Mysterious inhibitory cell regulator investigated and found likely to be secretogranin II related
In the context of a hunt for a postulated hormone that is tissue-mass inhibiting and reproductively associated, there is described probable relatedness to a granin protein. A 7–8 kDa polypeptide candidate (gels/MS) appeared in a bioassay-guided fractionation campaign involving sheep plasma. An N-terminal sequence of 14 amino acids was obtained for the polypeptide by Edman degradation. Bioinformatics and molecular biology failed to illuminate any ovine or non-ovine protein which might relate to this sequence. The N-terminal sequence was synthesized as the 14mer EPL001 peptide and surprisingly found to be inhibitory in an assay in vivo of compensatory renal growth in the rat and modulatory of nematode fecundity, in line with the inhibitory hormone hypothesis. Antibodies were raised to EPL001 and their deployment upheld the hypothesis that the EPL001 amino acid sequence is meaningful and relevant, notwithstanding bioinformatic obscurity. Immunohistochemistry (IHC) in sheep, rodents and humans yielded staining of seeming endocrine relevance (e.g. hypothalamus, gonads and neuroendocrine cells in diverse tissues), with apparent upregulation in certain human tumours (e.g. pheochromocytoma). Discrete IHC staining in Drosophila melanogaster embryo brain was seen in glia and in neuroendocrine cells, with staining likely in the corpus cardiacum. The search for the endogenous antigen involved immunoprecipitation (IP) followed by liquid chromatography and mass spectrometry (LC–MS). Feedstocks were PC12 conditioned medium and aqueous extract of rat hypothalamus—both of which had anti-proliferative and pro-apoptotic effects in an assay in vitro involving rat bone marrow cells, which inhibition was subject to prior immunodepletion with an anti-EPL001 antibody—together with fruit fly embryo material. It is concluded that the mammalian antigen is likely secretogranin II (SgII) related. The originally seen 7–8 kDa polypeptide is suggested to be a new proteoform of secretogranin II of ∼70 residues, SgII-70, with the anti-EPL001 antibody seeing a discontinuous epitope. The fly antigen is probably Q9W2X8 (UniProt), an uncharacterised protein newly disclosed as a granin and provisionally dubbed macrogranin I (MgI). SgII and Q9W2X8 merit further investigation in the context of tissue-mass inhibition.
Hart et al. (2017) Mysterious inhibitory cell regulator investigated and found likely to be secretogranin II related. PeerJ 5:e3833 https://doi.org/10.7717/peerj.3833
The intervals method: a new approach to analyse finite element outputs using multivariate statistics
Background In this paper, we propose a new method, named the intervals’ method, to analyse data from finite element models in a comparative multivariate framework. As a case study, several armadillo mandibles are analysed, showing that the proposed method is useful to distinguish and characterise biomechanical differences related to diet/ecomorphology. Methods The intervals’ method consists of generating a set of variables, each one defined by an interval of stress values. Each variable is expressed as a percentage of the area of the mandible occupied by those stress values. Afterwards these newly generated variables can be analysed using multivariate methods. Results Applying this novel method to the biological case study of whether armadillo mandibles differ according to dietary groups, we show that the intervals’ method is a powerful tool to characterize biomechanical performance and how this relates to different diets. This allows us to positively discriminate between specialist and generalist species. Discussion We show that the proposed approach is a useful methodology not affected by the characteristics of the finite element mesh. Additionally, the positive discriminating results obtained when analysing a difficult case study suggest that the proposed method could be a very useful tool for comparative studies in finite element analysis using multivariate statistical approaches.
Marcé-Nogué et al. (2017) The intervals method: a new approach to analyse finite element outputs using multivariate statistics. PeerJ 5:e3793 https://doi.org/10.7717/peerj.3793
Sediment microbial taxonomic and functional diversity in a natural salinity gradient challenge Remane’s “species minimum” concept
Several models have been developed for the description of diversity in estuaries and other brackish habitats, with the most recognized being Remane’s Artenminimum (“species minimum”) concept. It was developed for the Baltic Sea, one of the world’s largest semi-enclosed brackish water body with a unique permanent salinity gradient, and it argues that taxonomic diversity of macrobenthic organisms is lowest within the horohalinicum (5 to 8 psu). The aim of the present study was to investigate the relationship between salinity and sediment microbial diversity at a freshwater-marine transect in Amvrakikos Gulf (Ionian Sea, Western Greece) and assess whether species composition and community function follow a generalized concept such as Remane’s. DNA was extracted from sediment samples from six stations along the aforementioned transect and sequenced for the 16S rRNA gene using high-throughput sequencing. The metabolic functions of the OTUs were predicted and the most abundant metabolic pathways were extracted. Key abiotic variables, i.e., salinity, temperature, chlorophyll-a and oxygen concentration etc., were measured and their relation with diversity and functional patterns was explored. Microbial communities were found to differ in the three habitats examined (river, lagoon and sea) with certain taxonomic groups being more abundant in the freshwater and less in the marine environment, and vice versa. Salinity was the environmental factor with the highest correlation to the microbial community pattern, while oxygen concentration was highly correlated to the metabolic functional pattern. The total number of OTUs showed a negative relationship with increasing salinity, thus the sediment microbial OTUs in this study area do not follow Remane’s concept.
Pavloudi et al. (2017) Sediment microbial taxonomic and functional diversity in a natural salinity gradient challenge Remane’s “species minimum” concept. PeerJ 5:e3687 https://doi.org/10.7717/peerj.3687
Evaluation of plasma cytokines in patients with cocaine use disorders in abstinence identifies transforming growth factor alpha (TGFα) as a potential biomarker of consumption and dual diagnosis
Background Cocaine use disorder (CUD) is a complex health condition, especially when it is accompanied by comorbid psychiatric disorders (dual diagnosis). Dual diagnosis is associated with difficulties in the stratification and treatment of patients. One of the major challenges in clinical practice of addiction psychiatry is the lack of objective biological markers that indicate the degree of consumption, severity of addiction, level of toxicity and response to treatment in patients with CUD. These potential biomarkers would be fundamental players in the diagnosis, stratification, prognosis and therapeutic orientation in addiction. Due to growing evidence of the involvement of the immune system in addiction and psychiatric disorders, we tested the hypothesis that patients with CUD in abstinence might have altered circulating levels of signaling proteins related to systemic inflammation. Methods The study was designed as a cross-sectional study of CUD treatment-seeking patients. These patients were recruited from outpatient programs in the province of Malaga (Spain). The study was performed with a total of 160 white Caucasian subjects, who were divided into the following groups: patients diagnosed with CUD in abstinence (N = 79, cocaine group) and matched control subjects (N = 81, control group). Participants were clinically evaluated with the diagnostic interview PRISM according to the DSM-IV-TR, and blood samples were collected for the determination of chemokine C-C motif ligand 11 (CCL11, eotaxin-1), interferon gamma (IFNγ), interleukin-4 (IL-4), interleukin-8 (IL-8), interleukin-17α (IL-17α), macrophage inflammatory protein 1α (MIP-1α) and transforming growth factor α (TGFα) levels in the plasma. Clinical and biochemical data were analyzed in order to find relationships between variables. Results While 57% of patients with CUD were diagnosed with dual diagnosis, approximately 73% of patients had other substance use disorders. Cocaine patients displayed greater cocaine symptom severity when they were diagnosed with psychiatric comorbidity. Regarding inflammatory factors, we observed significantly lower plasma levels of IL-17α (p < 0.001), MIP-1α (p < 0.001) and TGFα (p < 0.05) in the cocaine group compared with the levels in the control group. Finally, there was a significant primary effect of dual diagnosis on the plasma concentrations of TGFα (p < 0.05) in the cocaine group, and these levels were lower in patients with dual diagnoses Discussion IL-17α, MIP-1α and TGFα levels are different between the cocaine and control groups, and TGFα levels facilitate the identification of patients with dual diagnosis. Because TGFα reduction is associated with enhanced responses to cocaine in preclinical models, we propose TGFα as a potential biomarker of complex CUD in humans.
Maza-Quiroga et al. (2017) Evaluation of plasma cytokines in patients with cocaine use disorders in abstinence identifies transforming growth factor alpha (TGFα) as a potential biomarker of consumption and dual diagnosis. PeerJ 5:e3926 https://doi.org/10.7717/peerj.3926
External validation of cut-off points for foveal thickness taking into account the intraretinal fluid using optical coherence tomography to diagnose diabetic macular oedema
Background In late 2015, cut-off points were published for foveal thickness to diagnose diabetic macular oedema taking into account the presence of intraretinal fluid using optical coherence tomography (OCT) in primary care patients (90 µm in the presence of intraretinal fluid and 310 µm otherwise). Methods This cross-sectional observational study was carried out on 134 eyes of diabetic patients treated in specialised ophthalmology services in a Spanish region in 2012–2013, to externally validate the aforementioned cut-off points. The main variable (Clinical Standard) was the diagnosis of macular oedema through indirect ophthalmoscopy and posterior segment slit-lamp biomicroscopy. As validation variables, both the foveal thickness and the presence of intraretinal fluid obtained by OCT were used. Validation was performed using bootstrapping by calculating the area under the ROC curve (AUC), sensitivity, specificity, positive likelihood ratio (PLR) and negative likelihood ratio (NLR). Results Forty-one eyes presented diabetic macular oedema (30.6%). The bootstrapping validation parameters were: AUC, 0.88; sensitivity, 0.75; specificity, 0.95; PLR, 14.31; NLR, 0.26. These values were very similar to those of the original publication. Conclusion We have externally validated in specialised care patients the cut-off points published for the diagnosis of diabetic macular oedema. We suggest that others carry out validation studies in their communities.
Hernández-Martínez et al. (2017) External validation of cut-off points for foveal thickness taking into account the intraretinal fluid using optical coherence tomography to diagnose diabetic macular oedema. PeerJ 5:e3922 https://doi.org/10.7717/peerj.3922
The complete chloroplast genome sequence of strawberry (Fragaria × ananassa Duch.) and comparison with related species of Rosaceae
Compared with other members of the family Rosaceae, the chloroplast genomes of Fragaria species exhibit low variation, and this situation has limited phylogenetic analyses; thus, complete chloroplast genome sequencing of Fragaria species is needed. In this study, we sequenced the complete chloroplast genome of F. × ananassa ‘Benihoppe’ using the Illumina HiSeq 2500-PE150 platform and then performed a combination of de novo assembly and reference-guided mapping of contigs to generate complete chloroplast genome sequences. The chloroplast genome exhibits a typical quadripartite structure with a pair of inverted repeats (IRs, 25,936 bp) separated by large (LSC, 85,531 bp) and small (SSC, 18,146 bp) single-copy (SC) regions. The length of the F. × ananassa ‘Benihoppe’ chloroplast genome is 155,549 bp, representing the smallest Fragaria chloroplast genome observed to date. The genome encodes 112 unique genes, comprising 78 protein-coding genes, 30 tRNA genes and four rRNA genes. Comparative analysis of the overall nucleotide sequence identity among ten complete chloroplast genomes confirmed that for both coding and non-coding regions in Rosaceae, SC regions exhibit higher sequence variation than IRs. The Ka/Ks ratio of most genes was less than 1, suggesting that most genes are under purifying selection. Moreover, the mVISTA results also showed a high degree of conservation in genome structure, gene order and gene content in Fragaria, particularly among three octoploid strawberries which were F. × ananassa ‘Benihoppe’, F. chiloensis (GP33) and F. virginiana (O477). However, when the sequences of the coding and non-coding regions of F. × ananassa ‘Benihoppe’ were compared in detail with those of F. chiloensis (GP33) and F. virginiana (O477), a number of SNPs and InDels were revealed by MEGA 7. Six non-coding regions (trnK-matK, trnS-trnG, atpF-atpH, trnC-petN, trnT-psbD and trnP-psaJ) with a percentage of variable sites greater than 1% and no less than five parsimony-informative sites were identified and may be useful for phylogenetic analysis of the genus Fragaria.
Cheng et al. (2017) The complete chloroplast genome sequence of strawberry (Fragaria × ananassa Duch.) and comparison with related species of Rosaceae. PeerJ 5:e3919 https://doi.org/10.7717/peerj.3919
Mouse spermatozoa with higher fertilization rates have thinner nuclei
Background Although spermatozoa with normal morphology are assumed to have uniform fertilization ability, recent data show that even normal spermatozoa have considerable variation in their head shape which is associated with differences in fertilization ability. Appropriate quantitative indicators for good sperm morphology, however, remain unidentified. Methods Therefore, in an effort to identify such an indicator, we compared the nuclear contour of normal mouse spermatozoa by quantitative multivariate analysis using elliptic Fourier descriptors combined with principal component analysis. The spermatozoa were obtained from different strains and collection sites which have been shown to be associated with different fertilization abilities. Results We found that the head was 5.7% thinner in spermatozoa from the B6D2F1 (BDF1) strain, known to have a higher fertilization rate, than in those from the C57BL/6N (B6N) strain, which has a lower fertilization rate. Moreover, zona-penetrated spermatozoa in the perivitelline space consistently had 5.4% thinner heads than those isolated from the epididymis before ejaculation. The aspect ratio, which represents the sperm head thinness, uniquely distinguished these sperm populations, confirming its validity as a morphological indicator. Discussion Because aspect ratio has also been shown to characterize human spermatozoa, this unique morphometric indicator might be applicable to compare normal spermatozoa among multiple patients, which will greatly facilitate and enhance current reproductive technologies.
Mashiko et al. (2017) Mouse spermatozoa with higher fertilization rates have thinner nuclei. PeerJ 5:e3913 https://doi.org/10.7717/peerj.3913
Cellobiohydrolase B of Aspergillus niger over-expressed in Pichia pastoris stimulates hydrolysis of oil palm empty fruit bunches
Background Aspergillus niger, along with many other lignocellulolytic fungi, has been widely used as a commercial workhorse for cellulase production. A fungal cellulase system generally includes three major classes of enzymes i.e., β-glucosidases, endoglucanases and cellobiohydrolases. Cellobiohydrolases (CBH) are vital to the degradation of crystalline cellulose present in lignocellulosic biomass. However, A. niger naturally secretes low levels of CBH. Hence, recombinant production of A. niger CBH is desirable to increase CBH production yield and also to allow biochemical characterisation of the recombinant CBH from A. niger. Methods In this study, the gene encoding a cellobiohydrolase B (cbhB) from A. niger ATCC 10574 was cloned and expressed in the methylotrophic yeast Pichia pastoris X-33. The recombinant CBHB was purified and characterised to study its biochemical and kinetic characteristics. To evaluate the potential of CBHB in assisting biomass conversion, CBHB was supplemented into a commercial cellulase preparation (Cellic® CTec2) and was used to hydrolyse oil palm empty fruit bunch (OPEFB), one of the most abundant lignocellulosic waste from the palm oil industry. To attain maximum saccharification, enzyme loadings were optimised by response surface methodology and the optimum point was validated experimentally. Hydrolysed OPEFB samples were analysed using attenuated total reflectance FTIR spectroscopy (ATR-FTIR) to screen for any compositional changes upon enzymatic treatment. Results Recombinant CBHB was over-expressed as a hyperglycosylated protein attached to N-glycans. CBHB was enzymatically active towards soluble substrates such as 4-methylumbelliferyl-β-D-cellobioside (MUC), p-nitrophenyl-cellobioside (pNPC) and p-nitrophenyl-cellobiotrioside (pNPG3) but was not active towards crystalline substrates like Avicel® and Sigmacell cellulose. Characterisation of purified CBHB using MUC as the model substrate revealed that optimum catalysis occurred at 50 °C and pH 4 but the enzyme was stable between pH 3 to 10 and 30 to 80 °C. Although CBHB on its own was unable to digest crystalline substrates, supplementation of CBHB (0.37%) with Cellic® CTec2 (30%) increased saccharification of OPEFB by 27%. Compositional analyses of the treated OPEFB samples revealed that CBHB supplementation reduced peak intensities of both crystalline cellulose Iα and Iβ in the treated OPEFB samples. Discussion Since CBHB alone was inactive against crystalline cellulose, these data suggested that it might work synergistically with other components of Cellic® CTec2. CBHB supplements were desirable as they further increased hydrolysis of OPEFB when the performance of Cellic® CTec2 was theoretically capped at an enzyme loading of 34% in this study. Hence, A. niger CBHB was identified as a potential supplementary enzyme for the enzymatic hydrolysis of OPEFB.
Woon et al. (2017) Cellobiohydrolase B of Aspergillus niger over-expressed in Pichia pastoris stimulates hydrolysis of oil palm empty fruit bunches. PeerJ 5:e3909 https://doi.org/10.7717/peerj.3909
Nests of red wood ants (Formica rufa-group) are positively associated with tectonic faults: a double-blind test
Ecological studies often are subjected to unintentional biases, suggesting that improved research designs for hypothesis testing should be used. Double-blind ecological studies are rare but necessary to minimize sampling biases and omission errors, and improve the reliability of research. We used a double-blind design to evaluate associations between nests of red wood ants (Formica rufa, RWA) and the distribution of tectonic faults. We randomly sampled two regions in western Denmark to map the spatial distribution of RWA nests. We then calculated nest proximity to the nearest active tectonic faults. Red wood ant nests were eight times more likely to be found within 60 m of known tectonic faults than were random points in the same region but without nests. This pattern paralleled the directionality of the fault system, with NNE–SSW faults having the strongest associations with RWA nests. The nest locations were collected without knowledge of the spatial distribution of active faults thus we are confident that the results are neither biased nor artefactual. This example highlights the benefits of double-blind designs in reducing sampling biases, testing controversial hypotheses, and increasing the reliability of the conclusions of research.
Del Toro et al. (2017) Nests of red wood ants (Formica rufa-group) are positively associated with tectonic faults: a double-blind test. PeerJ 5:e3903 https://doi.org/10.7717/peerj.3903
Seasonal and year-round use of the Kushiro Wetland, Hokkaido, Japan by sika deer (Cervus nippon yesoensis)
The sika deer (Cervus nippon yesoensis) population in the Ramsar-listed Kushiro Wetland has increased in recent years, and the Ministry of the Environment of Japan has decided to take measures to reduce the impact of deer on the ecosystem. However, seasonal movement patterns of the deer (i.e., when and where the deer inhabit the wetland) remain unclear. We examined the seasonal movement patterns of sika deer in the Kushiro Wetland from 2013 to 2015 by analyzing GPS location data for 28 hinds captured at three sites in the wetland. Seasonal movement patterns were quantitatively classified as seasonal migration, mixed, dispersal, nomadic, resident, or atypical, and the degree of wetland utilization for each individual was estimated. The area of overlap for each individual among intra-capture sites and inter-capture sites was calculated for the entire year and for each season. Our results showed that the movement patterns of these deer were classified not only as resident but also as seasonal migration, dispersal, and atypical. Approximately one-third of the individuals moved into and out of the wetland during the year as either seasonal migrants or individuals with atypical movement. Some of the individuals migrated to farmland areas outside the wetland (the farthest being 69.9 km away). Half of the individuals inhabited the wetland all or most of the year, i.e., 81–100% of their annual home range was within the wetland area. Even among individuals captured at the same site, different seasonal movement patterns were identified. The overlap areas of the home ranges of individuals from the same capture sites were larger than those for individuals from different capture sites (e.g., mean of annual home range overlap with intra-capture sites: 47.7% vs. inter-sites: 1.3%). To achieve more effective ecosystem management including deer management in the wetland, management plans should cover inside and outside of the wetland and separate the population into multiple management units to address the different movement patterns and wetland utilization of the population.
Takafumi et al. (2017) Seasonal and year-round use of the Kushiro Wetland, Hokkaido, Japan by sika deer (Cervus nippon yesoensis). PeerJ 5:e3869 https://doi.org/10.7717/peerj.3869