Cryptosporidium sp. of the starred lizardAgama stellio: Ultrastructure and life cycle

The life cycle ofCryptosporidium sp. in the gut of its reptilian hostAgama stellio was studied by electron microscopy. The parasite was located in a parasitophorous vacuole formed at the microvillous surface of the gut epithelium and was separated from the host-cell cytoplasm by a microfibrillar mesh and a dark band. One type of merogony was observed that produced eight merozoites. The microgamete lacked a flagellum and possessed a unique, anterior adhesive zone. The macrogamete had two types of wall-forming bodies corresponding to those of other coccidia. Sporulated oocysts possessed either a thick or a thin wall. Oocysts were similar in size to those ofCryptosporidium in other reptiles, including another agamed species, and the different life-cycle stages conformed ultrastructurally with those of isolates found in mammals and birds. This is the first detailed electron microscopic study ofCryptosporidium in a reptile.Abbreviations A amylopectin granules - AO attachment organelle - AZ adhesive zone - B basal body - C crystalline body - DB dense band - er endoplasmic reticulum - HC host cell - FM filamentous mesh - L lamellae - Lp lipid vacuole - M merozoites - Mg microgamete - Mn micronemes - Mt microtubules - N nucleus - Nu nucleolus - OL oocyst layer - Pm membrane of the parasitophorous envelope - PV parasitophorous vacuole - r rhoptries - R ring-shaped fusion - RB residual body - Sp sporozoite - Su suture - U oocyst outer membrane - WF ₁ wall-forming body type 1 - WF ₂ wall-forming body type 2     

Mutations in the structural gene for cytochrome c result in deficiency of both cytochromes aa 3 and c in Neurospora crassa

The cyt-12-12 mutant of Neurospora crassa is characterized by slow growth and a deficiency of spectrophotometrically-detectable cytochromes aa ₃ and c. Using a sib-selection procedure we have isolated the cyt-12 ⁺ allele from a cosmid library of N. crassa genomic DNA. Characterization of the cyt-12 ⁺ allele reveals that it encodes the structural gene for cytochrome c. DNA sequence analysis of the cyt-12-12 allele revealed a mutation in the cytochrome c coding sequence that results in replacement of a glycine residue, which is invariant in the cytochrome c of other species, with an aspartic acid. Genetic analysis confirms that cyt-12-12 is allelic with the previously-characterized cyc-1-1 mutant, which was also shown to affect the single locus encoding cytochrome c in N. crassa. We suggest that the amount of functional cytochrome c present in mitochondria influences the level of cytochrome aa ₃ .     

Further studies on resistance toFasciola hepatica andEchinostoma revolutum in mice infected withSchistosoma sp.

Schistosoma sp. induced cross-resistance to a challenge withFasciola hepatica andEchinostoma revolutum was studied in mice. Primary patent 56-days-oldS. intercalatum andS. bovis infections stimulated a statistically significant level of resistance to a challenge withF. hepatica, and primary patent 100-day-oldS. bovis infections induced an almost complete resistance to a challenge withE. revolutum. Primary single-sexS. mansoni infections, either male or female, aged 90 days did not stimulate any resistance to a challenge withE. revolutum.A primary infection withS. mansoni aged 70 days induced a marked reduction (94.1–100%) in theE. revolutum worm recovery already 2 h post-challenge as compared with that of theE. revolutum challenge control group and complete elimination of the echinostome worm population inS. mansoni infected mice had taken place 24 h after challenge. E. revolutum worm populations established in mice harbouring newly patent 36-day-oldS. mansoni infections persisted unchanged for a period of at least 33 days into the patent period of the schistosome infection in spite of development of a complete resistance to a challenge withE. revolutum metacercariae during this period.     

Akaike's information criterion for a measure of linkage disequilibrium

 The extent and distribution of linkage disequilibrium (LD) in humans is a current topic especially for gene mapping of complex diseases. Akaike's information criterion (AIC) was applied to estimate LD and compared with other standard LD measures, D′ and r ². By comparison of an independent model (IM; linkage equilibrium) and a dependent model (DM; linkage disequilibrium), the parsimonious model is the one with the smaller AIC score. Therefore, the extent of LD by AIC is expressed as AIC(IM) — AIC(DM)(AIC(LD)). A total of 39 single-nucleotide polymorphisms on a 1.6-Mb region of chromosome 21q22 were identified, and genotyped in 192 Japanese individuals. All possible pairs were analyzed to estimate LD and the analyses were compared. AIC(LD) became highly positive as the D′ value increased and was negative at D′ values of around 0.2. Because a negative value of AIC(LD) implies linkage equilibrium, D′ values below 0.2 should be regarded as linkage equilibrium. The LD estimate by AIC yielded results similar to those obtained by r ², indicating that AIC(LD) would be useful for fine gene mapping. Received: July 3, 2002 / Accepted: October 2, 2002     

The Drosophila projectin mutant,bent D,has reduced stretch activation and altered indirect flight muscle kinetics

Projectin is a ca. 900 kDa protein that is a member of the titin protein superfamily. In skeletal muscle titins are involved in the longitudinal reinforcement of the sarcomere by connecting the Z-band to the M-line. In insect indirect flight muscle (IFM), projectin is believed to form the connecting filaments that link the Z-band to the thick filaments and is responsible for the high relaxed stiffness found in this muscle type. The Drosophila mutant bent ^D (bt ^D ) has been shown to have a breakpoint close to the carboxy-terminal kinase domain of the projectin sequence. Homozygotes for bt ^D are embryonic lethal but heterozygotes (bt ^D /+) are viable. Here we show that bt ^D /+ flies have normal flight ability and a slightly elevated wing beat frequency (bt ^D /+ 223 ± 13 Hz; +/+203 ± 5 Hz, mean ± SD; P < 0.01). Electron microscopy of bt ^D /+ IFM show normal ultrastructure but skinned fiber mechanics show reduced stretch activation and oscillatory work. Although bt ^D /+ IFM power output was at wild-type levels, maximum power was achieved at a higher frequency of applied length perturbation (bt ^D /+ 151 ± 6 Hz; +/+ 102 ± 14 Hz; P < 0.01). Results were interpreted in the context of a viscoelastic model of the sarcomere and indicate altered cross-bridge kinetics of the power-producing step. These results show that the bt ^D mutation reduces oscillatory work in a way consistent with the proposed role of the connecting filaments in the stretch activation response of IFM.     

Pregnancy and lactation have anti-obesity and anti-diabetic effects in Ay/a mice

Aim: Dominant ‘yellow’ mutation at the mouse agouti locus (A^y) results in obesity. Pregnancy and lactation are characterized by large energy demand. The aim of this study was to investigate whether obesity would develop in pregnant and suckling A^y mice. Methods: Body weight and food intake in pregnancy, lactation, and after weaning, plasma leptin, insulin, corticosterone and blood glucose concentrations on days 7, 13 and 18 of pregnancy, days 1, 10, 21 and 80 postpartum, glucose and insulin tolerance on pregnancy days 7 and 18 were measured in C57Bl/6J mice of a/a (normal metabolism) and A^y/a genotypes. The same parameters were also measured in age-matched virgin females. Results: Virgin A^y/a females exhibited hyperphagia, enhanced body weight, glucose intolerance and normal blood parameters at the mating age. With age, they developed obesity, hyperleptinaemia, hyperinsulinaemia and hyperglycaemia. Obesity did not develop in mated A^y/a mice; during suckling, they had equal food intake and body weight as a/a mice. During pregnancy, glucose tolerance was enhanced in A^y/a mice and became equal in both genotypes. In both genotypes, concentrations of hormones increased, and glucose decreased from pregnancy day 7 to day 18 and returned to normal values after parturition. A^y/a mice did not differ from a/a in corticosterone, insulin and glucose levels during pregnancy and lactation, in leptin levels during suckling; however, A^y/a mice had two times higher leptin levels than a/a during pregnancy. After weaning, A^y/a mice began to eat and weigh more than a/a exhibiting normal metabolic parameters for 50 days. Conclusion: Pregnancy and lactation retard obesity and diabetes development in A^y mice.     

Polymorphism around cog extends into adjacent structural genes

The recombination hotspot cog overlaps a highly polymorphic 950-bp region of linkage group I in Neurospora crassa. The sequence of this region in the four strains, Lindegren 25a, Lindegren A, Emerson A and St. Lawrence 74A, each differs from the others by 1.4% or more. Comparison of the sequence of St. Lawrence 74A and Lindegren 25a each side of cog shows a high level of sequence heterology extending in both directions, including the coding sequences for his-3 and a putative gene lpl with homology to yeast lysophospholipase. The St. Lawrence 74A and Lindegren 25a sequences of his-3, centromere-proximal to cog, differ at 14 nucleotides, resulting in six amino-acid variations between the predicted protein sequences. In lpl, distal from cog, the sequences differ at 19 nucleotides leading to five amino-acid differences between the predicted proteins. Sequence heterology between St. Lawrence 74A and Lindegren 25a peaks either side of cog and then declines with distance. At the am locus on linkage group V, heterology is much less but peaks close to a weak recombination hotspot 5′ of the coding sequence. Uneven distribution of polymorphism along chromosomes has been explained by a hitch-hiking hypothesis in which selection for advantageous mutations causes local fixation of unselected variation. We suggest that new mutations arising from errors in recombination also contribute to the uneven distribution of polymorphism. Received: 15 October 1998 / 4 February 1999     

Identification and cloning of a mobile transposon fromAspergillus niger var.awamori

Aspergillus niger var.awamori contains multiple copies of a transposable element, Vader. This element was detected as a 437-bp insertion in four independently isolated spontaneous mutants of theniaD (nitrate reductase) gene. The Vader element is present in approximately 15 copies in bothA. niger var.awamori andA. niger. A single copy of Vader was detected from only one of the two laboratory strains ofA. nidulans which were also examined. Insertion of the Vader element into theniaD gene ofA. niger var.awamori caused a 2-bp duplication (TA) of the target sequence. The Vader element is flanked by a 44-bp inverted repeat. The genetic stabilities of the inserted Vader elements atniaD were examined by studying reversion frequencies resulting in colonies able to grow on nitrate as a sole nitrogen source. MutantsniaD392 andniaD436 reverted at a frequency of 9x10^-3 and 4x10^-2, respectively. Two of the mutants,niaD587 andniaD410, reverted at a lower frequency of 6x10^-4.     

Expression and stability of a Rhizobium leguminosarum bv. trifolii Sym plasmid in R. meliloti and Agrobacterium tumefaciens and its effect on clover-bacterium symbiosis

The Rhizobium leguminosarum bv. trifolii Sym plasmid pRtr5a was transferred by conjugation to R. meliloti strains deleted or mutated in the nod and fix genes, and also to a plasmid-less strain of Agrobacterium tumefaciens. The plasmid transfer led to the elimination of the resident cryptic plasmid of the R. meliloti recipients. The Sym plasmid of the recipients was only eliminated in the case of a deletion. In some cases the transferred plasmid was instable. The nod and nif/fix genes of R. leguminosarum bv. trifolii were expressed in R. meliloti and somewhat delayed in A. tumefaciens. A high level of nitrogen fixation on clover was only found in R. meliloti containing pRtr5a. However, this occurred only if the nif/fix genes of the recipient had been deleted before. Inoculation of clover with this R. meliloti strain resulted in a higher dry matter production in comparison to clover inoculated with the donor strain.     

Metathesis polymerization of exo,exo-, endo,exo- and endo,endo-5,6-dimethylbicyclo[2.2.1]hept-2-enes; microstructure of the polymers and their hydrogenated products

Polymers of exo,exo-, endo,exo, and endo,endo-5,6-dimethylbicyclo[2.2.1]hept-2-ene monomers were prepared using a selection of eight metathesis catalysts and their structures examined by ¹³C NMR spectroscopy. Tacticities could be determined for the polymers of the endo, endo and endo,exo monomers and for all three hydrogenated polymers. RuCl₃ gave hightrans atactic polymers while ReCl₅ gave high-cis syndiotactic polymers, but WCl₆/Bu₄Sn gave a high-cis atactic polymer of the endo,endo monomer. The endo,exo monomer was unusual in giving polymers of relatively low cis content (fraction of cis double bonds σ_c ≤ 0,3), and with strong exo,endo (XN) bias except in the case of RuCl₃. The parallel with the head, tail (HT) biased polymers of 1-methylbicyclo[2.2.1]hept-2-ene is noted. Polymers of the exo,exo and endo,endo monomers have a blocky cis/trans double bond distribution at moderate cis content. The results are discussed in terms of a mechanism involving propagation by metal-carbene, metal-carbene-olefin and metallacyclobutane complexes.     

Exercise breathing pattern during chronic altitude exposure

Summary Breathing pattern in response to maximal exercise was examined in four subjects during a 7-day acclimatisation to a simulated altitude of 4247 m (barometric pressure,PB = 59.5 kPa). Graded exercise tests to exhaustion were performed during normoxia (day 0), and on days 2 and 7 of hypoxia, respectively. Ventilation was significantly augmented in the hypoxic environment, as were both the mean inspiratory flow (V _T/T _I) and inspiratory duty cycle (T _I/T _TOT) components of it.V _I/T _I was increased due to a significant increase in tidal volume (VT) and a corresponding decrease in inspiratory time duration (TI). Throughout a range of exercise ventilation,T _I/T _TOT was increased due to an apparently greater decrease in expiratory time duration (T _E) with respect to TI. In all cases, the relation betweenV _T andT _I displayed a typical range 2 behaviour, with evidence of a range 3 occurring at very high ventilatory rates. There was essentially no difference observed in theV _T-T _I relation during exercise between the normoxic and hypoxic conditions. No significant changes were observed in the breathing pattern in response to exercise within the exposure period (from day 2 to day 7), although there was a discernible tendency to a higher stage 3 plateau by day 7 of altitude exposure.     

Effect of oral creatine ingestion on parameters of the work rate-time relationship and time to exhaustion in high-intensity cycling

The relationship between work rate (W˙) and time to exhaustion (t) during intense exercise is commonly described by either a hyperbolic function (NLin), t=W ^′/(W˙?W˙ _cp), or by its linear equivalent (LinW) W _lim =W ^′+W˙ _cp(t). The parameter W˙<INF _cp (critical power) has been described as an inherent characteristic of the aerobic energy system, while W?′ has been shown to be a ralid estimate of anaerobic work capacity. Recent studies have demonstrated that oral supplementation of creatine monohydrate (CrH₂O) increases total muscle creatine stores, and have linked these increases to improved performances in intense intermittent exercise. This study was conducted to determine the effect of CrH₂O supplementation on estimates of W?′ and W˙<INF _cp derived from the NLin and LinW equations, and to determine the effect of CrH₂O on t in exhaustive constant power exercise of different intensities. Fifteen active but untrained university students completed three phases of testing on a cycle ergometer: (1) familiarization, three learning trials, (2) baseline determination of W?′ and W˙<INF _cp, four bouts performed at a W˙ selected to elicit fatigue in 90–600?s, and (3) experimental determination of W?′ and W˙ _cp, four bouts performed at the same W˙ as baseline, but performed after 5 days of ingesting either a placebo (4?×?6?g of glucose/day) or CrH₂O (4?×?5?g of CrH₂O and 1?g glucose/day). Testing was administered in a double-blind manner. Analyses of covariance revealed a significant effect for CrH₂O on both estimates of W?′ (NLin, P=0.04; LinW, P<0.01), but not on estimates of W˙ _cp (NLin, P=0.37; LinW; P=0.30). Within groups, t was significantly different for only CrH₂O at the two highest W˙s (P=0.04). It is concluded that oral ingestion of CrH₂O increases estimates of W?′ due to an improved t at the shorter, more intense exercise bouts.

     

Die Entwicklung des Paraganglion caroticum und der Paraganglien am Herzen des Schweines

Ohne ZusammenfassungErklärung der Abkürzungen auf den Abbildungen a Aorta - a.a. Aorta ascendens - a.b.c. A. brachiocephalica - a.d. Aorta descendens - a.o. A. occipitalis - a.s. Ansa N. sympathici - aur.d. Rechtes Herzohr - aur.s. Linkes Herzohr - c.c. A. carotis communis - c.cd. R. cardiacus caudalis X - e.cr. R. cardiacus cranialis X - c.e. A. carotis externa - c.i. A. carotis interna - C.K. Carotisknötchen - d.a. Ductus arteriosus - E.Z. Gefäßendothelzellen - g.e. Ganglion extracraniale IX - g.n. Ganglion nodosum X - g.s..cd. Ganglion cervicale caudale N. sympathici - g.s.cr. Ganglion cervicale craniale N. sympathici - G. Blutgefäß - M.Z. Mesenchymzellen - n.i. Nervus intercaroticus (IX) - N.Z. Nervenzellen - N.Z.l. Nervenzellen der linken Seite - N.Z.r. Nervenzellen der rechten Seite - oe. Oesophagus - p.c. Paraganglion caroticum - p.d. A. pulmonalis dextra - p.di. A. pulmonalis distalis - p.p. A. pulmonalis proximalis - p.s. A. pulmonalis sinistra - p.sc. Paraganglion supracardiale superius - pe. Perikard - pl.c. Plexus caroticus communis - pl.e. Plexus caroticus externus - P.Z. Paraganglionäre Zellen - r.d. N. recurrens dexter - r.ph. R. pharyngicus - r.s. Rr. N. sympathici - R.Z. Zellen der Nebennierenrinde - scl.d. A. subclavia dextra - scl.s. A. subclavia sinistra - t.bi. Truncus bicaroticus - t.s. Truncus N. sympathici - tr. Trachea - v.cd. V. cava caudalis - v.cr.d. V. cava cranialis dextra - v.cr.s. V. cava cranialis sinistra - v.p. V. pulmonalis - IX N. glossopharyngicus - X N. vagus - XI N. accessorius - XII N. hypoglossus Mit 14 Textabbildungen.     

Analysis of a bone metastasis gene expression signature in patients with bone metastasis from solid tumors

Bone is a major target for metastases in the most frequent solid tumors, which result in severe complications and are a major cause of pain. A bone metastasis gene expression signature was identified using human breast cancer cells in a mouse model. The bone metastasis-related genes encode secretory and cell surface proteins implicated in bone-homing (CXCR4), angiogenesis (CTGF and FGF5), invasion (MMP-1 and ADAMTS1), and osteoclast recruitment (IL11). This signature superimposes on the 70-gene poor prognosis gene expression signature for breast cancer, and only ADAMTS1, CTGF and IL11 were found to be overexpressed in human primary breast cancers with bone relapse. We analyzed the expression of the six bone metastasis-related genes in bone metastases from patients with different types of solid tumors, to assess its relevance in human clinical samples. MMP-1, CXCR4, FGF5 and CTGF were found to be overexpressed in tumor cells of human bone metastases when compared to a human normal epithelial cell line. All the analyzed genes were overexpressed in the tumor cells of breast cancer bone metastases when compared to normal breast tissue. We did not detect any differences between the expression of these genes in bone metastases from breast cancer or from other types of solid tumors. Importantly, there was a significant correlation between the expressions of IL11/CTGF, IL11/ADAMTS1, CTGF/CXCR4, CTGF/ADAMTS1, and MMP-1/ADAMTS1, supporting the cooperative function of these proteins in the bone microenvironment, and the potential functional role of these genes in the establishment of bone metastases in vivo.     

Roles of SGS1, MUS81, and RAD51 in the repair of lagging-strand replication defects in Saccharomyces cerevisiae

Yeast cells lacking the SGS1 DNA helicase and the MUS81 structure-specific endonuclease display a synthetic lethality that is suppressed by loss of the RAD51 recombinase. This epistatic interaction suggests that the primary function of SGS1 or MUS81, or both genes, is downstream of RAD51. To identify RAD51-independent functions of SGS1 and MUS81, a synthetic-lethal screen was performed on the sgs1 mus81 rad51triple mutant. We found that mutation of RNH202, which encodes a subunit of the hetero-trimeric RNase H2, generates a profound synthetic-sickness in this background. RNase H2 is thought to play a non-essential role in Okazaki fragment maturation. Cells lacking RNH202 showed synthetic growth defects when combined with either mus81 or sgs1 alone. But, whereas the loss of RAD51 had little effect on rnh202 sgs1 double mutants, it strongly inhibited the growth of rnh202 mus81 cells. These data indicate that the primary function of SGS1, but not MUS81, is downstream of RAD51. SGS1 must have some RAD51-independent function, however, since the growth of rnh202 mus81 rad51cells was further compromised by the loss of SGS1. Consistent with these results, we show that rnh202 cells display a sensitivity to DNA-damaging agents that is exacerbated in the absence of RAD51 or MUS81. These data support a model in which defects in lagging-strand replication are repaired by the Mus81 endonuclease or through a pathway dependent on Rad51 and Sgs1.