(3) PEPCase isozymes. The C(4) characteristics of the PEPCase of A. pungens were accompanied by the presence of the C(4)-invariant serine residue at position 775 reinforcing that a serine at this position is essential for being a C(4) PEPCase (Svensson et al. 2003). Genomic Southern blot experiments and sequence analysis of the 3' untranslated regions of these genes indicated the existence of PEPCase multigene family in all three plants which can be grouped into three classes named ppcA, ppcB and ppcC.
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Amaranthaceae/enzimología , Amaranthaceae/genética , Evolución Molecular , Fosfoenolpiruvato Carboxilasa/genética , Amaranthaceae/fisiología , Secuencia de Aminoácidos , Southern Blotting , Paseo de Cromosoma , ADN Complementario/metabolismo , Isoenzimas/clasificación , Isoenzimas/genética , Isoenzimas/metabolismo , Cinética , Datos de Secuencia Molecular , Familia de Multigenes/genética , Fosfoenolpiruvato Carboxilasa/clasificación , Fosfoenolpiruvato Carboxilasa/metabolismo , Fotosíntesis/genética , Proteínas del Complejo del Centro de Reacción Fotosintética/clasificación , Proteínas del Complejo del Centro de Reacción Fotosintética/genética , Proteínas del Complejo del Centro de Reacción Fotosintética/metabolismo , Filogenia , Hojas de la Planta/enzimología , Proteínas Recombinantes , Alineación de Secuencia , Homología de Secuencia de AminoácidoRESUMEN
Resistance of the pathogenic yeast Candida albicans to the antifungal agent fluconazole is often caused by the overexpression of genes that encode multidrug efflux pumps ( CDR1, CDR2, or MDR1). We have undertaken a proteomic approach to gain further insight into the regulatory network controlling efflux pump expression and drug resistance in C. albicans. Three pairs of matched fluconazole-susceptible and resistant clinical C. albicans isolates, in which drug resistance correlated with stable activation of MDR1 or CDR1/2, were analyzed for differences in their protein expression profiles. In two independent, MDR1-overexpressing, strains, additional up-regulated proteins were identified, which are encoded by the YPR127 gene and several members of the IFD ( YPL088) gene family. All are putative aldo-keto reductases of unknown function. These proteins were not up-regulated in a fluconazole-resistant strain that overexpressed CDR1 and CDR2 but not MDR1, indicating that expression of the various efflux pumps of C. albicans is controlled by different regulatory networks. To investigate the possible role of YPR127 in the resistance phenotype of the clinical isolates, we constitutively overexpressed the gene in a C. albicans laboratory strain. In addition, the gene was deleted in a C. albicans laboratory strain and in one of the drug-resistant clinical isolates in which it was overexpressed. Neither forced overexpression nor deletion of YPR127 affected the susceptibility of the strains to drugs and other toxic substances, suggesting that the regulatory networks which control the expression of efflux pumps in C. albicans also control genes involved in cellular functions not related to drug resistance.
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Antifúngicos/farmacología , Candida albicans/efectos de los fármacos , Farmacorresistencia Fúngica/genética , Fluconazol/farmacología , Resistencia a Múltiples Medicamentos , Proteínas Fúngicas/genética , Eliminación de Gen , Expresión Génica , Regulación Fúngica de la Expresión Génica , Estrés Oxidativo/genética , Transformación GenéticaRESUMEN
The influence of the two anthracyclin antibiotics daunomycin and adriamycin on the proteome of Bacillus subtilis was investigated. They intercalate in the double helix causing strand breaks. Both compounds induce proteins related to DNA damage and oxidative stress as indicated by the induction of some members of the PerR and the DinR-regulon. The mild induction of some members of the HrcA- and the CtsR-regulon may indicate protein oxidation as well. Furthermore, an influence on the sigmaB-dependent general stress response was shown. These data show that the proteomic signature is a valuable experimental tool for a comprehensive evaluation of the action of different drugs.