Mixed-model of ANOVA for measurement reproducibility in proteomics

This work is a statistical analysis of reproducibility of a MALDI-TOF mass spectrometry experiment. Its aim is to evaluate measurement variability and compare peak intensities from two types of MALDI-TOF platforms. We compared and commented on the abilities of Principal Component Analysis and mixed-model analysis of variance to evaluate the biological variability and the technical variability of peak intensities in different patients. The properties and hypotheses of both methods are summarized and applied to spectra from plasma of patients with Hodgkin lymphoma. Principal Component Analysis checks rapidly the balance between the two variabilities; however, a mixed-model analysis of variance is necessary to quantify the biological and technical components of the experimental variance as well as their interactions and to split the total variance into between-subjects and within-subject components. The latter method helped to assess the reproducibility of measurements from two MALDI-TOF platforms and to decompose the technical variability according to the experimental design.     

Repeated therapeutic dosing selects macrolide‐resistant Campylobacter spp. in a turkey facility

Aims: This study assessed the effects of the therapeutic use of Tylan^® in a large‐scale turkey production facility on the selection of macrolide‐resistant Campylobacter. Methods and Results: A flock of production turkeys (c. 30 000 birds) was followed from brooding to slaughter, and the effects of macrolide application was assessed in one half of the flock from finishing stage to final product and compared against the control barn where no macrolide was used. Overall, Campylobacter prevalence in turkeys was almost 100% by 4 weeks of age. When Campylobacter prevalence was assessed in relation to treatment, high levels of macrolide resistance were evident in this group following treatment, with Campylobacter coli becoming the dominant strain type. Over time, and in the absence of a selection agent, the population of resistant strains decreased suggesting that there was a fitness cost associated with macrolide resistance carriage and persistence. Macrolide resistance was detected in the control barn at a very low level (four isolates recovered during the study), suggesting that the creation or selection of macrolide‐resistant Campylobacter was correlated with the treatment regime used. Molecular analysis of a selection of macrolide‐resistant Campylobacter recovered was assessed using PCR, RFLP and sequence analysis of the 23S rRNA. The majority of isolates displaying high‐level macrolide resistance (>256 μg ml^?1) possessed an A2075G transition mutation in the 23S rRNA and the CmeABC efflux pump. Conclusions: These studies suggest that macrolide resistance can be promoted through the application of treatment during the grow‐out phase and once established in a production facility has the potential to persist and be transferred to final product. Significance and Impact of the Study: The study highlights the prudent use of antimicrobials in treatment of disease in poultry. Of significance is the presence of macrolide‐resistant Campylobacter in poultry production and finished product as a consequence of macrolide usage.     

Design and synthesis of APTCs (aminopyrrolidinetricarboxylic acids): identification of a new group III metabotropic glutamate receptor selective agonist

A new family of mGlu receptor orthosteric ligands called APTCs was designed and synthesized using a parallel chemistry approach. Amongst 65 molecules tested on mGlu4, mGlu6 and mGlu8 subtypes, (2S,4S)-4-amino-1-[(E)-3-carboxyacryloyl]pyrrolidine-2,4-dicarboxylic acid (8a06-FP0429) has been shown to be a full mGlu4 agonist and a partial mGlu8 agonist. In addition, 8a06 was shown to be selective versus group I and II mGlu subtypes. A possible explanation for this efficacy difference is proposed by docking experiment performed with molecular model of the receptor.     

Recording Behavioral Responses to Reflection in Crayfish

Social behavior depends on sensory input from the visual, mechanical and olfactory systems. One important issue concerns the relative roles of each sensory modality in guiding behavior. The role of visual inputs has been examined by isolating visual stimuli from mechanical and chemosensory stimuli. In some studies (Bruski & Dunham, 1987: Delgado-Morales et al., 2004) visual inputs have been removed with blindfolds or low light intensity, and effects of remaining sensory modalities have been elucidated. An alternative approach is to study the effects of visual inputs in the absence of any appropriate mechanical and chemosensory cues. This approach aims to identify the exclusive role of visual inputs. We have used two methods to provide visual stimuli to crayfish without providing chemical and mechanical cues. In one method, crayfish are videotaped in an aquarium where half of the walls are covered in mirrors to provide a reflective environment, and the other half are covered in a non-reflective (matte finish) plastic. This gives the crayfish a choice between reflective and non-reflective environments. The reflective environment provides visual cues in the form of reflected images of the crayfish as it moves throughout half of the tank; these visual cues are missing from the non-reflective half of the tank. An alternative method is to videotape the behavior of crayfish in an aquarium separated by a smaller chamber at each end, with a crayfish in one small chamber providing visual cues and an inert object in the opposite small chamber providing visual input from a non-moving, non-crayfish source. Our published results indicate that responses of crayfish to the reflective environment depend on socialization and dominance rank. Socialized crayfish spent more time in the reflective environment and exhibited certain behaviors more frequently there than in the non-reflective environment; isolated crayfish showed no such differences. Crayfish that were housed in same-sex pairs developed a social rank of either dominant or subordinate. Responses to reflection differed between dominant and subordinate crayfish (May & Mercier, 2006; May & Mercier, 2007). Dominant crayfish spent more time on the reflective side, entered reflective corners more frequently and spent more time in reflective corners compared to the non-reflective side. Subordinate crayfish walked in reverse more often on the reflective side than on the non-reflective side. Preliminary data suggest similar effects from visual cues provided by a crayfish in a small adjoining chamber (May et al., 2008).Download video file.^{(129M, mp4)}     

Additional haplogroups of Toxoplasma gondii out of Africa: population structure and mouse-virulence of strains from Gabon

Background

Toxoplasma gondii is found worldwide, but distribution of its genotypes as well as clinical expression of human toxoplasmosis varies across the continents. Several studies in Europe, North America and South America argued for a role of genotypes in the clinical expression of human toxoplasmosis. Genetic data concerning T. gondii isolates from Africa are scarce and not sufficient to investigate the population structure, a fundamental analysis for a better understanding of distribution, circulation, and transmission.

Methodology/Principal Findings

Seropositive animals originating from urban and rural areas in Gabon were analyzed for T. gondii isolation and genotyping. Sixty-eight isolates, including one mixed infection (69 strains), were obtained by bioassay in mice. Genotyping was performed using length polymorphism of 13 microsatellite markers located on 10 different chromosomes. Results were analyzed in terms of population structure by Bayesian statistical modeling, Neighbor-joining trees reconstruction based on genetic distances, F _ST and linkage disequilibrium. A moderate genetic diversity was detected. Three haplogroups and one single genotype clustered 27 genotypes. The majority of strains belonged to one haplogroup corresponding to the worldwide Type III. The remaining strains were distributed into two haplogroups (Africa 1 and 3) and one single genotype. Mouse virulence at isolation was significantly different between haplogroups. Africa 1 haplogroup was the most virulent.

Conclusion

Africa 1 and 3 haplogroups were proposed as being new major haplogroups of T. gondii circulating in Africa. A possible link with strains circulating in South and Central America is discussed. Analysis of population structure demonstrated a local spread within a rural area and strain circulation between the main cities of the country. This circulation, favored by human activity could lead to genetic exchanges. For the first time, key epidemiological questions were addressed for the West African T. gondii population, using the high discriminatory power of microsatellite markers, thus creating a basis for further epidemiological and clinical investigations.     

Neuroprotective Effect of Inhaled Nitric Oxide on Excitotoxic-Induced Brain Damage in Neonatal Rat

Background

Inhaled nitric oxide (iNO) is one of the most promising therapies used in neonates. However, little information is known about its impact on the developing brain submitted to excitotoxic challenge.

Methodology/Principal Findings

We investigated here the effect of iNO in a neonatal model of excitotoxic brain lesions. Rat pups and their dams were placed in a chamber containing 20 ppm NO during the first week of life. At postnatal day (P)5, rat pups were submitted to intracranial injection of glutamate agonists. At P10, rat pups exposed to iNO exhibited a significant decrease of lesion size in both the white matter and cortical plate compared to controls. Microglia activation and astrogliosis were found significantly decreased in NO-exposed animals. This neuroprotective effect was associated with a significant decrease of several glutamate receptor subunits expression at P5. iNO was associated with an early (P1) downregulation of pCREB/pAkt expression and induced an increase in pAkt protein concentration in response to excitotoxic challenge (P7).

Conclusion

This study is the first describe and investigate the neuroprotective effect of iNO in neonatal excitotoxic-induced brain damage. This effect may be mediated through CREB pathway and subsequent modulation of glutamate receptor subunits expression.     

AKT/FOXO signaling enforces reversible differentiation blockade in myeloid leukemias

AKT activation is associated with many malignancies, where AKT acts, in part, by inhibiting FOXO tumor suppressors. We show a converse role for AKT/FOXOs in acute myeloid leukemia (AML). Rather than decreased FOXO activity, we observed that FOXOs are active in ～40% of AML patient samples regardless of genetic subtype. We also observe this activity in human MLL-AF9 leukemia allele-induced AML in mice, where either activation of Akt or compound deletion of FoxO1/3/4 reduced leukemic cell growth, with the latter markedly diminishing leukemia-initiating cell (LIC) function in vivo and improving animal survival. FOXO inhibition resulted in myeloid maturation and subsequent AML cell death. FOXO activation inversely correlated with JNK/c-JUN signaling, and leukemic cells resistant to FOXO inhibition responded to JNK inhibition. These data reveal a molecular role for AKT/FOXO and JNK/c-JUN in maintaining a differentiation blockade that can be targeted to inhibit leukemias with a range of genetic lesions.     

Evidence of dietary feedback in a facultative association between deep-sea gastropods and sea anemones

While epibiotic associations between macrobenthic invertebrates are common, the role they play in the feeding ecology of intervening species is often incompletely understood. The diets of epibiotic sea anemones Allantactis parasitica and their gastropod hosts were analyzed using digestive tract contents, lipid biomarkers and observations of live specimens in an attempt to detect dietary feedback from the facultative association. Comparisons were made using symbiotic individuals and asymbiotic counterparts collected at depths of 191-627 m from three neighbouring areas in the northwest Atlantic. Gastropods carrying one or two epibionts had higher stomach indices than those harbouring three epibionts or no epibiont. The diet of symbiotic gastropods was also more diversified based on stomach contents and lipid analysis. Among other things, symbiotic gastropods contained four times more lipids and a greater proportion of Σn−3 fatty acids. Gastrovascular cavity content indices of asymbiotic sea anemones were generally lower than those of symbiotic counterparts. Their cavities were more often empty, and their diet less diversified with fewer benthic items, suggesting that foraging of gastropods through the sediments makes more food available to sea anemones living as epibionts. Lipid analysis showed some disparities between symbiotic and asymbiotic sea anemones at the regional scale, including in percent phospholipids and in the proportion of certain fatty acids. Together these findings indicate that mutual protection against predators leads to prolonged and more efficient foraging for gastropods and increased time spent deployed (feeding) in food-rich areas for sea anemones, thus enabling both partners to fully exploit food resources that may be limited at bathyal depths.     

Contrasting reproductive strategies in three deep-sea octocorals from eastern Canada: Primnoa resedaeformis, Keratoisis ornata, and Anthomastus grandiflorus

Various aspects of reproduction were studied in three deep-sea octocorals belonging to the order Alcyonacea that co-occur at bathyal depths on the continental edge and the slope of eastern Canada. The main goals were to expand knowledge of deep-water heterotrophic corals and ascertain whether reproductive strategies could explain the known patterns of occurrence. Anthomastus grandiflorus is a gonochoric species with a female-biased sex ratio that exhibits internal fertilization and brooding of planula larvae. Conversely, Primnoa resedaeformis and Keratoisis ornata rely on broadcast spawning and external fertilization; their sexuality remains undetermined as spermatocysts were not found. In P. resedaeformis, the presence of mixed size classes of oocytes in samples from all months, depths, and locations studied suggests continuous oogenesis or overlapping development of oocyte cohorts, indicative of a gametogenic cycle spanning more than a year. No evidence of periodicity was found in this species, although it could have been masked by the striking bathymetric variation in potential relative fecundity (oocytes polyp⁻¹). The two other octocorals displayed a clear annual breeding pattern. Spawning in K. ornata and larval release in A. grandiflorus occurred in late summer and fall, respectively, possibly in response to environmental factors, as supported by shifts in the reproductive peak of A. grandiflorus across latitudes. The three species are presumed to share a nonfeeding larval mode, and data on their reproductive potential do not present any striking disparities. Published data on bycatches and video surveys in Atlantic Canada indicate that the gonochoric brooder A. grandiflorus is more widely distributed than the two free spawners, P. resedaeformis and K. ornata, which is contrary to common dispersal potential paradigms.     

FANCM-associated proteins MHF1 and MHF2, but not the other Fanconi anemia factors,limit meiotic crossovers

Genetic recombination is important for generating diversity and to ensure faithful segregation of chromosomes at meiosis. However, few crossovers (COs) are formed per meiosis despite an excess of DNA double-strand break precursors. This reflects the existence of active mechanisms that limit CO formation. We previously showed that AtFANCM is a meiotic anti-CO factor. The same genetic screen now identified AtMHF2 as another player of the same anti-CO pathway. FANCM and MHF2 are both Fanconi Anemia (FA) associated proteins, prompting us to test the other FA genes conserved in Arabidopsis for a role in CO control at meiosis. This revealed that among the FA proteins tested, only FANCM and its two DNA-binding co-factors MHF1 and MHF2 limit CO formation at meiosis.