Hydrogenase 3 but not hydrogenase 4 is major in hydrogen gas production by Escherichia coli formate hydrogenlyase at acidic pH and in the presence of external formate

Fermenting Escherichia coli is able to produce formate and molecular hydrogen (H₂) when grown on glucose. H₂ formation is possessed by two hydrogenases, 3 (Hyd-3) and 4 (Hyd-4), those, in conjunction with formate dehydrogenase H (Fdh-H), constitute distinct membrane-associated formate hydrogenylases. At slightly alkaline pH (pH 7.5), the production of H₂ was found to be dependent on Hyd-4 and the F₀F₁-adenosine triphosphate (ATPase), whereas external formate increased the activity of Hyd-3. In this study with cells grown without and with external formate H₂ production dependent on pH was investigated. In both types of cells, H₂ production was increased after lowering of pH. At acidic pH (pH 5.5), this production became insensitive either to N,N′-dicyclohexylcarbodiimide or to osmotic shock and it became largely dependent on Fdh-H and Hyd-3 but not Hyd-4 and the F₀F₁-ATPase. The results indicate that Hyd-3 has a major role in H₂ production at acidic pH independently on the F₀F₁-ATPase.     

PHENOPSIS, an automated platform for reproducible phenotyping of plant responses to soil water deficit in Arabidopsis thaliana permitted the identification of an accession with low sensitivity to soil water deficit

The high-throughput phenotypic analysis of Arabidopsis thaliana collections requires methodological progress and automation. Methods to impose stable and reproducible soil water deficits are presented and were used to analyse plant responses to water stress. Several potential complications and methodological difficulties were identified, including the spatial and temporal variability of micrometeorological conditions within a growth chamber, the difference in soil water depletion rates between accessions and the differences in developmental stage of accessions the same time after sowing. Solutions were found. Nine accessions were grown in four experiments in a rigorously controlled growth-chamber equipped with an automated system to control soil water content and take pictures of individual plants. One accession, An1, was unaffected by water deficit in terms of leaf number, leaf area, root growth and transpiration rate per unit leaf area. Methods developed here will help identify quantitative trait loci and genes involved in plant tolerance to water deficit.     

Capacités reproductrices et descendance de poulets ayant subi un transfert de cellules germinales primordiales durant la vie embryonnaire

Résumé Des cellules germinales primordiales de Dindon, transférées par injection intravasculaire à des embryons de Poulet préalablement stérilisés, peuvent subir une maturation complète dans les gonades de l'hôte et fournir des gamètes plus ou moins aptes à la fécondation.Les spermatozoïdes résultants ont fécondé des oeufs de poule avec une fréquence plus grande que les spermatozoïdes normaux de Dindon, mais sans permettre un développement embryonnaire plus important ou plus normal. Cependant, on n'est pas parvenu à leur faire féconder des oeufs de dinde.Les ufs résultants ont montré parfois un vitellus anormal et n'ont été pondus que durant 7 mois. Mis en présence de spermatozoïdes de coq, ils ont été quelquefois fécondés (ou peutêtre simplement activés), mais ils n'ont jamais fourni d'embryons. Fécondés par des spermatozoïdes de Dindon, ils ont fourni des embryons, parfois anormaux, qui ont atteint dans le meilleur cas le 15è jour d'incubation (stade 38 HH). Certainespraepennae de l'embryon qui a atteint ce stade montraient une pigmentation rouge brun qui ne peut pas être déterminée par le génotype du zygote (celui d'un Dindon de race blanche) et qui rappelle le phénotype de la mère nourricière (une poule rouge brun).A la suite de transferts intraspécifiques de cellules germinales primordiales, on a pu obtenir la maturation d'ovocytes de Rhode Island Red à l'intérieur d'un ovaire de Wyandotte Blanche (chez deux poules) et vice versa (chez une poule). La ponte a également été possible, mais à une fréquence souvent inférieure à la normale.Lorsqu'une poule Wyandotte Blanche, portant des ovocytes de Rhode Island Red a été accouplée à un coq Rhode Island Red normal, le duvet de leurs descendants s'est montré plus clair que celui des poussins Rhode Island Red dans un cas, mais il a été remplacé ensuite par un plumage rouge brun conforme au génotype.Lorsqu'une poule Rhode Island Red, portant des ovocytes de Wyandotte Blanche a été accouplée à un coq Wyandotte Blanche normal, le duvet de leurs descendants n'a jamais été conforme au génotype. Il présentait toujours du pigment noir plus ou moins étendu et, dans un cas, du pigment rouge brun, qui sont tous deux présents chez la mère nourricière. On ne connaît ni l'origine ni le mécanisme d'un tel transfert de pigments, qui ne représente peut-être qu'un effet transitoire affectant le duvet de la première génération. La crête, quant à elle, s'est toujours montrée conforme au génotype d'origine.

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Reproductive capacity and offspring of chickens submitted to a transfer of primordial germ cells during embryonic life

Summary Turkey primordial germ cells transfered by intravascular injection to previously sterilized chick embryos can undergo complete maturation inside the host's gonads and can give rise to gametes which are more or less suitable for fertilization.The resulting spermatozoa fertilized hen eggs at a higher frequency than normal turkey spermatozoa, but without allowing a longer or a more normal development. However, it was impossible to fertilize turkey eggs with them.The resulting eggs sometimes had an abnormal-looking yolk and were laid during the first 7 months only. Brought in contact with chicken spermatozoa, they were fertilized (or perhaps merely activated), but they never gave rise to embryos. Fertilized by turkey spermatozoa, they developed into embryos, sometimes abnormal, which in the best case reached the 15th day of incubation (stage 38 HH). Somepraepennae of the latter embryo showed a red-brown pigment which cannot be determined by the genotype of the zygote (a white turkey's) and which resembled the phenotype of the foster mother (a red-brown hen).After intraspecific transfer of primordial germ cells, maturation of Rhode Island Red oöcytes inside a Wyandotte White ovary (in two hens) and vice versa (in one hen) was achieved. Laying was also possible but often at a lower frequency than normal.When a Wyandotte White hen bearing Rhode Island Red oöcytes was mated with a normal Rhode Island cock, the down of their offspring looked brighter than Rhode Island Red chicken's in one case, but it was subsequently replaced by red-brown feathers according to the genotype.When a Rhode Island Red hen bearing Wyandotte White oöcytes was mated with a normal Wyandotte White cock, the down of their offspring was never in agreement with the genotype. It always showed a black pigment over more or less large areas and, in one case, a red-brown pigment, both of which were present in the foster mother. The origin and the mechanism of such a transfer of pigments are not understood. It might represent merely a temporary effect acting upon the down of the first generation. As far as the comb is concerned, it was always in agreement with the original genotype.

     

Stages of Infection during the Tripartite Interaction between Xenorhabdus nematophila, Its Nematode Vector, and Insect Hosts

Bacteria of the genus Xenorhabdus are mutually associated with entomopathogenic nematodes of the genus Steinernema and are pathogenic to a broad spectrum of insects. The nematodes act as vectors, transmitting the bacteria to insect larvae, which die within a few days of infection. We characterized the early stages of bacterial infection in the insects by constructing a constitutive green fluorescent protein (GFP)-labeled Xenorhabdus nematophila strain. We injected the GFP-labeled bacteria into insects and monitored infection. We found that the bacteria had an extracellular life cycle in the hemolymph and rapidly colonized the anterior midgut region in Spodoptera littoralis larvae. Electron microscopy showed that the bacteria occupied the extracellular matrix of connective tissues within the muscle layers of the Spodoptera midgut. We confirmed the existence of such a specific infection site in the natural route of infection by infesting Spodoptera littoralis larvae with nematodes harboring GFP-labeled Xenorhabdus. When the infective juvenile (IJ) nematodes reached the insect gut, the bacterial cells were rapidly released from the intestinal vesicle into the nematode intestine. Xenorhabdus began to escape from the anus of the nematodes when IJs were wedged in the insect intestinal wall toward the insect hemolymph. Following their release into the insect hemocoel, GFP-labeled bacteria were found only in the anterior midgut region and hemolymph of Spodoptera larvae. Comparative infection assays conducted with another insect, Locusta migratoria, also showed early bacterial colonization of connective tissues. This work shows that the extracellular matrix acts as a particular colonization site for X. nematophila within insects.     

Power to detect risk alleles using genome-wide tag SNP panels

Advances in high-throughput genotyping and the International HapMap Project have enabled association studies at the whole-genome level. We have constructed whole-genome genotyping panels of over 550,000 (HumanHap550) and 650,000 (HumanHap650Y) SNP loci by choosing tag SNPs from all populations genotyped by the International HapMap Project. These panels also contain additional SNP content in regions that have historically been overrepresented in diseases, such as nonsynonymous sites, the MHC region, copy number variant regions and mitochondrial DNA. We estimate that the tag SNP loci in these panels cover the majority of all common variation in the genome as measured by coverage of both all common HapMap SNPs and an independent set of SNPs derived from complete resequencing of genes obtained from SeattleSNPs. We also estimate that, given a sample size of 1,000 cases and 1,000 controls, these panels have the power to detect single disease loci of moderate risk (λ ~ 1.8–2.0). Relative risks as low as λ ~ 1.1–1.3 can be detected using 10,000 cases and 10,000 controls depending on the sample population and disease model. If multiple loci are involved, the power increases significantly to detect at least one locus such that relative risks 20%–35% lower can be detected with 80% power if between two and four independent loci are involved. Although our SNP selection was based on HapMap data, which is a subset of all common SNPs, these panels effectively capture the majority of all common variation and provide high power to detect risk alleles that are not represented in the HapMap data.     

Does Optic Nerve Head Surface Topography Change Prior to Loss of Retinal Nerve Fiber Layer Thickness: A Test of the Site of Injury Hypothesis in Experimental Glaucoma

Purpose

To test the hypothesis that optic nerve head (ONH) deformation manifesting as changes in its mean surface height precedes thinning of the peripapillary retinal nerve fiber layer (RNFL) in experimental glaucoma (EG).

Methods

68 rhesus macaque monkeys each had three or more baseline imaging sessions under manometric intraocular pressure (IOP) control to obtain average RNFL thickness (RNFLT) and the ONH surface topography parameter mean position of the disc (MPD). Laser photocoagulation was then applied to the trabecular meshwork of one eye to induce chronic, mild-to-moderate IOP elevation and bi-weekly imaging continued. Event analysis was applied to determine for each parameter when an ‘endpoint’ occurred (signficant change from baseline) for eight different endpoint criteria. Specificity was assessed in the group of 68 fellow control eyes. Classical signal detection theory and survival analysis were used to compare MPD with RNFLT.

Results

Regardless of the endpoint criterion, endpoints were always more frequent for MPD than for RNFLT. The discriminability index (d’) was 2.7 ± 0.2 for MPD and 1.9 ± 0.2 for RNFLT (p<0.0001). Endpoints were reached by MPD an average of 1-2 months earlier than by RNFLT (p<0.01). At the onset of the first specific, detectable MPD change in EG eyes, there was still no significant change in RNFLT on average (p=0.29) and only 25% of individual eyes exhibited signficant reduction. In contrast, at onset of signficant RNFLT change, MPD had already changed an average of 101 µm from baseline (p<0.0001) and 71% of the individual eyes had exhibited significant change. The magnitude of MPD change was more than could be explained on the basis of axon loss alone.

Conclusions

This study demonstrates that the average surface height of the ONH changes prior to any detectable loss of average peripapillary RNFL thickness in non-human primate eyes with experimental glaucoma.