Can We Use Tree Rings of Black Alder to Reconstruct Lake Levels? A Case Study for the Mecklenburg Lake District,Northeastern Germany

In this study, we explore the potential to reconstruct lake-level (and groundwater) fluctuations from tree-ring chronologies of black alder (Alnus glutinosa L.) for three study lakes in the Mecklenburg Lake District, northeastern Germany. As gauging records for lakes in this region are generally short, long-term reconstructions of lake-level fluctuations could provide valuable information on past hydrological conditions, which, in turn, are useful to assess dynamics of climate and landscape evolution. We selected black alder as our study species as alder typically thrives as riparian vegetation along lakeshores. For the study lakes, we tested whether a regional signal in lake-level fluctuations and in the growth of alder exists that could be used for long-term regional hydrological reconstructions, but found that local (i.e. site-specific) signals in lake level and tree-ring chronologies prevailed. Hence, we built lake/groundwater-level reconstruction models for the three study lakes individually. Two sets of models were considered based on (1) local tree-ring series of black alder, and (2) site-specific Standardized Precipitation Evapotranspiration Indices (SPEI). Although the SPEI-based models performed statistically well, we critically reflect on the reliability of these reconstructions, as SPEI cannot account for human influence. Tree-ring based reconstruction models, on the other hand, performed poor. Combined, our results suggest that, for our study area, long-term regional reconstructions of lake-level fluctuations that consider both recent and ancient (e.g., archaeological) wood of black alder seem extremely challenging, if not impossible.     

Criteria Used in Clinical Practice to Guide Immunosuppressive Treatment in Patients with Primary Sclerosing Cholangitis

Background & Aims

Current guidelines recommend immunosuppressive treatment (IT) in patients with primary sclerosing cholangitis (PSC) and elevated aminotransferase levels more than five times the upper limit of normal and elevated serum IgG-levels above twice the upper limit of normal. Since there is no evidence to support this recommendation, we aimed to assess the criteria that guided clinicians in clinical practice to initiate IT in patients with previously diagnosed PSC.

Methods

This is a retrospective analysis of 196 PSC patients from seven German hepatology centers, of whom 36 patients had received IT solely for their liver disease during the course of PSC. Analyses were carried out using methods for competing risks.

Results

A simplified autoimmune hepatitis (AIH) score >5 (HR of 36, p<0.0001) and a modified histological activity index (mHAI) greater than 3/18 points (HR 3.6, p = 0.0274) were associated with the initiation of IT during the course of PSC. Of note, PSC patients who subsequently received IT differed already at the time of PSC diagnosis from those patients, who did not receive IT during follow-up: they presented with increased levels of IgG (p = 0.004) and more frequently had clinical signs of cirrhosis (p = 0.0002).

Conclusions

This is the first study which investigates the parameters associated with IT in patients with PSC in clinical practice. A simplified AIH score >5 and a mHAI score >3, suggesting concomitant features of AIH, influenced the decision to introduce IT during the course of PSC. In German clinical practice, the cutoffs used to guide IT may be lower than recommended by current guidelines.     

Developmental Exposure to Fluoxetine Modulates the Serotonin System in Hypothalamus

The selective serotonin reuptake inhibitor (SSRI) fluoxetine (FLU, Prozac®) is commonly prescribed for depression in pregnant women. This results in SSRI exposure of the developing fetus. However, there are knowledge gaps regarding the impact of SSRI exposure during development. Given the role of serotonin in brain development and its cross-talk with sex hormone function, we investigated effects of developmental exposure to pharmacologically relevant concentrations of FLU (3 and 30 nM (measured)) on brain neurotransmitter levels, gonadal differentiation, aromatase activity in brain and gonads, and the thyroid system, using the Xenopus tropicalis model. Tadpoles were chronically exposed (8 weeks) until metamorphosis. At metamorphosis brains were cryosectioned and levels of serotonin, dopamine, norepinephrine, and their metabolites 5-hydroxyindoleacetic acid, 3,4-dihydroxyphenylacetic acid, and homovanillic acid were measured in discrete regions (telencephalon, hypothalamus and the reticular formation) of the cryosections using high-performance liquid chromatography. Exposure to 30 nM FLU increased the concentration of 5-hydroxyindoleacetic acid in hypothalamus compared with controls. FLU exposure did not affect survival, time to metamorphosis, thyroid histology, gonadal sex differentiation, or aromatase activity implying that the effect on the serotonergic neurotransmitter system in the hypothalamus region was specific. The FLU concentration that impacted the serotonin system is lower than the concentration measured in umbilical cord serum, suggesting that the serotonin system of the developing brain is highly sensitive to in utero exposure to FLU. To our knowledge this is the first study showing effects of developmental FLU exposure on brain neurochemistry. Given that SSRIs are present in the aquatic environment the current results warrant further investigation into the neurobehavioral effects of SSRIs in aquatic wildlife.     

Raccoons (Procyon lotor) in Germany as potential reservoir species for Lyssaviruses

Raccoons can be found almost everywhere in Germany since their first successful introduction in 1934. Although the animal is a well-known reservoir species for rabies in the USA, during the last European fox rabies epizootic, only a few rabid raccoons were reported from Germany. In recent years, the raccoon population density has increased tremendously, especially in (semi) urban settings. Presently, Germany is free of terrestrial wildlife rabies. To assess the potential risk that the raccoon population in Germany could act as a reservoir species upon reemergence of rabies, the susceptibility of the local raccoon population was investigated. Wild-caught animals were inoculated with the most likely lyssavirus variants to infect the local population. It was shown that the raccoons were fully susceptible for a dog and raccoon rabies virus isolate. Also, five of six raccoons inoculated with a fox rabies virus isolate showed clinical signs. However, none of the raccoons infected with European Bat Lyssavirus type 1 succumbed to rabies; meanwhile, all these raccoons seroconverted. It is concluded that the highest risk for the raccoon population in Germany to become infected with lyssaviruses is through the importation of rabies infected dogs.     

The predictive value of trill performance in a large repertoire songbird,the nightingale Luscinia megarhynchos

In animal communication, elaborate signals have been shown to be under sexual selection and often to reliably indicate a signaler's quality, condition, or motivation. For instance, the performance of physically challenging signals such as trills – i.e. rapidly repeated elements of broad frequency bandwidth – is considered to reflect signaler quality. Nightingales Luscinia megarhynchos are renowned for their outstanding song repertoire sizes, and most songs include a variety of complex trills. In the present study, we examined whether performance of trills can reliably reflect male quality. We show that vocal performance of trills predicts the age of a male. Older males sang trills that were closer to the performance limit than did younger males. Moreover, males with narrower beaks sang more consistent trills than did males with wider beaks. Vocal performance of trills, however, did not significantly predict other measures of biometric quality such as body size or body condition of the males. The findings suggest that receivers could benefit from the predictive value of physically demanding song traits in assessing age as an important quality component of potential mates or rivals. Particularly in species with high singing versatility, signaler assessment based on readily assessable structures may be adaptive, as this will allow receivers to quickly gather relevant information about the singer without attending to the full song repertoire.     

Vertical distribution and cyst production of Peridiniella catenata (Dinophyceae) during a spring bloom in the Baltic Sea

Vertical distribution and cyst production of the chain-forming,spring dinoflagellate Peridiniella catenata were studied throughoutthe spring season of 2000 in the coastal Gulf of Finland. Numbersof cells were monitored in the water column, and cyst sedimentationwas recorded using multiple sediment traps moored at three discretedepths. At the onset of the spring bloom, most of the populationwas situated in the euphotic zone. When the bloom progressed,the population was more evenly dispersed throughout the watercolumn. Coinciding with the decline of the spring bloom, afternitrogen depletion, a general reduction of cell size of P. catenataand a break-up of chains were observed. Resting cysts startedto appear shortly after the peak of the bloom, in sedimentationtraps moored at 30 and 40 m depth. Cysts were only retrievedfrom the uppermost sediment trap on three of the six samplingoccasions, constituting only a small proportion of all cystsproduced by P. catenata during spring. Our results suggest thatcyst production of this vertically migrating organism takesplace to a large extent in deep water layers and emphasizesthe necessity of whole water column monitoring in studies aimingto understand in situ life-cycle transformations of verticallymigrating dinoflagellates.     

Multivariate curve resolution of time course microarray data

Background  

Modeling of gene expression data from time course experiments often involves the use of linear models such as those obtained from principal component analysis (PCA), independent component analysis (ICA), or other methods. Such methods do not generally yield factors with a clear biological interpretation. Moreover, implicit assumptions about the measurement errors often limit the application of these methods to log-transformed data, destroying linear structure in the untransformed expression data.     

Extending the Limits of Quantitative Proteome Profiling with Data-Independent Acquisition and Application to Acetaminophen-Treated Three-Dimensional Liver Microtissues

The data-independent acquisition (DIA) approach has recently been introduced as a novel mass spectrometric method that promises to combine the high content aspect of shotgun proteomics with the reproducibility and precision of selected reaction monitoring. Here, we evaluate, whether SWATH-MS type DIA effectively translates into a better protein profiling as compared with the established shotgun proteomics.We implemented a novel DIA method on the widely used Orbitrap platform and used retention-time-normalized (iRT) spectral libraries for targeted data extraction using Spectronaut. We call this combination hyper reaction monitoring (HRM). Using a controlled sample set, we show that HRM outperformed shotgun proteomics both in the number of consistently identified peptides across multiple measurements and quantification of differentially abundant proteins. The reproducibility of HRM in peptide detection was above 98%, resulting in quasi complete data sets compared with 49% of shotgun proteomics.Utilizing HRM, we profiled acetaminophen (APAP)¹-treated three-dimensional human liver microtissues. An early onset of relevant proteome changes was revealed at subtoxic doses of APAP. Further, we detected and quantified for the first time human NAPQI-protein adducts that might be relevant for the toxicity of APAP. The adducts were identified on four mitochondrial oxidative stress related proteins (GATM, PARK7, PRDX6, and VDAC2) and two other proteins (ANXA2 and FTCD).Our findings imply that DIA should be the preferred method for quantitative protein profiling.Quantitative mass spectrometry is a powerful and widely used approach to identify differentially abundant proteins, e.g. for proteome profiling and biomarker discovery (1). Several tens of thousands of peptides and thousands of proteins can be routinely identified from a single sample injection in shotgun proteomics (2). Shotgun proteomics, however, is limited by low analytical reproducibility. This is due to the complexity of the samples that results in under sampling (supplemental Fig. 1) and to the fact that the acquisition of MS2 spectra is often triggered outside of the elution peak apex. As a result, only 17% of the detectable peptides are typically fragmented, and less than 60% of those are identified. This translates in reliable identification of only 10% of the detectable peptides (3). The overlap of peptide identification across technical replicates is typically 35–60% (4), which results in inconsistent peptide quantification. Alternatively to shotgun proteomics, selected reaction monitoring (SRM) enables quantification of up to 200–300 peptides at very high reproducibility, accuracy, and precision (5–8).Data-independent acquisition (DIA), a novel acquisition type, overcomes the semistochastic nature of shotgun proteomics (9–18). Spectra are acquired according to a predefined schema instead of dependent on the data. Targeted analysis of DIA data was introduced with SWATH-MS (19). For the originally published SWATH-MS, the mass spectrometer cycles through 32 predefined, contiguous, 25 Thomson wide precursor windows, and records high-resolution fragment ion spectra (19). This results in a comprehensive measurement of all detectable precursors of the selected mass range. The main novelty of SWATH-MS was in the analysis of the collected DIA data. Predefined fragment ions are extracted using precompiled spectrum libraries, which results in SRM-like data. Such targeted analyses are now enabled by several publicly available computational tools, in particular Spectronaut², Skyline (20), and OpenSWATH (21). The accuracy of peptide identification is evaluated based on the mProphet method (22).We introduce a novel SWATH-MS-type DIA workflow termed hyper reaction monitoring (HRM) (reviewed in (23)) implemented on a Thermo Scientific Q Exactive platform. It consists of comprehensive DIA acquisition and targeted data analysis with retention-time-normalized spectral libraries (24). Its high accuracy of peptide identification and quantification is due to three aspects. First, we developed a novel, improved DIA method. Second, we reimplemented the mProphet (22) approach in the software Spectronaut (www.spectronaut.org). Third, we developed large, optimized, and retention-time-normalized (iRT) spectral libraries.We compared HRM and state-of-the-art shotgun proteomics in terms of ability to discover differentially abundant proteins. For this purpose, we used a “profiling standard sample set” with 12 non-human proteins spiked at known absolute concentrations into a stable human cell line protein extract. This resulted in quasi complete data sets for HRM and the detection of a larger number of differentially abundant proteins as compared with shotgun proteomics. We utilized HRM to identify changes in the proteome in primary three-dimensional human liver microtissues after APAP exposure (25–27). These primary hepatocytes exhibit active drug metabolism. With a starting material of only 12,000 cells per sample, the abundance of 2,830 proteins was quantified over an APAP concentration range. Six novel NAPQI-cysteine proteins adducts that might be relevant for the toxicity of APAP were found and quantified mainly on mitochondrion-related proteins.