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1.  Effect of the soil type on the microbiome in the rhizosphere of field-grown lettuce 
The complex and enormous diversity of microorganisms associated with plant roots is important for plant health and growth and is shaped by numerous factors. This study aimed to unravel the effects of the soil type on bacterial communities in the rhizosphere of field-grown lettuce. We used an experimental plot system with three different soil types that were stored at the same site for 10 years under the same agricultural management to reveal differences directly linked to the soil type and not influenced by other factors such as climate or cropping history. Bulk soil and rhizosphere samples were collected 3 and 7 weeks after planting. The analysis of 16S rRNA gene fragments amplified from total community DNA by denaturing gradient gel electrophoresis and pyrosequencing revealed soil type dependent differences in the bacterial community structure of the bulk soils and the corresponding rhizospheres. The rhizosphere effect differed depending on the soil type and the plant growth developmental stage. Despite the soil type dependent differences in the bacterial community composition several genera such as Sphingomonas, Rhizobium, Pseudomonas, and Variovorax were significantly increased in the rhizosphere of lettuce grown in all three soils. The number of rhizosphere responders was highest 3 weeks after planting. Interestingly, in the soil with the highest numbers of responders the highest shoot dry weights were observed. Heatmap analysis revealed that many dominant operational taxonomic units were shared among rhizosphere samples of lettuce grown in diluvial sand, alluvial loam, and loess loam and that only a subset was increased in relative abundance in the rhizosphere compared to the corresponding bulk soil. The findings of the study provide insights into the effect of soil types on the rhizosphere microbiome of lettuce.
doi:10.3389/fmicb.2014.00144
PMCID: PMC3986527  PMID: 24782839
Lactuca sativa; bacterial communities; 16S rRNA gene analysis; DGGE; pyrosequencing; rhizosphere responders
2.  NAC changes the course of cerebral small vessel disease in SHRSP and reveals new insights for the meaning of stases - a randomized controlled study 
Background
N-Acetylcystein (NAC) reduces the reperfusion injury and infarct size in experimental macroangiopathic stroke. Here we now investigate the impact of NAC on the development of the histopathology of microangiopathic cerebrovascular disease including initial intravasal erythrocyte accumulations, blood–brain-barrier (BBB)-disturbances, microbleeds and infarcts.
Methods
Spontaneously Hypertensive Stroke-Prone Rats (SHRSP) were treated with NAC (12 mg/kg body weight, daily oral application for three to 30 weeks) and compared to untreated SHRSP. In all rats the number of microbleeds, thromboses, infarcts and stases were quantified by HE-staining. Exemplary brains were stained against von Willebrand factor (vWF), IgG, Glutathione and GFAP.
Results
NAC animals exhibited significant more microbleeds, a greater number of vessels with BBB-disturbances, but also an elevation of Glutathione-levels in astrocytes surrounding small vessels. NAC-treatment reduced the numbers of thromboses, infarcts and arteriolar stases.
Conclusions
NAC reduces the frequency of thromboses and infarcts to the expense of an increase of small microbleeds in a rat model of microangiopathic cerebrovascular disease. We suppose that NAC acts via an at least partial inactivation of vWF resulting in an insufficient sealing of initial endothelial injury leading to more small microbleeds. By elevating Glutathione-levels NAC most likely exerts a radical scavenger function and protects small vessels against extended ruptures and subsequent infarcts. Finally, it reveals that stases are mainly caused by endothelial injuries and restricted thromboses.
doi:10.1186/2040-7378-5-5
PMCID: PMC3661381  PMID: 23587288
Animal model; Blood–brain barrier; Cerebral microbleed; Cerebral small vessel disease; von Willebrand factor
3.  The pathologic cascade of cerebrovascular lesions in SHRSP: is erythrocyte accumulation an early phase? 
Cerebral small vessel disease (CSVD) is associated with vessel wall changes, microbleeds, blood–brain barrier (BBB) disturbances, and reduced cerebral blood flow (CBF). As spontaneously hypertensive stroke-prone rats (SHRSP) may be a valid model of some aspects of human CSVD, we aimed to identify whether those changes occur in definite temporal stages and whether there is an initial phenomenon beyond those common vascular alterations. Groups of 51 SHRSP were examined simultaneously by histologic (Hematoxylin–Eosin, IgG-Immunohistochemistry, vessel diameter measurement) and imaging methods (Magnetic Resonance Imaging, 201-Thallium-Diethyldithiocarbamate/99m-Technetium-HMPAO Single Photon Emission Computed Tomography conducted as pilot study) at different stages of age. Vascular pathology in SHRSP proceeds in definite stages, whereas an age-dependent accumulation of erythrocytes in capillaries and arterioles represents the homogeneous initial step of the disease. Erythrocyte accumulations are followed by BBB disturbances and microbleeds, both also increasing with age. Microthromboses, tissue infarctions with CBF reduction, and disturbed potassium uptake represent the final stage of vascular pathology in SHRSP. Erythrocyte accumulations—we parsimoniously interpreted as stases—without cerebral tissue damage represent the first step of vascular pathology in SHRSP. If that initial phenomenon could be identified in patients, these erythrocyte accumulations might be a promising target for implementing prophylactic and therapeutic strategies in human CSVD.
doi:10.1038/jcbfm.2011.122
PMCID: PMC3272595  PMID: 21878945
animal models; blood–brain barrier; microcirculation; SPECT; white matter disease
4.  A prospective longitudinal cohort study: evolution of GERD symptoms during the course of pregnancy 
BMC Gastroenterology  2012;12:131.
Background
Symptoms of gastro-esophageal reflux disease (GERD) in pregnancy are reported with a prevalence of 30–80%. The aim of this study was to assess the prevalence and severity of GERD symptoms during the course of pregnancy. Furthermore current practice in medical care for GERD during pregnancy was assessed.
Methods
We performed a prospective longitudinal cohort study on 510 pregnant women (mean age 28.12, SD 5.3). Investigations for reflux symptoms where based on the use of validated reflux-disease questionnaire (RDQ). Additional information was collected about the therapy. A group of non-pregnant women (mean age 24.56, SD 5.7) was included as controls. Frequency and severity of reflux symptoms were recorded in each trimester of pregnancy.
Results
The prevalence of GERD symptoms in pregnant women increased from the first trimester with 26.1 to 36.1% in the second trimester and to 51.2% in the third trimester of pregnancy. The prevalence of GERD symptoms in the control group was 9.3%.
Pregnant women received medication for their GERD symptoms in 12.8% during the first, 9.1% during the second and 15.7% during the third trimester. Medications used >90% antacids, 0% PPI.
Conclusion
GERD symptoms occur more often in pregnant women than in non-pregnant and the frequency rises in the course of pregnancy. Medical therapy is used in a minority of cases and often with no adequate symptom relief.
doi:10.1186/1471-230X-12-131
PMCID: PMC3499455  PMID: 23006768
Gastro-esophageal reflux disease; Pregnancy; Heartburn; Regurgitation; GERD symptoms
5.  Role of tight junction proteins in gastroesophageal reflux disease 
BMC Gastroenterology  2012;12:128.
Background
Gastroesophageal reflux disease (GERD) is associated with impaired epithelial barrier function that is regulated by cell-cell contacts. The aim of the study was to investigate the expression pattern of selected components involved in the formation of tight junctions in relation to GERD.
Methods
Eighty-four patients with GERD-related symptoms with endoscopic signs (erosive: n = 47) or without them (non-erosive: n = 37) as well as 26 patients lacking GERD-specific symptoms as controls were included. Endoscopic and histological characterization of esophagitis was performed according to the Los Angeles and adapted Ismeil-Beigi criteria, respectively. Mucosal biopsies from distal esophagus were taken for analysis by histopathology, immunohistochemistry and quantitative reverse-transcription polymerase chain reaction (RT-PCR) of five genes encoding tight junction components [Occludin, Claudin-1, -2, Zona occludens (ZO-1, -2)].
Results
Histopathology confirmed GERD-specific alterations as dilated intercellular spaces in the esophageal mucosa of patients with GERD compared to controls (P < 0.05). Claudin-1 and −2 were 2- to 6-fold upregulation on transcript (P < 0.01) and in part on protein level (P < 0.015) in GERD, while subgroup analysis of revealed this upregulation for ERD only. In both erosive and non-erosive reflux disease, expression levels of Occludin and ZO-1,-2 were not significantly affected. Notably, the induced expression of both claudins did not correlate with histopathological parameters (basal cell hyperplasia, dilated intercellular spaces) in patients with GERD.
Conclusions
Taken together, the missing correlation between the expression of tight junction-related components and histomorphological GERD-specific alterations does not support a major role of the five proteins studied in the pathogenesis of GERD.
doi:10.1186/1471-230X-12-128
PMCID: PMC3503771  PMID: 22994974
Gastroesophageal reflux disease; Tight junction; Claudins; Esophagitis; Inflammation
6.  Multitrophic Interaction in the Rhizosphere of Maize: Root Feeding of Western Corn Rootworm Larvae Alters the Microbial Community Composition 
PLoS ONE  2012;7(5):e37288.
Background
Larvae of the Western Corn Rootworm (WCR) feeding on maize roots cause heavy economical losses in the US and in Europe. New or adapted pest management strategies urgently require a better understanding of the multitrophic interaction in the rhizosphere. This study aimed to investigate the effect of WCR root feeding on the microbial communities colonizing the maize rhizosphere.
Methodology/Principal Findings
In a greenhouse experiment, maize lines KWS13, KWS14, KWS15 and MON88017 were grown in three different soil types in presence and in absence of WCR larvae. Bacterial and fungal community structures were analyzed by denaturing gradient gel electrophoresis (DGGE) of the16S rRNA gene and ITS fragments, PCR amplified from the total rhizosphere community DNA. DGGE bands with increased intensity were excised from the gel, cloned and sequenced in order to identify specific bacteria responding to WCR larval feeding. DGGE fingerprints showed that the soil type and the maize line influenced the fungal and bacterial communities inhabiting the maize rhizosphere. WCR larval feeding affected the rhiyosphere microbial populations in a soil type and maize line dependent manner. DGGE band sequencing revealed an increased abundance of Acinetobacter calcoaceticus in the rhizosphere of several maize lines in all soil types upon WCR larval feeding.
Conclusion/Significance
The effects of both rhizosphere and WCR larval feeding seemed to be stronger on bacterial communities than on fungi. Bacterial and fungal community shifts in response to larval feeding were most likely due to changes of root exudation patterns. The increased abundance of A. calcoaceticus suggested that phenolic compounds were released upon WCR wounding.
doi:10.1371/journal.pone.0037288
PMCID: PMC3358342  PMID: 22629377
7.  Why (not) go east? Comparison of findings from FDA Investigational New Drug study site inspections performed in Central and Eastern Europe with results from the USA, Western Europe, and other parts of the world 
Since the mid-1990s, investigational sites in the countries of Central and Eastern Europe (CEE) have been increasingly utilized by pharmaceutical companies because of their high productivity in terms of patient enrolment into clinical trials. Based on the FDA’s publicly accessible Clinical Investigator Inspection List, we present an analysis of findings and outcome classifications from FDA inspections during Investigational New Drug (IND) studies and compare the results for the CEE region to those from Western European countries and the USA. Data from all 5531 FDA clinical trials inspections that occurred between 1994 (when the FDA first performed inspections in CEE) and the end of 2010 were entered into the database for comparative analysis. Of these, 4865 routine data audit (DA) inspections were analyzed: 401 from clinical trials performed in Western Europe, 230 in CEE, 3858 in the USA, and 376 in other countries. The average number of deficiencies per inspection ranged between 0.99 for CEE and 1.97 in Western Europe. No deficiencies were noted during 16.6%, 39.0%, and 21.5% of the inspections in Western Europe, CEE and USA, respectively. The percentages of inspections after which no follow-up action was indicated were 36.9% for Western Europe, 55.7% for CEE, and 44.3% for US sites. CEE was also the region with the lowest percentage of inspections that required official or voluntary action. On the basis of FDA inspection data, the high productivity of CEE sites appears to be accompanied by regulatory compliance as well as by data quality standards that are not inferior to those in Western regions.
doi:10.2147/DDDT.S30109
PMCID: PMC3340105  PMID: 22563236
clinical trials; inspection; Central and Eastern Europe (CEE); data quality; deficiencies
8.  Clinical evaluation study of the German network of disorders of sex development (DSD)/intersexuality: study design, description of the study population, and data quality 
BMC Public Health  2009;9:110.
Background
The German Network of Disorders of Sex Development (DSD)/Intersexuality carried out a large scale clinical evaluation study on quality of life, gender identity, treatment satisfaction, coping, and problems associated with diagnoses and therapies in individuals with disorders of sex development (DSD). DSD are a heterogeneous group of various genetic disorders of sex determination or sex differentiation, all of which are rare conditions. In about half of all cases the molecular genetic diagnosis is unknown and diagnosis rests on clinical features.
Methods and design
The multi-centre clinical evaluation study includes short-term follow-up in some and cross-sectional assessments in all age and diagnostic groups fitting the criteria of DSD. Recruitment was from January 2005 until December 2007 in whole Germany and, additionally, in 2007 in Austria and German-speaking Switzerland. The study consists of a psychosocial inquiry for children, adolescents and their parents, and adults with standardized instruments and the collection of DSD-specific medical data by the attending physician. The main goal was the description of clinical outcomes and the health-care situation of individuals with DSD using a broad generic definition of DSD including all conditions with a mismatch of chromosomal, gonadal and phenotypical sex. 439 children and adolescents, their parents and adults with DSD participated.
Discussion
The clinical evaluation study represents the most comprehensive study in this clinical field. The paper discusses the study protocol, the data management and data quality as well as the classification used, and it describes the study population. Given the lack of large datasets in rare conditions such as DSD and often biased results from small scale clinical case series, the study aims to generate concrete hypotheses for evidence-based guidelines, which should be tested in further studies.
doi:10.1186/1471-2458-9-110
PMCID: PMC2678119  PMID: 19383134
9.  Nonparametric relevance-shifted multiple testing procedures for the analysis of high-dimensional multivariate data with small sample sizes 
BMC Bioinformatics  2008;9:54.
Background
In many research areas it is necessary to find differences between treatment groups with several variables. For example, studies of microarray data seek to find a significant difference in location parameters from zero or one for ratios thereof for each variable. However, in some studies a significant deviation of the difference in locations from zero (or 1 in terms of the ratio) is biologically meaningless. A relevant difference or ratio is sought in such cases.
Results
This article addresses the use of relevance-shifted tests on ratios for a multivariate parallel two-sample group design. Two empirical procedures are proposed which embed the relevance-shifted test on ratios. As both procedures test a hypothesis for each variable, the resulting multiple testing problem has to be considered. Hence, the procedures include a multiplicity correction. Both procedures are extensions of available procedures for point null hypotheses achieving exact control of the familywise error rate. Whereas the shift of the null hypothesis alone would give straight-forward solutions, the problems that are the reason for the empirical considerations discussed here arise by the fact that the shift is considered in both directions and the whole parameter space in between these two limits has to be accepted as null hypothesis.
Conclusion
The first algorithm to be discussed uses a permutation algorithm, and is appropriate for designs with a moderately large number of observations. However, many experiments have limited sample sizes. Then the second procedure might be more appropriate, where multiplicity is corrected according to a concept of data-driven order of hypotheses.
doi:10.1186/1471-2105-9-54
PMCID: PMC2268654  PMID: 18221560
10.  Synaptic proteome changes in mouse brain regions upon auditory discrimination learning 
Proteomics  2012;12(15-16):2433-2444.
Changes in synaptic efficacy underlying learning and memory processes are assumed to be associated with alterations of the protein composition of synapses. Here, we performed a quantitative proteomic screen to monitor changes in the synaptic proteome of four brain areas (auditory cortex, frontal cortex, hippocampus striatum) during auditory learning. Mice were trained in a shuttle box GO/NO-GO paradigm to discriminate between rising and falling frequency modulated tones to avoid mild electric foot shock. Control-treated mice received corresponding numbers of either the tones or the foot shocks. Six hours and 24 h later, the composition of a fraction enriched in synaptic cytomatrix-associated proteins was compared to that obtained from naïve mice by quantitative mass spectrometry. In the synaptic protein fraction obtained from trained mice, the average percentage (±SEM) of downregulated proteins (59.9 ± 0.5%) exceeded that of upregulated proteins (23.5 ± 0.8%) in the brain regions studied. This effect was significantly smaller in foot shock (42.7 ± 0.6% down, 40.7 ± 1.0% up) and tone controls (43.9 ± 1.0% down, 39.7 ± 0.9% up). These data suggest that learning processes initially induce removal and/or degradation of proteins from presynaptic and postsynaptic cytoskeletal matrices before these structures can acquire a new, postlearning organisation. In silico analysis points to a general role of insulin-like signalling in this process.
doi:10.1002/pmic.201100669
PMCID: PMC3509369  PMID: 22696468
Animal proteomics; Auditory learning; Chemical synapse; Isotope-coded protein labelling; Learning and memory; Quantitative mass spectrometry

Results 1-10 (10)