Small proteins in bacteria – Big challenges in prediction and identification

Proteins with up to 100 amino acids have been largely overlooked due to the challenges associated with predicting and identifying them using traditional methods. Recent advances in bioinformatics and machine learning, DNA sequencing, RNA and Ribo-seq technologies, and mass spectrometry (MS) have greatly facilitated the detection and characterisation of these elusive proteins in recent years. This has revealed their crucial role in various cellular processes including regulation, signalling and transport, as toxins and as folding helpers for protein complexes. Consequently, the systematic identification and characterisation of these proteins in bacteria have emerged as a prominent field of interest within the microbial research community. This review provides an overview of different strategies for predicting and identifying these proteins on a large scale, leveraging the power of these advanced technologies. Furthermore, the review offers insights into the future developments that may be expected in this field.     

Acetylcholinesterases – the structural similarities and differences

Acetylcholinesterase (AChE) is a widely spread enzyme playing a very important role in nerve signal transmission. As AChE controls key processes, its inhibition leads to the very fast death of an organism, including humans. However, when this feature is to be used for killing of unwanted organisms (i.e. mosquitoes), one is faced with the question - how much do AChEs differ between species and what are the differences? Here, a theoretical point of view was utilized to identify the structural basis for such differences. The various primary and tertiary alignments show that AChEs are very evolutionary conserved enzymes and this fact could lead to difficulties, for example, in the search for inhibitors specific for a particular species.     

Root taxa identification in plant mixtures – current techniques and future challenges

Background

Studying root biomass, root system distribution and belowground interactions is essential for understanding the composition of plant communities, the impact of global change, and terrestrial biogeochemistry. Most soil samples and minirhizotron pictures hold roots of more than one species or plant individual. The identification of taxa by their roots would allow species-specific questions to be posed; information about root affiliation to plant individuals could be used to determine intra-specific competition.

Scope

Researchers need to be able to discern plant taxa by roots as well as to quantify abundances in mixed root samples. However, roots show less distinctive features that permit identification than aboveground organs. This review discusses the primary use of available methods, outlining applications, shortcomings and future developments.

Conclusion

Methods are either non-destructive, e.g. visual examination of root morphological criteria in situ, or require excavated and excised root samples. Among the destructive methods are anatomical keys, chemotaxonomic approaches and molecular markers. While some methods allow for discerning the root systems of individual plants, others can distinguish roots on the functional group or plant taxa level; methods such as IR spectroscopy and qPCR allow for quantifying the root biomass proportion of species without manual sorting.     

Two-component systems in Arabidopsis thaliana – A structural view

Plants respond to outside stimuli by initiating signaling cascades that regulate gene expression. Little structural information is available on the signaling proteins that are part of the two-component systems of plants, and how changes in phosphorylation translate into alterations of three-dimensional structures and changes in recognition domains remains largely mysterious. Our work is on deciphering aspects of these systems through the crystallization and structural analysis of two-component system proteins in Arabidopsis thaliana.     

Relationship between homoeologous regulatory and structural genes in allopolyploid genome – A case study in bread wheat

Background

Whole-genome physical maps facilitate genome sequencing, sequence assembly, mapping of candidate genes, and the design of targeted genetic markers. An automated protocol was used to construct a Vitis vinifera 'Cabernet Sauvignon' physical map. The quality of the result was addressed with regard to the effect of high heterozygosity on the accuracy of contig assembly. Its usefulness for the genome-wide mapping of genes for disease resistance, which is an important trait for grapevine, was then assessed.

Results

The physical map included 29,727 BAC clones assembled into 1,770 contigs, spanning 715,684 kbp, and corresponding to 1.5-fold the genome size. Map inflation was due to high heterozygosity, which caused either the separation of allelic BACs in two different contigs, or local mis-assembly in contigs containing BACs from the two haplotypes. Genetic markers anchored 395 contigs or 255,476 kbp to chromosomes. The fully automated assembly and anchorage procedures were validated by BAC-by-BAC blast of the end sequences against the grape genome sequence, unveiling 7.3% of chimerical contigs. The distribution across the physical map of candidate genes for non-host and host resistance, and for defence signalling pathways was then studied. NBS-LRR and RLK genes for host resistance were found in 424 contigs, 133 of them (32%) were assigned to chromosomes, on which they are mostly organised in clusters. Non-host and defence signalling genes were found in 99 contigs dispersed without a discernable pattern across the genome.

Conclusion

Despite some limitations that interfere with the correct assembly of heterozygous clones into contigs, the 'Cabernet Sauvignon' physical map is a useful and reliable intermediary step between a genetic map and the genome sequence. This tool was successfully exploited for a quick mapping of complex families of genes, and it strengthened previous clues of co-localisation of major NBS-LRR clusters and disease resistance loci in grapevine.     

Live bacterial vaccines – a review and identification of potential hazards

The use of live bacteria to induce an immune response to itself or to a carried vaccine component is an attractive vaccine strategy. Advantages of live bacterial vaccines include their mimicry of a natural infection, intrinsic adjuvant properties and their possibility to be administered orally. Derivatives of pathogenic and non-pathogenic food related bacteria are currently being evaluated as live vaccines. However, pathogenic bacteria demands for attenuation to weaken its virulence. The use of bacteria as vaccine delivery vehicles implies construction of recombinant strains that contain the gene cassette encoding the antigen. With the increased knowledge of mucosal immunity and the availability of genetic tools for heterologous gene expression the concept of live vaccine vehicles gains renewed interest. However, administration of live bacterial vaccines poses some risks. In addition, vaccination using recombinant bacteria results in the release of live recombinant organisms into nature. This places these vaccines in the debate on application of genetically modified organisms. In this review we give an overview of live bacterial vaccines on the market and describe the development of new live vaccines with a focus on attenuated bacteria and food-related lactic acid bacteria. Furthermore, we outline the safety concerns and identify the hazards associated with live bacterial vaccines and try to give some suggestions of what to consider during their development.     

StePSIM – a method for stepwise peak selection and identification of metabolites in 1 H NMR spectra

A method for stepwise selection of peaks in NMR spectra from multiple groups is described. This method is based on initial peak-finding among the spectra and uses jacknife classification performance as the basis for selection of peaks. The selection process is followed by the construction of correlation maps to identify sets of multiplets that are related to each of the selected peaks, aiding in the identification of metabolites that are responsible for differences among the groups. For illustrative purposes, this methodology is applied to a data set that contains 52 spectra from renal cell carcinoma and normal renal tissue samples. The new method is denoted as StePSIM, Stepwise Peak Selection and Identification of Metabolites. Research partially supported by NCI 1 R21 CA89671-01A1 and NIH NCRR 02584     

The identification of wingless Betula fruits in Weichselian sediments in the Gross Todtshorn borehole (Lower Saxony, Germany) – the occurrence of Betula humilis Schrank.

Betula fruits lacking their wings were recovered from the Gross Todtshorn borehole, Germany, covering the late Eemian interglacial, the Early Weichselian, and the Weichselian pleniglacial. In order to identify them, a data-set of seven size and shape variables was made from modern fruits of B. pubescens, B. pendula, B. nana, and B. humilis. These measurements were summarised by principal components analysis (PCA). The 140 measured fossil fruit-bodies were positioned passively on the PCA plot and identified by comparison with the dimensional envelopes of the modern species. Tree birches (B. pendula, B. pubescens) occurred in the Eemian and Early Weichselian interstadials, but not thereafter.B. nana was widespread during the intervening stadials and the Oerel interstadial. B. humilis occurred throughout the sequence, coexisting with tree birches in the Early Weichselian and with B. nana in the stadials. It usually grew on mire surfaces, in contrast to B. nana which mainly grew on mineral substrates. However, B. nana grew on peat with B. humilis in the pleniglacial Oerel interstadial, suggesting a more continental climate with cold winters. A modern analogue could be near the Ural mountains. B. humilis is presently rare in central Europe, but during the Weichselian it was well established in northern Germany. The PCA has enabled specific determination of the wingless Betula fruits, leading to vegetational and climatic reconstructions and contributing to the previously poorly known historical biogeography of Betula humilis. Received June 14, 2000 / Accepted January 2, 2001     

Pre-molecular identification – Ignorance was Bliss?

Use of molecular techniques to characterize microorganisms during the past 20 years has increased the numbers of anaerobic species and made identification using only phenotypic methods difficult. Some of the newly described species have been recovered from blood cultures, showing pathogenic potential, and posing a challenge for identification.     

Nectar resorption in the spur of Platanthera chlorantha Custer (Rchb.) Orchidaceae – structural and microautoradiographic study

 In the flowers of Platanthera chlorantha nectar is secreted and accumulated in a spur. Previous studies of this species revealed that after the period of secretion and cessation, rapid nectar resorption occurs. The aim of this study was the observation of nectar resorption on the structural level using tritium labelled sucrose. For this purpose, during the peak of nectar secretion, 10 μl of nectar accumulated in the spur was replaced with the same volume of labelled sucrose (10 μCi). Small fragments of spurs were sampled between 12–36 h of incubation, at the resorption phase. Afterwards, they were fixed and embedded in epoxy resin. Material for the microautoradiographic study was prepared with the dipping technique. The cells of secretory epidermis and unicellular secretory papillae had dense, strong stained cytoplasm, a large nucleus and small vacuoles. A characteristic feature of these cells was the presence of numerous starchless plastids, mitochondria and ER profiles. Many vesicles occurred in the close vicinity of the cell wall. In the cuticle covering cell walls no pores or cracks were observed. Presence of [³H] sucrose was detected mainly in the walls of the nectary cells, which would indicate an apoplastic route of resorbed nectar. Received August 3, 2002; accepted November 10, 2002 Published online: June 2, 2003     

PseudoGeneQuest – Service for identification of different pseudogene types in the human genome

Background  

Pseudogenes, nonfunctional copies of genes, evolve fast due the lack of evolutionary pressures and thus appear in several different forms. PseudoGeneQuest is an online tool to search the human genome for a given query sequence and to identify different types of pseudogenes as well as novel genes and gene fragments.     

Type A-trichothecenes — Quantitative analysis using LC-MS and occurrence in Austrian maize and oats

During this study a method was developed for the quantitative determination of diacetoxyscirpenol (DAS), T-2 toxin and HT-2 toxin using deuterated T-2 toxin (T-2 d3 toxin) as an internal standard. The described method involves a clean up step of maize extract by the use of Mycosep® 227 columns a chromatographic separation on a Zorbax® bonus-RP-column (2,1×150mm) and the detection and quantification step on a mass spectrometer in SIM (Selected Ion Monitoring) mode.Data on the occurrence of three type A trichothecenes in Austrian maize, maize silage and oats were collected. It could be shown that maize and silage samples harvested in 2002 were only contaminated to a small extent with T-2 toxin (8% of the maize, 0% of the silage) and with HT-2 toxin (30% of the maize, 18% of the silage). But most of the analysed oats samples showed significant levels of T-2 toxin (64%) or HT-2 toxin (82%).     

Review: SARS-CoV-2 infection in farmed minks – an overview of current knowledge on occurrence,disease and epidemiology

Coronaviruses (CoVs), which are enveloped, positive-sense RNA viruses, may cause infections in mammals and birds. Apart from the respiratory manifestations, CoVs are also responsible for infections of the gastrointestinal tract and nervous systems. Their propensity to recombine allows them to easily transmit and adapt to new hosts. The emergence of a new CoV in humans, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which is attributed to a zoonotic origin, has provoked numerous studies to assess its pathogenicity for different animal species (pets, farm and wild animals). Available results indicate that numerous animal species are susceptible to infection with SARS-CoV-2. From April 2020, when the first SARS-CoV-2 infection in minks was reported in the Netherlands, to the end of January 2021, further outbreaks have been confirmed in Denmark, Italy, Spain, Sweden, the United States, Greece, France, Canada, Lithuania and Poland. It has also been established that human-to-minks and minks-to-human transmission may occur. The results obtained to date indicate that the virus was originally introduced into the minks population by humans, possibly at the start of the pandemic and had been circulating in the population for several weeks before detection. Recent data indicate that minks are highly susceptible to SARS-CoV-2 infection, but the route or routes of virus transmission between farms, other than by direct contact with infected humans, have not been identified. In minks, infection can occur in clinical and subclinical form, making it possibly difficult to detect. Therefore, minks could represent potentially dangerous, not always recognized, animal reservoir for SARS-CoV-2. The current data indicate that further studies on minks and other Mustelidae are needed to clarify whether they may be a potential reservoir for SARS-CoV-2, and if so, how and whether this can be prevented.     

Micro- and Nanofluidics – Applications in Biotechnology

Cover illustration: Micro- and Nanofluidics – Applications in Biotechnology. This issue of BTJ edited by Hikmet Geckil and Utkan Demirci covers recent advances on BioMEMs (microelectromechanical systems used in biology) and lab-on-a-chip devices for cell and fluid manipulation, e.g. picoliter sequencing and PCR, as well as cytometry and imaging technologies. The cover image provided by Gurkan Yilmaz shows fluorescent particles moving inside a spiral channel in an dielectrophoretic particle separation experiment displayed by a fluorescent microscope. DOI: http://dx.doi.org/10.1002/biot.201000204     

Potassium in agriculture – Status and perspectives

In this review we summarize factors determining the plant availability of soil potassium (K), the role of K in crop yield formation and product quality, and the dependence of crop stress resistance on K nutrition. Average soil reserves of K are generally large, but most of it is not plant-available. Therefore, crops need to be supplied with soluble K fertilizers, the demand of which is expected to increase significantly, particularly in developing regions of the world. Recent investigations have shown that organic exudates of some bacteria and plant roots play a key role in releasing otherwise unavailable K from K-bearing minerals. Thus, breeding for genotypes that have improved mechanisms to gain access to this fixed K will contribute toward more sustainable agriculture, particularly in cropping systems that do not have access to fertilizer K. In K-deficient crops, the supply of sink organs with photosynthates is impaired, and sugars accumulate in source leaves. This not only affects yield formation, but also quality parameters, for example in wheat, potato and grape. As K has beneficial effects on human health, its concentration in the harvest product is a quality parameter in itself. Owing to its fundamental roles in turgor generation, primary metabolism, and long-distance transport, K plays a prominent role in crop resistance to drought, salinity, high light, or cold as well as resistance to pests and pathogens. Despite the abundance of vital roles of K in crop production, an improvement of K uptake and use efficiency has not been a major focus of conventional or transgenic breeding in the past. In addition, current soil analysis methods for K are insufficient for some common soils, posing the risk of imbalanced fertilization. A stronger prioritization of these areas of research is needed to counter declines in soil fertility and to improve food security.     

Characterization and sequence prediction of structural variations in α-helix

Background

Centralised resources such as GenBank and UniProt are perfect examples of the major international efforts that have been made to integrate and share biological information. However, additional data that adds value to these resources needs a simple and rapid route to public access. The Distributed Annotation System (DAS) provides an adequate environment to integrate genomic and proteomic information from multiple sources, making this information accessible to the community. DAS offers a way to distribute and access information but it does not provide domain experts with the mechanisms to participate in the curation process of the available biological entities and their annotations.

Results

We designed and developed a Collaborative Annotation System for proteins called DAS Writeback. DAS writeback is a protocol extension of DAS to provide the functionalities of adding, editing and deleting annotations. We implemented this new specification as extensions of both a DAS server and a DAS client. The architecture was designed with the involvement of the DAS community and it was improved after performing usability experiments emulating a real annotation task.

Conclusions

We demonstrate that DAS Writeback is effective, usable and will provide the appropriate environment for the creation and evolution of community protein annotation.