RESUMO
BACKGROUND: Clinically, coronavirus disease 2019 (COVID-19) is associated with a wide range of symptoms, which can range from mild complaints of an upper respiratory infection to life-threatening hypoxic respiratory insufficiency and multiorgan failure. OBJECTIVE: The initially identified pulmonary damage patterns, such as diffuse alveolar damage in acute lung failure, are accompanied by new findings that draw a more complex scenario. These include microvascular involvement and a wide range of associated pathologies of multiple organ systems. A back-scaling of microstructural vascular changes is possible via targeted correlation of pathological autopsy results with radiological imaging. MATERIAL AND METHODS: Radiological and pathological correlation as well as microradiological imaging to investigate microvascular involvement in fatal COVID-19. RESULTS: The cases of two COVID-19 patients are presented. Patient 1 showed a relative hypoperfusion in lung regions that did not have typical COVID-19 infiltrates; the targeted post-mortem correlation also showed subtle signs of microvascular damage even in these lung sections. Patient 2 showed both radiologically and pathologically advanced typical COVID-19 destruction of lung structures and the case illustrates the damage patterns of the blood-air barrier. The perfusion deficit of the intestinal wall shown in computed tomography of patient 2 could not ultimately clearly be microscopically attributed to intestinal microvascular damage. CONCLUSION: In addition to microvascular thrombosis, our results indicate a functional pulmonary vasodysregulation as part of the pathophysiology during the vascular phase of COVID-19. The clinical relevance of autopsies and the integration of radiological imaging findings into histopathological injury patterns must be emphasized for a better understanding of COVID-19.
Assuntos
Betacoronavirus , Infecções por Coronavirus , Pandemias , Pneumonia Viral , COVID-19 , Humanos , Microvasos , SARS-CoV-2RESUMO
Evolutionary transitions from outcrossing to selfing can strongly affect the genetic diversity and structure of species at multiple spatial scales. We investigated the genetic consequences of mating-system shifts in the North American, Pacific coast dune endemic plant Camissoniopsis cheiranthifolia (Onagraceae) by assaying variation at 13 nuclear (n) and six chloroplast (cp) microsatellite (SSR) loci for 38 populations across the species range. As predicted from the expected reduction in effective population size (Ne ) caused by selfing, small-flowered, predominantly selfing (SF) populations had much lower nSSR diversity (but not cpSSR) than large-flowered, predominantly outcrossing (LF) populations. The reduction in nSSR diversity was greater than expected from the effects of selfing on Ne alone, but could not be accounted for by indirect effects of selfing on population density. Although selfing should reduce gene flow, SF populations were not more genetically differentiated than LF populations. We detected five clusters of nSSR genotypes and three groups of cpSSR haplotypes across the species range consisting of parapatric groups of populations that usually (but not always) differed in mating system, suggesting that selfing may often initiate ecogeographic isolation. However, lineage-wide genetic variation was not lower for selfing clusters, failing to support the hypothesis that selection for reproductive assurance spurred the evolution of selfing in this species. Within three populations where LF and SF plants coexist, we detected genetic differentiation among diverged floral phenotypes suggesting that reproductive isolation (probably postzygotic) may help maintain the striking mating-system differentiation observed across the range of this species.
Assuntos
Ecossistema , Onagraceae/genética , Onagraceae/fisiologia , Teorema de Bayes , Análise por Conglomerados , Loci Gênicos , Variação Genética , Genética Populacional , Geografia , Haplótipos/genética , Repetições de Microssatélites/genética , Fenótipo , Densidade Demográfica , Análise de Regressão , ReproduçãoRESUMO
By reducing genetically effective population size and gene flow, self-fertilization should lead to strong spatial genetic structure (SGS). Although the short-lived plant Aquilegia canadensis produces large, complex, nectar-rich flowers, 75% of seed, on average, are self-fertilized. Previous experimental results are consistent with the fine-scale SGS expected in selfing populations. In contrast, key floral traits show no evidence of SGS, despite a significant genetic basis to phenotypic variation within populations. In this study, we attempt to resolve these contradictory results by hierarchically sampling plants from two plots nested within each of seven rock outcrops distributed over several km, and comparing the spatial pattern of phenotypic variation in four floral traits with neutral genetic variation at 10 microsatellite loci. For both floral and microsatellite variation, we detected only weak hierarchical structuring and no isolation by distance. The spatial pattern of variation in floral traits was on par with microsatellite polymorphisms. These results suggest regular long-distance gene flow via pollen. At much finer spatial scales within plots, estimates of relatedness were higher (albeit very low) between nearest neighbors than random plants, and declined with increasing distance between neighbors, which is consistent with highly localized seed dispersal. High selfing should yield SGS, but strong inbreeding depression in A. canadensis likely erodes SGS so that reproductive plants exhibit weak structure typical of outcrossers, especially given that outcrossing and consequent gene flow in this species are mediated by strong-flying hummingbirds and bumble bees.
Assuntos
Fertilização , Genes de Plantas , Variação Genética , Ranunculaceae/genética , Ranunculaceae/fisiologia , FloresRESUMO
Clostridium difficile can form biofilms. Thirty-seven strains were characterized for their ability to form a biofilm, adhesion on an inert surface and hydrophobicity. No correlation between the ability to form a biofilm and the strain virulence was highlighted. However, non-motile strains were not able to form a high biofilm.
Assuntos
Biofilmes/crescimento & desenvolvimento , Clostridioides difficile/fisiologia , Microbiologia Ambiental , Aderência Bacteriana , Clostridioides difficile/crescimento & desenvolvimento , Interações Hidrofóbicas e Hidrofílicas , Locomoção , Propriedades de Superfície , VirulênciaRESUMO
Calprotectin and lactoferrin are released by the gastrointestinal tract in response to infection and mucosal inflammation. Our objective was to assess the usefulness of quantifying faecal lactoferrin and calprotectin concentrations in Clostridium difficile infection (CDI) patients with or without free toxins in the stools. We conducted a single-centre 22-month case-control study. Patients with a positive CDI diagnosis were compared to two control groups: group 1 = diarrhoeic patients negative for C. difficile and matched (1:1) to CDI cases on the ward location and age, and group 2 = diarrhoeic patients colonised with a non-toxigenic strain of C. difficile. Faecal lactoferrin and calprotectin concentrations in faeces were determined for patients with CDI and controls. Of 135 patients with CDI, 87 (64.4%) had a positive stool cytotoxicity assay (free toxin) and 48 (35.6%) had a positive toxigenic culture without detectable toxins in the stools. The median lactoferrin values were 26.8 µg/g, 8.0 µg/g and 15.8 µg/g in CDI patients and groups 1 and 2, respectively. The median calprotectin values were 218.0 µg/g, 111.5 µg/g and 111.3 µg/g, respectively. Among patients with CDI, faecal lactoferrin and calprotectin levels were higher in those with free toxins in their stools (39.2 vs. 10.2 µg/g, p = 0.003 and 274.0 vs. 166.0 µg/g, p = 0.051, respectively). Both faecal calprotectin and lactoferrin were higher in patients with CDI, especially in those with detectable toxin in faeces, suggesting a correlation between intestinal inflammation and toxins in stools.
Assuntos
Clostridioides difficile , Infecções por Clostridium/metabolismo , Infecções por Clostridium/microbiologia , Fezes/química , Lactoferrina , Complexo Antígeno L1 Leucocitário , Idoso , Estudos de Casos e Controles , Feminino , Trato Gastrointestinal/metabolismo , Humanos , Lactoferrina/metabolismo , Complexo Antígeno L1 Leucocitário/metabolismo , Masculino , Pessoa de Meia-Idade , Curva ROCRESUMO
Lack of standardised Clostridium difficile testing is a potential confounder when comparing infection rates. We used an observational, systematic, prospective large-scale sampling approach to investigate variability in C. difficile sampling to understand C. difficile infection (CDI) incidence rates. In-patient and institutional data were gathered from 60 European hospitals (across three countries). Testing methodology, testing/CDI rates and case profiles were compared between countries and institution types. The mean annual CDI rate per hospital was lowest in the UK and highest in Italy (1.5 vs. 4.7 cases/10,000 patient bed days [pbds], p < 0.001). The testing rate was highest in the UK compared with Italy and France (50.7/10,000 pbds vs. 31.5 and 30.3, respectively, p < 0.001). Only 58.4 % of diarrhoeal samples were tested for CDI across all countries. Overall, only 64 % of hospitals used recommended testing algorithms for laboratory testing. Small hospitals were significantly more likely to use standalone toxin tests (SATTs). There was an inverse correlation between hospital size and CDI testing rate. Hospitals using SATT or assays not detecting toxin reported significantly higher CDI rates than those using recommended methods, despite testing similar testing frequencies. These data are consistent with higher false-positive rates in such (non-recommended) testing scenarios. Cases in Italy and those diagnosed by SATT or methods NOT detecting toxin were significantly older. Testing occurred significantly earlier in the UK. Assessment of testing practice is paramount to the accurate interpretation and comparison of CDI rates.
Assuntos
Clostridioides difficile/isolamento & purificação , Infecções por Clostridium/diagnóstico , Infecções por Clostridium/epidemiologia , Testes Diagnósticos de Rotina/métodos , Diarreia/diagnóstico , Diarreia/epidemiologia , Técnicas Microbiológicas/métodos , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Criança , Pré-Escolar , Infecções por Clostridium/microbiologia , Erros de Diagnóstico , Testes Diagnósticos de Rotina/normas , Diarreia/microbiologia , Monitoramento Epidemiológico , Feminino , França/epidemiologia , Hospitais , Humanos , Incidência , Lactente , Recém-Nascido , Itália/epidemiologia , Masculino , Técnicas Microbiológicas/normas , Pessoa de Meia-Idade , Política Organizacional , Projetos Piloto , Reino Unido/epidemiologia , Adulto JovemRESUMO
The Suzuki coupling reaction of basic nitrogen containing substrates (2-bromo- and 2-chloro-4-aminopyridine, and 2-bromo and 2-chloropyridine) with phenylboronic acid using Pd(TPP)2Cl2/K3PO4 in acetonitrile-water biphasic solvent systems under a CO2 or a N2 atmosphere is discussed. It was observed that 2-halo-4-aminopyridine produced quantitative yields of coupled products under a CO2 atmosphere while the yields for the 2-halopyridines were poor. In contrast, the yields of coupled products for the 2-halopyridines substrates were quantitative under a N2 atmosphere while only poor yields were realized for the 2-halo-4-aminopyridines under the same conditions. Evidence is presented which suggests that the presence of CO2 alters the pH of the aqueous phase of the reaction system and the accompanying efficiency of the coupling process. Using a series of buffers to adjust the pH of the aqueous phase, the pH dependence associated with the efficiency of the coupling process is illustrated.
Assuntos
Dióxido de Carbono/química , Nitrogênio/química , Pressão , Acetonitrilas/química , Concentração de Íons de Hidrogênio , Cinética , Fosfatos/química , Fosfinas/química , Compostos de Potássio/química , TemperaturaRESUMO
Keeping up to date with the latest clinical advances in prostate cancer can be challenging. We investigated the impact of guideline use on quality of treatment decisions as well as the impact of a novel, CE-certified clinical decision support tool (Siemens AIPC software) on the amount of time clinicians spend on decision-making in a multicenter setting. Ten urologists assessed ten clinical cases (screening and localized prostate cancer) in three settings: without support, using a digital version of the EAU guidelines, and with the AIPC tool, resulting in 300 clinical decisions. Comparison involved time spent, decision correct- and completeness. Using AIPC compared to digital guidelines led to a significant reduction of expenditure of time at a per case level (3.57 min and 0:14 min, p < 0.01) and for overall time per urologist (39.45 min and 02:20 min, p < 0.01). Decision options without guidelines support, online guideline usage and usage of AIPC were complete in 61%, 80% and 100%, respectively (p < 0.01). Decision making without guidelines support, online guideline usage and usage of AIPC was correct including all options in 28%, 66% and 100%, respectively (p < 0.01).Clinical decision support systems have the potential to reduces decision-making time and to enhance decision quality.
Assuntos
Tomada de Decisão Clínica , Sistemas de Apoio a Decisões Clínicas , Guias de Prática Clínica como Assunto , Neoplasias da Próstata , Software , Humanos , Masculino , Neoplasias da Próstata/terapia , Neoplasias da Próstata/diagnóstico , Fidelidade a Diretrizes/estatística & dados numéricos , Pessoa de Meia-Idade , IdosoRESUMO
Theory predicts that inbreeding depression (ID) should decline via purging in self-fertilizing populations. Yet, intraspecific comparisons between selfing and outcrossing populations are few and provide only mixed support for this key evolutionary process. We estimated ID for large-flowered (LF), predominantly outcrossing vs. small-flowered (SF), predominantly selfing populations of the dune endemic Camissoniopsis cheiranthifolia by comparing selfed and crossed progeny in glasshouse environments differing in soil moisture, and by comparing allozyme-based estimates of the proportion of seeds selfed and inbreeding coefficient of mature plants. Based on lifetime measures of dry mass and flower production, ID was stronger in nine LF populations [mean δ = 1-(fitness of selfed seed/fitness of outcrossed seed) = 0.39] than 16 SF populations (mean δ = 0.03). However, predispersal ID during seed maturation was not stronger for LF populations, and ID was not more pronounced under simulated drought, a pervasive stress in sand dune habitat. Genetic estimates of δ were also higher for four LF (δ = 1.23) than five SF (δ = 0.66) populations; however, broad confidence intervals around these estimates overlapped. These results are consistent with purging, but selective interference among loci may be required to maintain strong ID in partially selfing LF populations, and trade-offs between selfed and outcrossed fitness are likely required to maintain outcrossing in SF populations.
Assuntos
Endogamia , Onagraceae/fisiologia , Autofertilização , Alelos , Secas , Evolução Molecular , Flores/anatomia & histologia , Flores/genética , Flores/fisiologia , Variação Genética , Funções Verossimilhança , Onagraceae/genética , Polinização , Sementes/fisiologia , Autoincompatibilidade em Angiospermas , Solo/química , Especificidade da EspécieRESUMO
The BFM studies for relapsed childhood acute lymphoblastic leukemia (ALL) were started in 1983, at a time when cure rates for ALL were still lower and the number of children with ALL relapse equaled about the number of children with newly diagnosed neuroblastoma. Today, relapses have become relatively rare events in ALL although, because of the frequency of ALL, they are still a significant cause of death in children and adolescents. With currently used treatment modalities, cure rates of about 50% after relapse can be achieved, and, together with the improved results of front-line therapy, the survival rate of childhood ALL is now about 90%. Most children with extramedullary and late bone marrow (BM) relapses achieve a second CR; remission rates in patients with high-risk features, however, remain still unsatisfactory. With improved techniques allogeneic hematopoietic stem cell transplant (HSCT) has become a relatively safe treatment but is not necessary for all patients as postremission therapy. HSCT is not required in children with isolated extramedullary and late BM relapses with rapid response to induction therapy measured by molecular techniques (minimal residual disease, MRD) but absolutely indicated in patients with early BM relapses and systemic relapses of T-cell ALL. For patients with insufficient response innovative therapies such as small molecules or targeted immunological or pharmacological approaches are urgently required. Efforts have to be made, therefore, in order to detect potential biological or molecular targets, which can be used for individualized, more effective and hopefully less toxic therapies in the future.
Assuntos
Neoplasia Residual/tratamento farmacológico , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamento farmacológico , Adolescente , Protocolos de Quimioterapia Combinada Antineoplásica , Asparaginase , Medula Óssea/patologia , Criança , Terapia Combinada , Ciclofosfamida , Citarabina , Daunorrubicina , Intervalo Livre de Doença , Europa (Continente) , Transplante de Células-Tronco Hematopoéticas , Humanos , Mercaptopurina , Metotrexato , Estudos Multicêntricos como Assunto , Neoplasia Residual/mortalidade , Neoplasia Residual/patologia , Leucemia-Linfoma Linfoblástico de Células Precursoras/mortalidade , Leucemia-Linfoma Linfoblástico de Células Precursoras/patologia , Leucemia-Linfoma Linfoblástico de Células T Precursoras/tratamento farmacológico , Leucemia-Linfoma Linfoblástico de Células T Precursoras/mortalidade , Leucemia-Linfoma Linfoblástico de Células T Precursoras/patologia , Prednisona , Ensaios Clínicos Controlados Aleatórios como Assunto , Recidiva , Indução de Remissão , Retratamento , Taxa de Sobrevida , VincristinaRESUMO
Chromosomal rearrangements of the human KMT2A/MLL gene are associated with de novo as well as therapy-induced infant, pediatric, and adult acute leukemias. Here, we present the data obtained from 3401 acute leukemia patients that have been analyzed between 2003 and 2022. Genomic breakpoints within the KMT2A gene and the involved translocation partner genes (TPGs) and KMT2A-partial tandem duplications (PTDs) were determined. Including the published data from the literature, a total of 107 in-frame KMT2A gene fusions have been identified so far. Further 16 rearrangements were out-of-frame fusions, 18 patients had no partner gene fused to 5'-KMT2A, two patients had a 5'-KMT2A deletion, and one ETV6::RUNX1 patient had an KMT2A insertion at the breakpoint. The seven most frequent TPGs and PTDs account for more than 90% of all recombinations of the KMT2A, 37 occur recurrently and 63 were identified so far only once. This study provides a comprehensive analysis of the KMT2A recombinome in acute leukemia patients. Besides the scientific gain of information, genomic breakpoint sequences of these patients were used to monitor minimal residual disease (MRD). Thus, this work may be directly translated from the bench to the bedside of patients and meet the clinical needs to improve patient survival.
Assuntos
Histona-Lisina N-Metiltransferase , Leucemia Mieloide Aguda , Proteína de Leucina Linfoide-Mieloide , Leucemia-Linfoma Linfoblástico de Células Precursoras , Humanos , Lactente , Pré-Escolar , Criança , Adolescente , Adulto , Pessoa de Meia-Idade , Idoso , Leucemia Mieloide Aguda/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Histona-Lisina N-Metiltransferase/genética , Proteína de Leucina Linfoide-Mieloide/genética , Fusão GênicaRESUMO
This study compared a repetitive-element PCR (rep-PCR) method (DiversiLab system) to PCR ribotyping. The discriminatory power of rep-PCR was 0.997. Among the PCR ribotype 027 isolates tested, different rep types could be distinguished. rep-PCR showed a higher discriminatory power than PCR ribotyping. Nevertheless, this method requires technical skill, and visual interpretation of rep-PCR fingerprint patterns may be difficult.
Assuntos
Clostridioides difficile/classificação , Tipagem Molecular/métodos , Reação em Cadeia da Polimerase/métodos , Adulto , Clostridioides difficile/genética , Humanos , Sensibilidade e EspecificidadeRESUMO
The gold standards for the diagnosis of Clostridium difficile infections (CDIs) are the cytotoxicity assay and the toxigenic culture. However, both methods are time-consuming and the results are not available before 24-48 h. We developed and evaluated a multiplex in-house real-time polymerase chain reaction (PCR) assay for the simultaneous detection of toxigenic strains of C. difficile and the presumptive identification of the epidemic NAP1/027/BI strain from stools. Amplifications were performed using specific primers for tcdB and tcdC on an ABI Prism 7300 (Applied Biosystems). The detection of amplicons was done using TaqMan probes. The analytical sensitivity of the multiplex real-time PCR for detecting tcdB was estimated to 10 CFU/g of stools. This assay was assessed from 881 consecutive unformed stools from patients suspected of having CDI. The gold standard was the toxigenic culture for the diagnosis of CDI and PCR ribotyping for the identification of the NAP1/027/BI strain. The prevalence of positive toxigenic culture was 9.31%. Compared to the toxigenic culture, the sensitivity, specificity, and positive and negative predictive values were 86.59%, 97.43%, 78.02%, and 98.57%, respectively, for the real-time PCR and 70.73%, 100%, 100%, and 97.08%, respectively, for the cytotoxicity assay.
Assuntos
Técnicas Bacteriológicas/métodos , Clostridioides difficile/isolamento & purificação , Infecções por Clostridium/diagnóstico , Técnicas de Diagnóstico Molecular/métodos , Reação em Cadeia da Polimerase Multiplex/métodos , Reação em Cadeia da Polimerase em Tempo Real/métodos , Proteínas de Bactérias/genética , Toxinas Bacterianas/genética , Clostridioides difficile/genética , Infecções por Clostridium/microbiologia , Primers do DNA/genética , Fezes/microbiologia , Humanos , Sondas de Oligonucleotídeos/química , Sondas de Oligonucleotídeos/genética , Proteínas Repressoras/genética , Sensibilidade e EspecificidadeRESUMO
OBJECTIVES: We aimed to evaluate if malnutrition and rurality are associated with fall risk and future falls in community-dwelling older adults. DESIGN: Prospective Cohort. SETTING: Community, Vermont. PARTICIPANTS: Older adults receiving home support services who completed a health risk assessment (n=3,300; Mean age 79.6 years ±8.4, 75% female). Additional analysis was completed with a subset of 2,043 participants with two-years of consecutive health assessments. MEASUREMENTS: Fall Risk Questionnaire, DETERMINE Nutrition Risk Questionnaire, and fall history. RESULTS: Independently, high malnutrition risk and rurality were associated with fall risk (p<0.001) and high malnutrition risk was associated with rurality (p<0.001). After adjusting for age, sex, and physical function, individuals with high nutrition risk had a 66% increase in the odds of falling over the next year, but rurality was not significantly associated with a new fall. CONCLUSION: These findings suggest that falls are associated with malnutrition risk, but the relationship between falls and rurality is less evident. Further research is needed to identify services that may best alleviate malnutrition risk in older adults and aspects of nutrition that are most protective against fall risk.
Assuntos
Acidentes por Quedas , Idoso , Vida Independente , Desnutrição , Acidentes por Quedas/prevenção & controle , Atividades Cotidianas , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Desnutrição/complicações , Desnutrição/epidemiologia , Estudos ProspectivosRESUMO
In 2006 and 2007, a large outbreak of Clostridium difficile infections (CDIs) with PCR-ribotype 027 was identified in northern France. Overall, 38 healthcare facilities notified 529 CDIs over a 22-month period, including 281 laboratory-confirmed CDI 027 and 248 non-confirmed CDI 027 cases (incidence rate per 10,000 elective bed days: 1.63, range: 0.07 to 7.94). The cases occurred mainly in long-term care hospital facilities and nursing homes, near the border between France and Belgium. An active surveillance and prevention campaign was launched at the first epidemic peak including hygiene precautions for healthcare professionals, which supported healthcare facilities to improve care organisation. The outbreak was controlled at the end of 2007, but sporadic cases were identified until the end of 2009. A bundle of appropriate control measures may halt the spread of such outbreaks, provided that substantial human resources and financial support are available.
Assuntos
Clostridioides difficile/genética , Enterocolite Pseudomembranosa/epidemiologia , Idoso , Idoso de 80 Anos ou mais , Clostridioides difficile/isolamento & purificação , Surtos de Doenças/prevenção & controle , Enterocolite Pseudomembranosa/prevenção & controle , Feminino , França/epidemiologia , Instalações de Saúde , Humanos , Masculino , Reação em Cadeia da Polimerase , RibotipagemRESUMO
The magnitude of inbreeding depression, a central parameter in the evolution of plant mating systems, can vary depending on environmental conditions. However, the underlying genetic mechanisms causing environmental fluctuations in inbreeding depression, and the consequences of this variation for the evolution of self-fertilization, have been little studied. Here, we consider temporal fluctuations of the selection coefficient in an explicit genetic model of inbreeding depression. We show that substantial variance in inbreeding depression can be generated at equilibrium by fluctuating selection, although the simulated variance tends to be lower than has been measured in experimental studies. Our simulations also reveal that purging of deleterious mutations does not depend on the variance in their selection coefficient. Finally, an evolutionary analysis shows that, in contrast to previous theoretical approaches, intermediate selfing rates are never evolutionarily stable when the variation in inbreeding depression is due to fluctuations in the selection coefficient on deleterious mutations.
Assuntos
Evolução Biológica , Endogamia , Modelos Genéticos , Fenômenos Fisiológicos Vegetais/genética , Simulação por Computador , Variação GenéticaRESUMO
Pathogenesis of Clostridium difficile has been linked to production of toxins, including the large toxins A and B as well as the binary toxin CDT. Until recently, toxin A was only found in combination in clinical strains with the toxin B, unlike toxin B or CDT, which were found alone in toxigenic variants. New toxigenic variants of C. difficile detected in our laboratory from patients with diarrhoea or severe colitis, including a variant producing only toxin A, were tested for virulence in the hamster model, which displays the clinical features of C. difficile disease. Hamsters infected with a strain producing only toxin B induced similar clinical signs, time to death from infection and histologic damage compared to the hypervirulent strain 027. No mortality or clinical signs of infection but caecal histologic damage was found with the variant producing only toxin A. The C. difficile variant strain producing only CDT was able to kill one hamster out of seven; nevertheless, the surviving animals had few alteration of the caecum.
RESUMO
OBJECTIVES: Reported rates of community-acquired Clostridium difficile infections (CDIs) have been increasing. However, the true burden of the disease in general practice is unknown in France. Our objective was to determine the incidence of toxigenic C. difficile carriage and the percentage of stool samples prescribed by general practitioners (GPs) which contained free C. difficile toxins. METHODS: During an 11-month period, all stool samples submitted for any enteric pathogen detection to 15 different private laboratories in Paris and the surrounding areas were tested for C. difficile, irrespective of the GPs' request. A clinical questionnaire was completed for each patient. Stool samples were screened using a rapid simultaneous glutamate dehydrogenase and toxins A/B detection test: any positive result (glutamate dehydrogenase or toxin) was further confirmed by the stool cytotoxicity assay (CTA) on MRC-5 cells and by toxigenic culture (TC) at a central laboratory. The C. difficile isolates were characterized by PCR ribotyping. RESULTS: A total of 2541 patients (1295 female, 1246 male) were included. The incidences of patients with a positive toxigenic culture and a positive CTA were 3.27% (95% CI 2.61%-4.03%) and 1.81% (95% CI 1.33%-2.41%), respectively. GPs requested C. difficile testing in only 12.93% of the stool samples, detecting 52.30% of all TC-positive patients. The 83 toxigenic C. difficile strains belonged to 36 different PCR ribotypes. CONCLUSIONS: Toxigenic C. difficile carriage is frequent in general practice but remains under-recognized. It may affect young patients without previous antimicrobial therapy or hospitalization.
Assuntos
ADP Ribose Transferases/análise , Proteínas de Bactérias/análise , Portador Sadio/epidemiologia , Clostridioides difficile/isolamento & purificação , Infecções por Clostridium/epidemiologia , Infecções Comunitárias Adquiridas/epidemiologia , Medicina Geral , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Criança , Pré-Escolar , Clostridioides difficile/classificação , Clostridioides difficile/genética , Clostridioides difficile/metabolismo , Fezes/microbiologia , Feminino , Humanos , Incidência , Lactente , Recém-Nascido , Masculino , Pessoa de Meia-Idade , Paris/epidemiologia , Estudos Prospectivos , Ribotipagem , Adulto JovemRESUMO
CRISPR-Cas systems have revolutionized genome editing across a broad range of biotechnological endeavors. Many CRISPR-Cas nucleases have been identified and engineered for improved capabilities. Given the modular structure of such enzymes, we hypothesized that engineering chimeric sequences would generate non-natural variants that span the kinetic parameter landscape, and thus provide for the rapid selection of nucleases fit for a particular editing system. Here, we design a chimeric Cas12a-type library with approximately 560 synthetic chimeras, and select several functional variants. We demonstrate that certain nuclease domains can be recombined across distantly related nuclease templates to produce variants that function in bacteria, yeast, and human cell lines. We further characterize selected chimeric nucleases and find that they have different protospacer adjacent motif (PAM) preferences and the M44 chimera has higher specificity relative to wild-type (WT) sequences. This demonstration opens up the possibility of generating nuclease sequences with implications across biotechnology.
Assuntos
Sistemas CRISPR-Cas , Endonucleases/metabolismo , Edição de Genes/métodos , Proteínas Recombinantes de Fusão/metabolismo , Bactérias/genética , Biotecnologia/métodos , Endonucleases/genética , Biblioteca Gênica , Células HEK293 , Humanos , Mutação , Proteínas Recombinantes de Fusão/genética , Reprodutibilidade dos Testes , Leveduras/genéticaRESUMO
There is growing interest in quantifying genetic population structure across the geographical ranges of species to understand why species might exhibit stable range limits and to assess the conservation value of peripheral populations. However, many assertions regarding peripheral populations rest on the long-standing but poorly tested supposition that peripheral populations exhibit low genetic diversity and greater genetic differentiation as a consequence of smaller effective population size and greater geographical isolation relative to geographically central populations. We reviewed 134 studies representing 115 species that tested for declines in within-population genetic diversity and/or increases in among-population differentiation towards range margins using nuclear molecular genetic markers. On average, 64.2% of studies detected the expected decline in diversity, 70.2% of those that tested for it showed increased differentiation and there was a positive association between these trends. In most cases, however, the difference in genetic diversity between central and peripheral population was not large. Although these results were consistent across plants and animals, strong taxonomic and biogeographical biases in the available studies call for a cautious generalization of these results. Despite the large number of studies testing these simple predictions, very few attempted to test possible mechanisms causing reduced peripheral diversity or increased differentiation. Almost no study incorporated a phylogeographical framework to evaluate historical influences on contemporary genetic patterns. Finally, there has been little effort to test whether these geographical trends in putatively neutral variation at marker loci are reflected by quantitative genetic trait variation, which is likely to influence the adaptive potential of populations across the geographical range.