Identifications of pathogens - a bioinformatic point of view

Over the past 15 years, microbiology has undergone a momentous shift toward molecular methods. New sequences appear daily in the public databases and new computer tools and web servers are published on a regular basis. Major advances in molecular identifications of pathogens have been made because new biotechnology methods have appeared that often require a thorough in silico analysis of sequences. However, significant difficulties partly remain in developing efficient methods because the public databases contain many poorly annotated or partial sequences (often of environmental origin) and also because there are few dedicated web servers and curated databases.     

Dissociation of Increases in PGC-1α and Its Regulators from Exercise Intensity and Muscle Activation Following Acute Exercise

Muscle activation as well as changes in peroxisome proliferator-activated receptor gamma coactivator 1 alpha (PGC-1α) following high-intensity interval exercise (HIIE) were examined in young healthy men (n  = 8; age, 21.9±2.2 yrs; VO₂peak, 53.1±6.4 ml/min/kg; peak work rate, 317±23.5 watts). On each of 3 visits HIIE was performed on a cycle ergometer at a target intensity of 73, 100, or 133% of peak work rate. Muscle biopsies were taken at rest and three hours after each exercise condition. Total work was not different between conditions (∼730 kJ) while average power output (73%, 237±21; 100%, 323±26; 133%, 384±35 watts) and EMG derived muscle activation (73%, 1262±605; 100%, 2089±737; 133%, 3029±1206 total integrated EMG per interval) increased in an intensity dependent fashion. PGC-1α mRNA was elevated after all three conditions (p<0.05), with a greater increase observed following the 100% condition (∼9 fold, p<0.05) compared to both the 73 and 133% conditions (∼4 fold). When expressed relative to muscle activation, the increase in PGC-1α mRNA for the 133% condition was less than that for the 73 and 100% conditions (p<0.05). SIRT1 mRNA was also elevated after all three conditions (∼1.4 fold, p<0.05), with no difference between conditions. These findings suggest that intensity-dependent increases in PGC-1α mRNA following submaximal exercise are largely due to increases in muscle recruitment. As well, the blunted response of PGC-1α mRNA expression following supramaximal exercise may indicate that signalling mediated activation of PGC-1α may also be blunted. We also indentify that increases in PDK4, SIRT1, and RIP140 mRNA following acute exercise are dissociated from exercise intensity and muscle activation, while increases in EGR1 are augmented with supramaximal HIIE (p<0.05).     

Global Trophic Position Comparison of Two Dominant Mesopelagic Fish Families (Myctophidae,Stomiidae) Using Amino Acid Nitrogen Isotopic Analyses

The δ¹⁵N values of organisms are commonly used across diverse ecosystems to estimate trophic position and infer trophic connectivity. We undertook a novel cross-basin comparison of trophic position in two ecologically well-characterized and different groups of dominant mid-water fish consumers using amino acid nitrogen isotope compositions. We found that trophic positions estimated from the δ¹⁵N values of individual amino acids are nearly uniform within both families of these fishes across five global regions despite great variability in bulk tissue δ¹⁵N values. Regional differences in the δ¹⁵N values of phenylalanine confirmed that bulk tissue δ¹⁵N values reflect region-specific water mass biogeochemistry controlling δ¹⁵N values at the base of the food web. Trophic positions calculated from amino acid isotopic analyses (AA-TP) for lanternfishes (family Myctophidae) (AA-TP ∼2.9) largely align with expectations from stomach content studies (TP ∼3.2), while AA-TPs for dragonfishes (family Stomiidae) (AA-TP ∼3.2) were lower than TPs derived from stomach content studies (TP∼4.1). We demonstrate that amino acid nitrogen isotope analysis can overcome shortcomings of bulk tissue isotope analysis across biogeochemically distinct systems to provide globally comparative information regarding marine food web structure.     

Two Types of Meiotic Crossovers Coexist in Maize

We apply modeling approaches to investigate the distribution of late recombination nodules in maize (Zea mays). Such nodules indicate crossover positions along the synaptonemal complex. High-quality nodule data were analyzed using two different interference models: the “statistical” gamma model and the “mechanical” beam film model. For each chromosome, we exclude at a 98% significance level the hypothesis that a single pathway underlies the formation of all crossovers, pointing to the coexistence of two types of crossing-over in maize, as was previously demonstrated in other organisms. We estimate the proportion of crossovers coming from the noninterfering pathway to range from 6 to 23% depending on the chromosome, with a cell average of ∼15%. The mean number of noninterfering crossovers per chromosome is significantly correlated with the length of the synaptonemal complex. We also quantify the intensity of interference. Finally, we develop inference tools that allow one to tackle, without much loss of power, complex crossover interference models such as the beam film. The lack of a likelihood function in such models had prevented their use for parameter estimation. This advance will allow more realistic mechanisms of crossover formation to be modeled in the future.     

Improvement in Renal Hemodynamics following Combined Angiotensin II Infusion and AT1R Blockade in Aged Female Sheep following Fetal Unilateral Nephrectomy

Renin-angiotensin system (RAS) is a powerful modulator of renal hemodynamic and fluid homeostasis. Up-regulation in components of intra-renal RAS occurs with ageing. Recently we reported that 2 year old uninephrectomised (uni-x) female sheep have low renin hypertension and reduced renal function. By 5 years of age, these uni-x sheep had augmented decrease in renal blood flow (RBF) compared to sham. We hypothesised that this decrease in RBF in 5 year old uni-x sheep was due to an up-regulation in components of the intra-renal RAS. In this study, renal responses to angiotensin II (AngII) infusion and AngII type 1 receptor (AT1R) blockade were examined in the same 5 year old sheep. We also administered AngII in the presence of losartan to increase AngII bioavailability to the AT2R in order to understand AT2R contribution to renal function in this model. Uni-x animals had significantly lower renal cortical content of renin, AngII (∼40%) and Ang 1–7 (∼60%) and reduced cortical expression of AT1R gene than sham animals. In response to both AngII infusion and AT1R blockade via losartan, renal hemodynamic responses and tubular sodium excretion were significantly attenuated in uni-x animals compared to sham. However, AngII infusion in the presence of losartan caused ∼33% increase in RBF in uni-x sheep compared to ∼14% in sham (P<0.05). This was associated with a significant decrease in renal vascular resistance in the uni-x animals (22% vs 15%, P<0.05) without any changes in systemic blood pressure. The present study shows that majority of the intra-renal RAS components are suppressed in this model of low renin hypertension. However, increasing the availability of AngII to AT2R by AT1R blockade improved renal blood flow in uni-x sheep. This suggests that manipulation of the AT2R maybe a potential therapeutic target for treatment of renal dysfunction associated with a congenital nephron deficit.     

Epidemiological Survey of Quantitative Ultrasound in Risk Assessment of Falls in Middle-Aged and Elderly People

The risk assessment of falls is important, but still unsatisfactory and time-consuming. Our objective was to assess quantitative ultrasound (QUS) in the risk assessment of falls. Our study was designed as epidemiological cross-sectional study occurring from March 2009 to February 2010 by community survey at a medical center. The participants were collected from systemic sample of 1,200 community-dwelling people (Male/Female = 524/676) 40 years old and over in Yunlin County, Mid-Taiwan. Structural questionnaires including socioeconomic status, living status, smoking and drinking habits, exercise and medical history were completed. Quantitative ultrasound (QUS) at the non-dominant distal radial area (QUS-R) and the left calcaneal area (QUS-C) were measured. The overall prevalence of falls was 19.8%. In men, the independently associated factors for falls were age (OR: 1.04; 95%CI: 1.01∼1.06), fracture history (OR: 1.89; 95%CI: 1.12∼3.19), osteoarthritis history (OR: 3.66; 95%CI: 1.15∼11.64) and speed of sound (OR: 0.99; 95%CI: 0.99∼1.00; p<0.05) by QUS-R. In women, the independently associated factors for falls were current drinking (OR: 3.54; 95%CI: 1.35∼9.31) and broadband ultrasound attenuation (OR: 0.98; 95%CI: 0.97∼0.99; p<0.01) by QUS-C. The cutoffs at -2.5< T-score<-1 derived using QUS-R (OR: 2.85; 95%CI: 1.64∼4.96; p<0.01) in men or T-score ≦-2.5 derived using QUS-C (OR: 2.72; 95%CI: 1.42∼5.21; p<0.01) in women showed an independent association with falls. The lowest T-score derived using either QUS-R or QUS-C was also revealed as an independent factor for falls in both men (OR: 2.13; 95%CI: 1.03∼4.43; p<0.05) and women (OR: 2.36; 95%CI: 1.13∼4.91; p<0.05). Conclusions: Quantitative ultrasounds, measured either at the radial or calcaneal area, are convenient tools by which to assess the risk of falls in middle-aged and elderly people.     

Comparisons of GnRH Antagonist versus GnRH Agonist Protocol in Supposed Normal Ovarian Responders Undergoing IVF: A Systematic Review and Meta-Analysis

Objective

To evaluate the effectiveness and safety of GnRH antagonist and GnRH agonist in supposed normal ovarian responders undergoing IVF.

Methods

Data from 6 databases were retrieved for this study. The RCTs of GnRH agonist and GnRH antagonist use during IVF-EF therapy for patients with supposed normal ovarian response were included. A meta-analysis was performed with Revman 5.1software.

Results

Twenty-three RCTs met the inclusion criteria. The number of stimulation days (mean difference (MD): −0.66, 95% confidence interval (CI): −1.04∼−0.27), Gn amount (MD: −2.92, 95% CI: −5.0∼−0.85), E2 values on the day of HCG (MD: −330.39, 95% CI: −510.51∼−150.26), Number of oocytes retrieved (MD: −1.33, 95% CI: −2.02∼−0.64), clinical pregnancy rate (odds ratio (OR): 0.87, 95% CI: 0.75−1.0), and ovarian hyperstimulation syndrome (OHSS) incidence (OR: 0.59, 95% CI: 0.42∼0.82) were significantly lower in GnRH antagonist protocol than GnRH agonist protocol. However, the endometrial thickness on the day of HCG (MD: −0.04, 95% CI: −0.23∼0.14), the ongoing pregnancy rate (OR: 0.87, 95% CI: 0.74∼1.03), live birth rate (OR: 0.89, 95% CI: 0.64∼1.24), miscarriage rate (OR: 1.17, 95% CI: 0.85∼1.61), and cycle cancellation rate (OR: 1.11, 95% CI: 0.90∼1.37) did not significantly differ between the 2 groups.

Conclusions

During IVF treatment for patients with supposed normal responses, the incidence of OHSS were significantly lower, whereas the ongoing pregnancy and live birth rates were similar in the GnRH antagonist compared with the standard long GnRH agonist protocols.     

Electrical Stimulation Improves Rat Muscle Dysfunction Caused by Chronic Intermittent Hypoxia-Hypercapnia via Regulation of miRNA-Related Signaling Pathways

Skeletal muscle dysfunction in chronic obstructive pulmonary disease (COPD) patients is common. Neuromuscular Electrical Stimulation (NMES) is a powerful exercise training that may relieve muscle dysfunction in COPD. This study investigated whether electrical stimulation may have atypical adaptations via activation of miRNA related pathways in counteracting COPD muscle dysfunction. Forty-eight male Sprague-Dawley rats were randomly assigned to 3 groups. With the exception of the rats in the control group, the experimental rats were exposed to chronic intermittent hypoxia-hypercapnia (CIHH) (9∼11%O₂,5.5∼6.5%CO₂) for 2 or 4 weeks. Electrical stimulation was performed immediately after each CIHH session. Following assessment of the running capacity, biopsy samples were obtained from the gastrocnemius of the rats. The miR-1, miR-133a and miR-133b levels were measured, as well as their related proteins: phosphorylation of Akt (p-AKT), PGC-1alpha (PGC-1α), histone deacetylase 4 (HDAC4) and serum response factor (SRF). Myosin heavy chainⅡa (MHCⅡa) and myosin heavy chainⅡb (MHCⅡb) were also measured to assess fiber type changes. After 2 weeks, compared with the controls, only miR-1 and miR-133a were significantly increased (p<0.05) in the exposure group. After 4 weeks, the exposure group exhibited a decreased running distance (p = 0.054) and MHCⅡa-to-MHCⅡb shift (p<0.05). PGC-1α (p = 0.051), nuclear HDAC4 (p = 0.058), HDAC4, p-AKT, PGC-1α and SRF was also significantly decreased (p<0.05). In contrast, miR-1 and miR-133a were significantly increased (p<0.05). Four weeks of electrical stimulation can partly reversed those changes, and miR-133b exhibited a transient increase after 2 weeks electrical stimulation. Our study indicate miRNAs may have roles in the response of CIHH-impaired muscle to changes during electrical stimulation.     

Structure and Function of the Escherichia coli Tol-Pal Stator Protein TolR

TolR is a 15-kDa inner membrane protein subunit of the Tol-Pal complex in Gram-negative bacteria, and its function is poorly understood. Tol-Pal is recruited to cell division sites where it is involved in maintaining the integrity of the outer membrane. TolR is related to MotB, the peptidoglycan (PG)-binding stator protein from the flagellum, suggesting it might serve a similar role in Tol-Pal. The only structure thus far reported for TolR is of the periplasmic domain from Haemophilus influenzae in which N- and C-terminal residues had been deleted (TolR(62–133), Escherichia coli numbering). H. influenzae TolR(62–133) is a symmetrical dimer with a large deep cleft at the dimer interface. Here, we present the 1.7-Å crystal structure of the intact periplasmic domain of E. coli TolR (TolR(36–142)). E. coli TolR(36–142) is also dimeric, but the architecture of the dimer is radically different from that of TolR(62–133) due to the intertwining of its N and C termini. TolR monomers are rotated ∼180° relative to each other as a result of this strand swapping, obliterating the putative PG-binding groove seen in TolR(62–133). We found that removal of the strand-swapped regions (TolR(60–133)) exposes cryptic PG binding activity that is absent in the full-length domain. We conclude that to function as a stator in the Tol-Pal complex dimeric TolR must undergo large scale structural remodeling reminiscent of that proposed for MotB, where the N- and C-terminal sequences unfold in order for the protein to both reach and bind the PG layer ∼90 Å away from the inner membrane.     

Distribution and Features of the Six Classes of Peroxiredoxins

Peroxiredoxins are cysteine-dependent peroxide reductases that group into 6 different, structurally discernable classes. In 2011, our research team reported the application of a bioinformatic approach called active site profiling to extract active site-proximal sequence segments from the 29 distinct, structurally-characterized peroxiredoxins available at the time. These extracted sequences were then used to create unique profiles for the six groups which were subsequently used to search GenBank(nr), allowing identification of ∼3500 peroxiredoxin sequences and their respective subgroups. Summarized in this minireview are the features and phylogenetic distributions of each of these peroxiredoxin subgroups; an example is also provided illustrating the use of the web accessible, searchable database known as PREX to identify subfamily-specific peroxiredoxin sequences for the organism Vitis vinifera (grape).     

Yield Trends Are Insufficient to Double Global Crop Production by 2050

Several studies have shown that global crop production needs to double by 2050 to meet the projected demands from rising population, diet shifts, and increasing biofuels consumption. Boosting crop yields to meet these rising demands, rather than clearing more land for agriculture has been highlighted as a preferred solution to meet this goal. However, we first need to understand how crop yields are changing globally, and whether we are on track to double production by 2050. Using ∼2.5 million agricultural statistics, collected for ∼13,500 political units across the world, we track four key global crops—maize, rice, wheat, and soybean—that currently produce nearly two-thirds of global agricultural calories. We find that yields in these top four crops are increasing at 1.6%, 1.0%, 0.9%, and 1.3% per year, non-compounding rates, respectively, which is less than the 2.4% per year rate required to double global production by 2050. At these rates global production in these crops would increase by ∼67%, ∼42%, ∼38%, and ∼55%, respectively, which is far below what is needed to meet projected demands in 2050. We present detailed maps to identify where rates must be increased to boost crop production and meet rising demands.     

Evolution of Plastic Transmission Strategies in Avian Malaria

Malaria parasites have been shown to adjust their life history traits to changing environmental conditions. Parasite relapses and recrudescences—marked increases in blood parasite numbers following a period when the parasite was either absent or present at very low levels in the blood, respectively—are expected to be part of such adaptive plastic strategies. Here, we first present a theoretical model that analyses the evolution of transmission strategies in fluctuating seasonal environments and we show that relapses may be adaptive if they are concomitant with the presence of mosquitoes in the vicinity of the host. We then experimentally test the hypothesis that Plasmodium parasites can respond to the presence of vectors. For this purpose, we repeatedly exposed birds infected by the avian malaria parasite Plasmodium relictum to the bites of uninfected females of its natural vector, the mosquito Culex pipiens, at three different stages of the infection: acute (∼34 days post infection), early chronic (∼122 dpi) and late chronic (∼291 dpi). We show that: (i) mosquito-exposed birds have significantly higher blood parasitaemia than control unexposed birds during the chronic stages of the infection and that (ii) this translates into significantly higher infection prevalence in the mosquito. Our results demonstrate the ability of Plasmodium relictum to maximize their transmission by adopting plastic life history strategies in response to the availability of insect vectors.     

Prediction of heterosis using genome-wide SNP-marker data: application to egg production traits in white Leghorn crosses

Prediction of heterosis has a long history with mixed success, partly due to low numbers of genetic markers and/or small data sets. We investigated the prediction of heterosis for egg number, egg weight and survival days in domestic white Leghorns, using ∼400 000 individuals from 47 crosses and allele frequencies on ∼53 000 genome-wide single nucleotide polymorphisms (SNPs). When heterosis is due to dominance, and dominance effects are independent of allele frequencies, heterosis is proportional to the squared difference in allele frequency (SDAF) between parental pure lines (not necessarily homozygous). Under these assumptions, a linear model including regression on SDAF partitions crossbred phenotypes into pure-line values and heterosis, even without pure-line phenotypes. We therefore used models where phenotypes of crossbreds were regressed on the SDAF between parental lines. Accuracy of prediction was determined using leave-one-out cross-validation. SDAF predicted heterosis for egg number and weight with an accuracy of ∼0.5, but did not predict heterosis for survival days. Heterosis predictions allowed preselection of pure lines before field-testing, saving ∼50% of field-testing cost with only 4% loss in heterosis. Accuracies from cross-validation were lower than from the model-fit, suggesting that accuracies previously reported in literature are overestimated. Cross-validation also indicated that dominance cannot fully explain heterosis. Nevertheless, the dominance model had considerable accuracy, clearly greater than that of a general/specific combining ability model. This work also showed that heterosis can be modelled even when pure-line phenotypes are unavailable. We concluded that SDAF is a useful predictor of heterosis in commercial layer breeding.     

Identification of rare alleles and their carriers using compressed se(que)nsing

Identification of rare variants by resequencing is important both for detecting novel variations and for screening individuals for known disease alleles. New technologies enable low-cost resequencing of target regions, although it is still prohibitive to test more than a few individuals. We propose a novel pooling design that enables the recovery of novel or known rare alleles and their carriers in groups of individuals. The method is based on a Compressed Sensing (CS) approach, which is general, simple and efficient. CS allows the use of generic algorithmic tools for simultaneous identification of multiple variants and their carriers. We model the experimental procedure and show via computer simulations that it enables the recovery of rare alleles and their carriers in larger groups than were possible before. Our approach can also be combined with barcoding techniques to provide a feasible solution based on current resequencing costs. For example, when targeting a small enough genomic region (∼100 bp) and using only ∼10 sequencing lanes and ∼10 distinct barcodes per lane, one recovers the identity of 4 rare allele carriers out of a population of over 4000 individuals. We demonstrate the performance of our approach over several publicly available experimental data sets.     

Ascorbate Synthesis Pathway: DUAL ROLE OF ASCORBATE IN BONE HOMEOSTASIS*

Using mouse gene knock-out models, we identify aldehyde reductase (EC 1.1.1.2, Akr1a4 (GR)) and aldose reductase (EC 1.1.1.21, Akr1b3 (AR)) as the enzymes responsible for conversion of d-glucuronate to l-gulonate, a key step in the ascorbate (ASC) synthesis pathway in mice. The gene knock-out (KO) mice show that the two enzymes, GR and AR, provide ∼85 and ∼15% of l-gulonate, respectively. GRKO/ARKO double knock-out mice are unable to synthesize ASC (>95% ASC deficit) and develop scurvy. The GRKO mice (∼85% ASC deficit) develop and grow normally when fed regular mouse chow (ASC content = 0) but suffer severe osteopenia and spontaneous fractures with stresses that increase ASC requirements, such as pregnancy or castration. Castration greatly increases osteoclast numbers and activity in GRKO mice and promotes increased bone loss as compared with wild-type controls and additionally induces proliferation of immature dysplastic osteoblasts likely because of an ASC-sensitive block(s) in early differentiation. ASC and the antioxidants pycnogenol and resveratrol block osteoclast proliferation and bone loss, but only ASC feeding restores osteoblast differentiation and prevents their dysplastic proliferation. This is the first in vivo demonstration of two independent roles for ASC as an antioxidant suppressing osteoclast activity and number as well as a cofactor promoting osteoblast differentiation. Although humans have lost the ability to synthesize ASC, our mouse models suggest the mechanisms by which suboptimal ASC availability facilitates the development of osteoporosis, which has important implications for human osteoporosis.     

Biological SOAP servers and web services provided by the public sequence data bank

A number of biological data resources (i.e. databases and data analytical tools) are searchable and usable on-line thanks to the internet and the World Wide Web (WWW) servers. The output from the web server is easy for us to browse. However, it is laborious and sometimes impossible for us to write a computer program that finds a useful data resource, sends a proper query and processes the output. It is a serious obstacle to the integration of distributed heterogeneous data resources. To solve the issue, we have implemented a SOAP (Simple Object Access Protocol) server and web services that provide a program-friendly interface. The web services are accessible at http://www.xml.nig.ac.jp/.     

Bi-Epitope SPR Surfaces: A Solution to Develop Robust Immunoassays

Surface plasmon resonance (SPR)-based immunoassays have numerous applications and require high affinity reagents for sensitive and reliable measurements. We describe a quick approach to turn low affinity antibodies into appropriate capture reagents. We used antibodies recognizing human ephrin type A receptor 2 (EphA2) and a ProteOn XPR36 as a model system. We generated so-called ‘bi-epitope’ sensor surfaces by immobilizing various pairs of anti-EphA2 antibodies using standard amine coupling. The apparent binding affinities to EphA2 and EphA2 detection sensitivities of the bi-epitope and ‘single-epitope’ surfaces were then compared. For all antibody pairs tested, bi-epitope surfaces exhibited an ∼10–100-fold improvement in apparent binding affinities when compared with single-epitope ones. When pairing 2 antibodies of low intrinsic binding affinities (∼10⁻⁸ M) and fast dissociation rates (∼10⁻² s⁻¹), the apparent binding affinity and dissociation rate of the bi-epitope surface was improved up to ∼10^–10 M and 10⁻⁴ s⁻¹, respectively. This led to an ∼100–200-fold enhancement in EphA2 limit of detection in crude cell supernatants. Our results show that the use of antibody mixtures in SPR applications constitutes a powerful approach to develop sensitive immunoassays, as previously shown for non-SPR formats. As SPR-based assays have significantly expanded their reach in the last decade, such an approach promises to further accelerate their development.     

A Highlights from MBoC Selection: Quantitative Analysis of the Mechanism of Endocytic Actin Patch Assembly and Disassembly in Fission Yeast

We used quantitative confocal microscopy to measure the numbers of 16 proteins tagged with fluorescent proteins during assembly and disassembly of endocytic actin patches in fission yeast. The peak numbers of each molecule that accumulate in patches varied <30–50% between individual patches. The pathway begins with accumulation of 30–40 clathrin molecules, sufficient to build a hemisphere at the tip of a plasma membrane invagination. Thereafter precisely timed waves of proteins reach characteristic peak numbers: endocytic adaptor proteins (∼120 End4p and ∼230 Pan1p), activators of Arp2/3 complex (∼200 Wsp1p and ∼340 Myo1p) and ∼300 Arp2/3 complexes just ahead of a burst of actin assembly into short, capped and highly cross-linked filaments (∼7000 actins, ∼200 capping proteins, and ∼900 fimbrins). Coronin arrives last as all other components disperse upon patch internalization and movement over ∼10 s. Patch internalization occurs without recruitment of dynamins. Mathematical modeling, described in the accompanying paper (Berro et al., 2010, MBoC 21: 2803–2813), shows that the dendritic nucleation hypothesis can account for the time course of actin assembly into a branched network of several hundred filaments 100–200 nm long and that patch disassembly requires actin filament fragmentation in addition to depolymerization from the ends.