The role of learning by a predator, Rivulus hartii, in the rare-morph survival advantage in guppies.

Negative frequency-dependent selection (NFDS), where rare types are favoured by selection, can maintain diversity. However, the ecological processes that mediate NFDS are often not known. Male guppies (Poecilia reticulata) exhibit extreme diversity of colour patterning and, in a previous field experiment, rare morphs had a survival advantage. Here, we test the hypothesis that predators impose NFDS because they are efficient at capturing familiar prey morphs, but are less efficient at capturing unfamiliar morphs. Over a series of trials, we presented Rivulus hartii, a natural predator of guppies, with male guppies with the same colour patterning (A trials); then, for a second series of trials, we presented the rivulus with guppies with a new colour pattern (B trials). The success of rivulus at capturing guppies on the first attack increased over successive A trials. First attack success decreased significantly for the early B trials, and then increased during successive B trials, eventually reaching the same level as in the best A trials. This experiment demonstrates that learning, perhaps through long-term search image formation, plays a role in predation success on familiar vs. unfamiliar prey morphs. These results support the hypothesis that predator learning contributes to the maintenance of the extreme male guppy polymorphism seen in nature.

Nonallelic behavior of rabbit variable-region allotypes.

Group a allotypes not detected by qualitative typing or anticipated from breeding data (latent allotypes) were detected at low levels in 50% of normal rabbit sera tested. The latent allotypes, which were serologically identical to allotypes of pooled IgG, were detected in sera from rabbits with all possible combinations of group a allotypes and their occurrence in individual rabbits was transitory and sporadic. These findings give reason to question the assumption that the synthesis of immunoglobulin allotypes is directed by allelic structural genes.

Selection at the MHC class IIB locus across guppy (Poecilia reticulata) populations.

The highly diverse genes of the major histocompatibility complex (MHC) are important in the adaptive immune system and are expected to be under selection from pathogens. Thus, the MHC genes provide an exceptional opportunity to investigate patterns of selection within and across populations. In this study, we analyzed genetic variation at the MHC class IIB gene and six microsatellite loci across 10 populations of guppies (Poecilia reticulata) in the northern range of Trinidad. We found a high level of diversity at the MHC, with a total of 43 alleles in 142 individuals. At the population level, we found that neutral evolution could not fully account for the variability found at the MHC. Instead, we found that MHC F(ST) statistics were lower than F(ST) derived from the microsatellite loci; 33 of 45 population pairwise estimates for the MHC were significantly lower than those for the microsatellite loci, and MHC F(ST) estimates were consistently lower than those predicted by a coalescent model of neutral evolution. These results suggest a similar selection acting across populations, and we discuss the potential roles of directional and balancing selection. At the sequence level, we found evidence for both positive and purifying selection. Furthermore, positive selection was detected within and adjacent to the putative peptide-binding region (PBR) of the MHC. Surprisingly, we also found a purifying selection at two sites within the putative PBR. Overall, our data provide evidence for selection for functional diversity at the MHC class IIB gene at both the population and nucleotide levels of guppy populations.

MHC class IIB additive and non-additive effects on fitness measures in the guppy Poecilia reticulata.

The genetic architecture of fitness at the class IIB gene of the major histocompatibility complex (MHC) in the guppy Poecilia reticulata was analysed. Diversity at the MHC is thought to be maintained by some form of balancing selection; heterozygote advantage, frequency-dependent selection or spatially and temporally fluctuating selection. Here these hypotheses are evaluated by using an algorithm that partitions the effect of specific MHC allele and genotypes on fitness measures. The effect of MHC genotype on surrogate measures of fitness was tested, including growth rate (at high and low bulk food diets), parasite load following a parasite challenge and survival. The number of copies of the Pore_a132 MHC allele was inversely related to infection by Gyrodactylus flukes and it appeared to be positively related to faster growth. Also, genotypes combining the Pore_a132 or other relatively common alleles paired with rare MHC alleles produced both advantageous and detrimental non-additive effects. Thus, the genetic architecture underlying fitness at the MHC is complex in the P. reticulata.

Analysis of the effects of polyphenols on human spermatozoa reveals unexpected impacts on mitochondrial membrane potential, oxidative stress and DNA integrity; implications for assisted reproductive technology.

The need to protect human spermatozoa from oxidative stress during assisted reproductive technology, has prompted a detailed analysis of the impacts of phenolic compounds on the functional integrity of these cells. Investigation of 16 individual compounds revealed a surprising variety of negative effects including: (i) a loss of mitochondrial membrane potential (Δψm) via mechanisms that were not related to opening of the permeability transition pore but associated with a reduction in thiol expression, (ii) a decline in intracellular reduced glutathione, (iii) the stimulation of pro-oxidant activity including the induction of ROS generation from mitochondrial and non-mitochondrial sources, (iv) stimulation of lipid peroxidation, (v) the generation of oxidative DNA damage, and (vi) impaired sperm motility. For most of the polyphenolic compounds examined, the loss of motility was gradual and highly correlated with the induction of lipid peroxidation (r=0.889). The exception was gossypol, which induced a rapid loss of motility due to its inherent alkylating activity; one consequence of which was a marked reduction in carboxymethyl lysine expression on the sperm tail; a post-translational modification that is known to play a key role in the regulation of sperm movement. The only polyphenols that did not appear to have adverse effects on spermatozoa were resveratrol, genistein and THP at doses below 100µM. These compounds could, therefore, have some therapeutic potential in a clinical setting.

Identification of alpha-carboxamidated and carboxy-terminal glycine forms of peptides in bovine hypothalamus, bovine pituitary and porcine heart extracts.

Two chemical assays have been developed for identifying and quantifying peptides which either could be biologically active by virtue of their alpha-carboxamidation or could be substrates for peptidylglycine alpha-amidating mono-oxygenase. The first assay is specific for the alpha-carboxamide of peptides. Using bis[trifluoroacetoxy]iodobenzene, the alpha-carboxamide was converted via a Hoffman reaction into a primary amine, which was then quantified by ninhydrin. The second assay is specific for glycine at the carboxy-terminus of a peptide. Glycine at the carboxy-terminus was derivatized to form 2-thiohydantoin, which was then separated and quantified by reverse phase HPLC. These assays were used to detect peptides in HPLC-separated extracts of bovine hypothalamus, bovine anterior lobe pituitary and porcine heart which may be of biological interest.

HPLC purification and separation of 5,5'-substituted-2,4-imidazolidinedithiones.

Several 5,5'-substituted-2,4-imidazolidinedithiones, synthesized from aldehydes or ketones, have been purified by HPLC using poly(styrene-divinylbenzene) packings. Purified 5,5'-substituted-2,4-imidazolidinedithiones have been identified in column effluent by UV absorbance and corroborated by mass spectrometry. Several silica-based, polymeric, and poly(styrene-divinylbenzene)-based packings were evaluated as matrices for resolution of a mixture of purified 5,5'-substituted-2,4-imidazolidinedithiones.

The relation of aberrant vasculogenesis to skeletal malformation in the hamster fetus.

Oral administration of teratogenic doses of retinoic acid to pregnant hamsters on day 10 of gestation is associated with dysmorphogenesis of the appendicular skeleton. During the 24 h following retinoic acid treatment, the developing limb bud vasculature was disorganized, with blood vessels encroaching on areas where mesenchymal condensation of the skeletal blastemata normally occurs. Large, branching marginal folds and endothelial cell vesiculations protruded into the blood vessel lumina. It is suggested that the vascular changes observed may affect the concurrent early development of the skeleton and contribute to the skeletal malformation seen in near-term fetuses.

Synthesis and fast-atom-bombardment-mass spectrometry of N-acetylmuramoyl-L-alanyl-D-isoglutamine (MDP).

N-Acetylmuramoyl-L-alanyl-D-isoglutamine (MDP) was synthesized by a series of condensations of appropriate reagents, followed by hydrogenolysis. Each intermediate step resulted in a stable, crystalline product. D-Isoglutamine 4-benzyl ester was condensed with N-(tert-butoxycarbonyl)-L-alanine N-hydroxysuccinimide ester, to give N-(tert-butoxycarbonyl)-L-alanyl-D-isoglutamine benzyl ester. Condensation of L-alanyl-D-isoglutamine benzyl ester with N-acetyl-1-O-benzyl-4,6-O-benzylidenemuramic acid, followed by hydrogenolysis, gave MDP. The synthetic scheme was shown to be capable of producing gram quantities of highly pure MDP, as well as a few of its analogs. The synthetic MDP was characterized by analytical and biological methods, and it was found that the use of fast-atom-bombardment-mass spectrometry may greatly simplify the characterization process.

The structure of the repeating oligosaccharide unit of the pneumococcal capsular polysaccharide type 18C.

The structure of the repeating oligosaccharide of the pneumococcal capsular polysaccharide type 18C has been investigated. The repeating oligosaccharide, isolated from an aqueous hydrofluoric acid hydrolyzate of the polysaccharide, was shown, by fast atom bombardment-mass spectrometry, to have a molecular weight of 928, and to contain an O-acetyl group and a glycerol residue. Information about the sequence in the per-O-methylated oligosaccharide was derived from electron-impact mass spectrometry. Supporting data were obtained from methylation analysis, periodate and chromium trioxide oxidations, and enzymic and acid hydrolyses of the oligosaccharide. These studies indicated that the polysaccharide consists of the following pentasaccharide repeating unit. (formula see text)

Mass-spectral studies of isomeric D-ribofuranosylribitol disaccharides from the capsular polysaccharides of Haemophilus influenzae type b and Escherichia coli K 100.

Two isomeric-D-ribofuranosylribitols, derived from capsular polysaccharides of Haemophilus influenzae type b and Escherichia coli K 100, were methylated or acetylated, and the products analyzed by gas-liquid chromatography-mass spectrometry. The marked difference in the mass spectra of the methyl ethers of these disaccharides allowed clear distinction between 1- and 2-O-D-ribofuranosylribitol was characteristic for this disaccharide; its isomer, the (1 leads to 2)-linked species, has a base peak at m/e 57. The difference in the base peaks is attributable to fragmentation of the methylated ribitol, as both spectra display common ions characteristic of the methylated D-ribofuranosyl group. For the acetylated disaccharides, the mass spectra displayed common ions characteristic of the acetylated D-ribofuranosyl group. However, no ions similar to those found for the methylated ribitol allowed ready differentiation between the two acetates. Instead, their spectra displayed similar ions, differing somewhat in relative abundance; the M-1 ion, m/e 577, was obtained for both. Comparison of the relative abundance of m/e 139, 259, and 303 in the spectra of the two acetates did allow distinction between them.

Evaluation of physical therapy service for severely mentally impaired students with cerebral palsy.

The effects of two types of physical therapy service were studied over a five-month period in 19 severely mentally impaired cerebral palsied students aged 3 to 22 years. The students were paired and assigned to either a direct therapy treatment group or a supervised therapy management group. Ten similar students in a comparison group received no physical therapy. The study showed no significant difference in development of mature developmental reflexes, improvement of gross motor skills, or increase of passive joint motion among similar students placed in the direct, supervised, and comparison groups. This pilot study is not intended to decide the issue of physical therapy for these students. It does, however, indicate that objective tests may be used reliably to measure three major areas of sensory motor development. More extensive and longer-term studies are needed to determine the most beneficial form of physical therapy for this double-disability population.

Development of a physical characteristics assessment (PCA): a checklist for determining appropriate computer access for individuals with cerebral palsy.

The physical characteristics assessment (PCA), a newly developed illustrated checklist, is designed for use by allied health and education professionals to identify a viable method for an individual with cerebral palsy to access a computer. The PCA was developed in response to a documented void of such assessments in the literature. The value of PCA was assessed with respect to content validity, clinical utility, reliability (inter-rater), and opinion of persons with cerebral palsy and health and education professionals. Professionals (n = 72) and persons with cerebral palsy (n = 78) from 20 sites in Canada, Chile, England, Israel, Spain, and the United States participated in the study. Statistically, the PCA produced a highly significant (p < 0.001) assessment time savings compared with non-PCA methods. A statistically significant difference in the number of user/computer interface changes during five post-assessment computer training sessions was not shown between the PCA and non-PCA methods. Inter-rater reliability ranged between 90% and 100% (n = 33 pairs). Both persons with cerebral palsy assessed via the PCA and professionals using the PCA gave positive opinions. Overall results support the value of the PCA.

Anaphylaxis to cisatracurium following negative skin testing.

General anaesthesia for the patient with a history of anaesthesia-related anaphylaxis is challenging. Precautions against anaphylaxis and the use of skin test negative drugs can reduce but not eliminate the risk. In the majority of such cases, subsequent anaesthesia is uneventful. However, the absence of a clearly identified triggering agent increases the difficulties facing the anaesthetist. We present a case of anaphylaxis to cisatracurium following a negative skin test.

Vasculogenesis and chondrogenesis in the chick wing bud.

Stage 22 to stage 24 chick wing buds were labeled with sulphate-35 to identify chondrogenic areas; these areas were then related to the presence of blood vessels. Blood vessels were found in areas of increased sulphate uptake, indicating that there is no avascular area formed in the limb bud mesenchyme prior to the onset of chondrogenesis, as indicated by increased sulphate-35 uptake. It is, therefore, unlikely that the vasculature plays an initial role in the determination of chondrogenic areas in the wing bud.

Peptide studies using a fast atom bombardment high field mass spectrometer and data system. 4. Disulfide-containing peptides.

Ten peptides containing one, two or three disulfides were examined to determine their behavior under fast atom bombardment (FAB) mass spectrometric conditions. The mass spectra for the disulfide and the reduced disulfide forms of each peptide were compared. Several factors were examined that contribute to the fast atom bombardment mass spectra of these peptides: components of the FAB matrix such as alkali cations, acids, bases and reducing agents, the intrinsic molecular properties of the intact peptide, and the effect of reducing conditions on sensitivity. The FAB mass spectra of the disulfide-containing peptides examined in this study displayed accurate molecular weight information and fragmentation which indicated the position of the disulfide in the amino acid sequence.

Ultimobranchial body cysts in the human foetal thyroid: pathological implications.

In a histological study of 28 human foetal thyroids, ultimobranchial body cysts were found in seven foetuses. Large oval cells were seen adjacent to or within the cysts. There is a morphological resemblance between the cells associated with the ultimobranchial cysts and those of medullary carcinoma of the thyroid gland. This similarity, in humans, substantiates the hypothesis that medullary carcinoma arises from the parafollicular cells, which themselves are known to differentiate from ultimobranchial body tissue.

Use of immobilized exopeptidases and volatile buffers for analysis of peptides by fast atom bombardment mass spectrometry.

beta-Lipotrophin (62-77) or Ac-gastrin releasing peptide was incubated with immobilized carboxypeptidase Y or aminopeptidase M. Subsequent aliquots of each incubation mixture were analysed by fast atom bombardment mass spectrometry using a dithiothreitol/dithioerythritol liquid matrix. The use of immobilized enzymes and volatile buffers for exopeptidase digestions enabled rapid and facile separation of enzyme from digestion products. This approach to mass spectral peptide analysis reduced spectral background arising from a glycerol matrix, buffer salts, or enzyme proteins and contaminants, enabling analysis of as little as 200 picomoles of a suitable peptide.

The effect of retinoic acid on chondrogenesis in the fetal hamster tibia in vivo.

Chondrogenesis in the fetal hamster tibia was examined 24, 36 and 48 h after maternal administration of retinoic acid at a known teratogenic dose (80 mg/kg) on day 10 of gestation. Twenty-four hours after treatment the tibial chondroblasts were more closely packed with less intervening fibrillar or granular matrix than in the controls. By 36 h post treatment, an intercellular matrix containing wide clumped fibrils unassociated with matrix granules had begun to accumulate, while the condroblasts had a relatively poorly developed Golgi apparatus and smooth, non-scalloped cell outlines. Cells frequently maintained contact chondroblasts appeared similar to controls of the same age. It is suggested that the paucity and abnormality of the extracellular matrix prevents the chondroblasts from assuming their normal spatial relations with each other, resulting in a small skeletal blastema.

Analysis of peptides containing oxidized methionine and/or tryptophan by fast atom bombardment mass spectrometry.

Treatment of peptides containing methionine and/or tryptophan with dimethylsulfoxide/hydrochloric acid/acetic acid resulted in oxidation of these amino acids respectively to methionine sulfoxide and oxyindolalanine. This reaction was monitored by fast atom bombardment mass spectrometry using a dithiothreitol/dithioerythritol liquid matrix. Under these conditions, only methionine and tryptophan were oxidized. Comparison of mass spectra of a sample before and after oxidation should provide a rapid screening procedure for determination of these residues in peptides.