Behavioural cues of reproductive status in seahorses Hippocampus abdominalis.

A method is described to assess the reproductive status of male Hippocampus abdominalis on the basis of behavioural traits. The non-invasive nature of this technique minimizes handling stress and reduces sampling requirements for experimental work. It represents a useful tool to assist researchers in sample collection for studies of reproduction and development in viviparous syngnathids, which are emerging as important model species.

Identification of a uniquely southern African clade of coastal pipefishes Syngnathus spp.

The taxonomic status of two southern African coastal pipefish species, Syngnathus temminckii and Syngnathus watermeyeri, was investigated using a combination of morphological and genetic data. Morphological data showed that S. temminckii is distinct from the broadly distributed European pipefish Syngnathus acus, and a molecular phylogeny reconstructed using mitochondrial DNA recovered S. temminckii and S. watermeyeri as sister taxa. The southern African species share an evolutionary origin with north-eastern Atlantic Ocean and Mediterranean Sea species, including S. acus. These data support the existence of a distinct southern African clade of Syngnathus pipefishes that has diverged in situ to form the two species present in the region today.

The evolutionary origins of Syngnathidae: pipefishes and seahorses.

Despite their importance as evolutionary and ecological model systems, the phylogenetic relationships among gasterosteiforms remain poorly understood, complicating efforts to understand the evolutionary origins of the exceptional morphological and behavioural diversity of this group. The present review summarizes current knowledge on the origin and evolution of syngnathids, a gasterosteiform family with a highly developed form of male parental care, combining inferences based on morphological and molecular data with paleontological evidence documenting the evolutionary history of the group. Molecular methods have provided new tools for the study of syngnathid relationships and have played an important role in recent conservation efforts. Despite recent insights into syngnathid evolution, however, a survey of the literature reveals a strong taxonomic bias towards studies on the species-rich genera Hippocampus and Syngnathus, with a lack of data for many morphologically unique members of the family. The study of the evolutionary pressures responsible for generating the high diversity of syngnathids would benefit from a wider perspective, providing a comparative framework in which to investigate the evolution of the genetic, morphological and behavioural traits of the group as a whole.

Sexual size dimorphism predicts the frequency of multiple mating in the sex-role reversed pipefish Syngnathus typhle.

The sex-role reversed pipefish Syngnathus typhle is a member of the Syngnathidae, a family of fishes in which males brood embryos on their body surface. As in most ectotherms, embryonic development is highly temperature dependent in syngnathids and male brooding periods are extended when water temperatures are reduced. The influence of temperature on reproduction is expected to effectively truncate the breeding season and reduce fecundity in cold waters, potentially enhancing the opportunity for both fecundity and sexual selection. We studied spatial variation in the morphology and reproductive biology of S. typhle in five European populations which vary in latitude and water temperature. Microsatellite analyses indicated that the average number of male mates per population ranged between 1.3 and 3.7. The frequency of multiple mating by males was negatively correlated with the degree of sexual size dimorphism in each population, suggesting that disproportionate increases in female fecundity may be able to compensate for increased male brood pouch capacity. Both sexes were larger and males had an increased brood size where water temperatures during the breeding season were lower. Morphological variation among populations may be mediated by differences in fecundity selection associated with different optimal reproductive strategies in cold and warm water environments.

Prediction of reduction in aggressive behaviour of growing pigs using skin lesion traits as selection criteria.

Aggression at regrouping is a common issue in pig farming. Skin lesions are genetically and phenotypically correlated with aggression and have been shown to have a significant heritable component. This study predicts the magnitude of reduction in complex aggressive behavioural traits when using lesion numbers on different body regions at two different time points as selection criteria, to identify the optimum skin lesion trait for selection purposes. In total, 1146 pigs were mixed into new social groups, and skin lesions were counted 24 h (SL24h) and 3 weeks (SL3wk) post-mixing, on the anterior, centre and posterior regions of the body. An animal model was used to estimate genetic parameters for skin lesion traits and 14 aggressive behavioural traits. Estimated breeding values (EBVs) and phenotypic values were scaled and standardised to allow direct comparison across multiple traits. Individuals with SL24h and SL3wk EBVs in the least aggressive 10% of the population were compared with the population mean to predict the expected genetic and phenotypic response in aggressive behaviour to selection. At mixing, selection for low anterior lesions was predicted to affect substantially more behavioural traits of aggressiveness than lesions obtained on other body parts, with EBVs between -0.21 and -1.17 SD below the population mean. Individuals with low central SL24h EBVs also had low EBVs for aggressive traits (-0.33 to -0.55). Individuals with high SL3wk EBVs had low EBVs for aggression at mixing (between -0.24 and -0.53 SD below the population mean), although this was predicted to affect fewer traits than selection against SL24h. These results suggest that selection against anterior SL24h would result in the greatest genetic and phenotypic reduction in aggressive behaviour recorded at mixing. Selection for increased SL3wk was predicted to reduce aggression at mixing; however, current understanding about aggressive behaviour under stable social conditions is insufficient to recommend using this trait for selection purposes.

The relationship between body dimensions of living pigs and their carcass composition.

The performance of a visual image analysis (VIA) system was tested with regards to its potential to determine in vivo carcass composition and conformation, either alone, or in conjunction with other in vivo measures such as live weight and backfat depth. Pigs of both sexes of a commercial type were reared and slaughtered at weights ranging from 50 to 120 kg. Feeding was ad libitum on diets ranging from 0.14 to 0.19 kgkg(-1) crude protein content to produce animals of a range of body condition. Two analyses were carried out: the first analysis addressed the relationship between dimensionless carcass and VIA indices; the second analysis assessed the relationship between carcass composition and VIA body shape using detrended carcass and VIA data, which were produced by removal of allometric growth trends. A statistically significant relationship (P<0.05) between in vivo VIA body size and shape and carcass muscle dimensions and composition was found for most body regions. Adjusted R(2) statistics ranged between 0.13 and 0.54 for relative fat weights and between 0.14 and 0.51 for relative lean weights. The predictive power of the regression models, indicated by R(2)-like statistics for prediction, was approximately 70% of the adjusted R(2) values. The descriptive and predictive powers of the corresponding models generally strengthened if VIA indices were combined with other in vivo measurements. The relationships between in vivo and carcass measures remained statistically significant (P<0.05) after removal of the growth trends, although adjusted R(2) statistics generally decreased. The predictive power of models corresponding to the detrended measures was, however, weak. The results show in vivo VIA measurements to be useful in the estimation of muscle size, carcass conformation and composition, all of which are of significant importance to the pig production, marketing and processing industries.

Genetic associations of short- and long-term aggressiveness identified by skin lesion with growth, feed efficiency, and carcass characteristics in growing pigs.

The objective of this study was to investigate the genetic relationships between skin lesion traits in group housed growing pigs as a measure of short- (in a newly mixed group) and long- (in a socially stable group) term aggression and commonly used commercial performance measures: growth, feed intake, feed efficiency, and carcass traits. Data on 2,413 growing pigs (138 groups) were available. Pigs were mixed into new social groups of 18 animals, and skin lesions were counted 24 h (SL24h) and 5 wk (SL5wk) postmixing. The animal model was used to estimate genetic parameters for skin lesion traits, test daily gain, lifetime daily gain, daily feed intake, feed efficiency (calculated as test daily gain divided by daily feed intake), loin depth, back fat, and HCW. Skin lesions had a heritable component, ranging from 0.08 for anterior SL24h to 0.22 for central SL5wk and would, therefore, be suitable as a method of phenotyping aggression for selection purposes. Significant positive genetic correlations were found between SL24h and SL5wk (0.46 to 0.81). Positive genetic correlations were also found between SL24h (central and posterior body regions) or SL5wk (all body regions) and the production traits lifetime daily gain, test daily gain, and HCW (0.29 to 0.54). Central SL24h, anterior SL5wk, and posterior SL5wk were found to correlate positively with feed efficiency (0.39 to 0.50), suggesting that pigs with more lesions convert feed more efficiently. Where significant, the magnitude of phenotypic correlations was low but positive (0.07 to 0.10). These results suggest that, genetically, animals that receive many lesions show improved performance compared to those with few lesions, except for anterior SL24h, which had previously been shown to be genetically positively correlated with the initiation of nonreciprocal attacks. It may, therefore, be possible, via selection against anterior skin lesions at mixing, to reduce this form of 1-sided aggression without adversely affecting production traits.

Increased affinity of guinea pig thymocytes and thymus-dependent lymphocytes for papain-treated rabbit erythrocytes compared to untreated erythrocytes.

The affinity of guinea pig thymocytes and peripheral T lymphocytes for rabbit erythrocytes was found to be enhanced following treatment of the erythrocytes with papain. By increasing the numbers of rosette-forming cells and giving stronger, more stable rosettes, this procedure increases the usefulness of the reaction as a T cell marker in guinea pigs.

Discrimination between innate and cytophilic immunoglobulin on human peripheral blood lymphocytes: analysis by the direct antiglobulin rosette-forming reaction.

Bovine red blood cells linked to polyclonal or monoclonal anti-immunoglobulin antibodies are used in the direct antiglobulin rosetting reaction to detect surface-Ig on human lymphocytes. The sensitivity of this test is markedly increased by pretreating the red cells with trypsin. Enzyme-treated red cells, coupled to anti-human Fab or anti-light chain antibodies, react not only with innate Ig on B lymphocytes but also with smaller amounts of passively adsorbed, cytophilic Ig on up to 25% of freshly prepared peripheral blood (non-B) lymphocytes. In contrast, trypsinized red cells carrying anti-Ig isotype-specific antibodies react exclusively with B cell surface-Ig. Cytophilic Ig is abnormally firmly bound to lymphocytes separated on Ficoll-Hypaque at 20 degrees C or below, and is released very slowly during 3 h or more at 37 degrees C in vitro. Lymphocytes are free of detectable cytophilic Ig when isolated on Ficoll-Hypaque at 37 degrees C, and very little Ig is retained by non-B cells in suspensions purified on Percoll which, unlike Ficoll, does not increase Ig binding affinity. These lymphocyte separation procedures are recommended as a preliminary to B cell assays by sensitive antiglobulin techniques.

Artefactual variations in the B and T subpopulations of rabbit blood lymphocytes depending on method of isolation, and blocking of C3 receptors due to in vitro activation of complement.

Marked differences were found in the proportions of lymphocyte subpopulations in rabbit peripheral blood depending on the techniques used for the purification of lymphocytes. Rosette-forming reactions were used to find the numbers of T lymphocytes, Ig-bearing cells and cells with receptors for C3 or IgG-Fc. Some of the methods used for lymphocyte separation altered the relative numbers of T and B lymphocytes, through a disproportionate loss of T cells. Other changes were due to in vitro activation of complement detectable by the presence of C3 on the lymphocyte cell-membrane and causing partial blocking of C3 receptors. Highest yields of lymphocytes were obtained from defibrinated blood treated with carbonyl iron to remove phagocytes and methyl cellulose to sediment erythrocytes. This procedure was accompanied by in vitro activation of complement, with the consequences mentioned. Complement activation was inhibited by taking the blood into either EDTA or citrate. As EDTA was cytotoxic for rabbit T lymphocytes, citrate was considered best although the resulting lymphocyte suspensions were contaminated with up to 25% granulocytes and monocytes owing to the inhibition of carbonyl iron uptake by the prior exposure to citrate.

Direct antibody rosette-forming reactions using monoclonal markers of lymphocyte subpopulations. Methodology and applications illustrated by investigations with rat pan-T antibodies of the CAMPATH series.

A sensitive direct antibody rosette assay has been developed for the detection of antigens on the lymphocyte cell membrane. Indicator cells for rosette tests were prepared by chromic chloride coupling of rat or mouse monoclonal IgG or IgM anti-lymphocyte antibodies to untreated or trypsinized bovine red blood cells. The monoclonal antibodies used were reactive with a range of cell surface markers which identify various lymphocyte subpopulations, including T cell antigens, HLA class II (Ia-like antigens), Leu-7 (HNK-1) and VEP 13, a determinant of Fc gamma receptors on large granular lymphocytes. Results obtained by direct rosette formation correlated well with those of parallel tests using indirect immunofluorescent antibodies staining. Several applications of the direct antibody rosetting procedure are described in further investigations with a series of pan-T monoclonal (CAMPATH) antibodies. These include the morphological examination of antibody-binding cells in cytocentrifuge smears, the separation of lymphocyte subsets by density gradient centrifugation, and the use of a rosette inhibition assay to identify monoclonal antibodies binding to the same (or closely associated) epitopes of the lymphocyte cell membrane.

A competitive inhibition ELISA for the quantification of human interferon-gamma.

A competitive enzyme immunoassay has been developed for the measurement of human interferon-gamma (IFN-gamma) in cell culture supernatants. The assay is based on the dose-dependent inhibitory effect of liquid phase IFN-gamma on the binding of a specific monoclonal antibody to recombinant IFN-gamma (rIFN-gamma) immobilized on microtitre plate wells. The extent of monoclonal anti-IFN-gamma inhibition was determined by the uptake of alkaline phosphatase-conjugated goat anti-mouse IgG and the subsequent development of enzyme substrate colour. Absorbance readings were taken and results for test samples were extrapolated from standard rIFN-gamma inhibition curves constructed as logit-log plots. Assay performance was assessed using three different monoclonal antibodies (clones 20G7, H-22 and GZ-4). Optimum sensitivity was achieved with the antibodies of higher affinity, 20G7 and H-22, which gave reliable quantification of IFN-gamma over a wide range of concentrations from 0.4 ng/ml (3.4 IU/ml), or less, to 250 ng/ml approximately 2000 IU/ml). The inhibition assay incorporates the advantages of specificity, reproducibility and convenience of performance which are the hallmarks of monoclonal antibody-based ELISAs. However, compared to the sandwich ELISAs previously described for human IFN-gamma, it is considerably more economical in its use of monoclonal anti-IFN-gamma, requiring < 50 ng of a single antibody per 96 well plate. It also uses relatively small volumes of test samples (50 microliters/well) which is particularly advantageous where limited amounts of cell culture supernatant are available for cytokine assays.

Incipient speciation in sympatric Nicaraguan crater lake cichlid fishes: sexual selection versus ecological diversification.

The growing body of empirical evidence for sympatric speciation has been complemented by recent theoretical treatments that have identified evolutionary conditions conducive to speciation in sympatry. The Neotropical Midas cichlid (Amphilophus citrinellum) fits both of the key characteristics of these models, with strong assortative mating on the basis of a colour polymorphism coupled with trophic and ecological differentiation derived from a polymorphism in their pharyngeal jaws. We used microsatellite markers and a 480 bp fragment of the mitochondrial DNA control region to study four polymorphic populations of the Midas cichlid from three crater lakes and one large lake in Nicaragua in an investigation of incipient sympatric speciation. All populations were strongly genetically differentiated on the basis of geography. We identified strong genetic separation based on colour polymorphism for populations from Lake Nicaragua and one crater lake (Lake Apoyo), but failed to find significant genetic structuring based on trophic differences and ecological niche separation in any of the four populations studied. These data support the idea that sexual selection through assortative mating contributes more strongly or earlier during speciation in sympatry than ecological separation in these cichlids. The long-term persistence of divergent cichlid ecotypes (as measured by the percentage sequence divergence between populations) in Central American crater lakes, despite a lack of fixed genetic differentiation, differs strikingly from the patterns of extremely rapid speciation in the cichlids in Africa, including its crater lakes. It is unclear whether extrinsic environmental factors or intrinsic biological differences, e.g. in the degree of phenotypic plasticity, promote different mechanisms and thereby rates of speciation of cichlid fishes from the Old and New Worlds.

Determination of CD59 protein in normal human serum by enzyme immunoassay, using octyl-glucoside detergent to release glycosyl-phosphatidylinositol-CD59 from lipid complex.

In this study we have optimised the enzyme immunoassay (ELISA) to quantify CD59 antigen in human serum or plasma. The glycosyl-phosphatidylinositol (GPI)-linked form of CD59 is known to complex with serum high-density lipoprotein. For ELISA optimisation, therefore, we investigated the effect of detergents, added to the sample diluent, on the determined values of CD59. Values obtained in the presence of octyl-glucoside (OG) for 20 adults aged 18-35 years and 17 children 1-5 years old were, respectively, 33-119 ng/ml (mean +/- S.D.: 66+/-22 ng/ml) and 37-143 ng/ml (76+/-33 ng/ml). These results were higher than those measured without OG and were in contrast with published results showing absence, or eight to nine times lower levels, of the protein in serum. A known range for serum concentrations of CD59 in healthy individuals will establish an important reference point for clinical work and for the investigation of diseases involving the complement membrane attack complex (MAC) and its regulation.

Factors influencing the inhibitory action of anti-inflammatory drugs on carrageenin induced oedema.

1. In a series of experiments carried out over 2(1/2) years, the inflammatory response of the rat hind paw to injected carrageenin was relatively constant, but the anti-inflammatory activity of phenylbutazone was subject to wide variation.2. Phenylbutazone, aspirin and indomethacin were all well absorbed after oral administration, and the resultant plasma drug concentrations were closely similar to those produced when the drugs were administered intraperitoneally.3. The anti-inflammatory effect of the drugs on carrageenin oedema was variable and poorly correlated with log dose and plasma concentration.4. There was a pronounced and linearly correlated increase in the anti-inflammatory activity of phenylbutazone with increasing ambient temperature in the range 5-30 degrees C.5. Variations in relative humidity at constant temperature (20 degrees C) did not influence the anti-oedema potency of phenylbutazone.6. For the precise evaluation of anti-inflammatory activity in carrageenin oedema tests, it is concluded that accurate control of temperature is essential.

Quantitative analysis on isolated organs of the autonomic blocking properties of indoramin hydrochloride (Wy 21901).

1. Indoramin is a competitive alpha-adrenoceptor blocking agent on the guinea-pig isolated vas deferens and aortic strip; the pA(2) value on the aorta is 7.4.2. Indoramin is devoid of beta-adrenoceptor blocking activity on the guinea-pig isolated trachea (10(-4)M) and the Langendorff preparation of the rabbit heart (10(-6)M). It also has no adrenergic neurone blocking action on the Finkleman preparation of the rabbit small intestine (10(-5)M).3. On the Langendorff preparation of the rabbit isolated heart, indoramin (above 10(-6)M) has cardio-inhibitory properties similar to those of propranolol.4. On the guinea-pig isolated ileum (10(-5)M) and trachea (10(-6)M), indoramin is devoid of anticholinergic activity, but has a potent antihistamine action which satisfies the criteria for competitive antagonism; the pA(2) value for this antagonism on the ileum is 8.2.5. Indoramin antagonizes 5-hydroxytryptamine on the rat isolated fundus and ileum; the pA(2) value for the antagonism on the ileum is 6.0.

The evaluation of an intervention program for young schoolchildren with acting-out problems.

The rationale and nature of a program to train nonprofessional child-aides for helping interactions with young acting-out school children are described. Pre/post teacher measures of children's problems and competencies, aide measures of problem behavior, and school mental health professionals' change in behavior estimates were used to evaluate the program's effectiveness. Children seen by trained aides showed significantly greater reductions both in acting-out problems and in overall maladjustment than did similar children seen by comparable aides who did not have additional training, or by themselves before training. Implications were considered for optimizing outcomes both in the specific school intervention project in question and in other, broader types of clinical interventions.

Lateral rotatory dislocation of the ankle without fracture.

A 34-year-old woman sustained a closed lateral rotatory dislocation of the ankle without a fracture. Treatment was by closed reduction and cast immobilization for 6 weeks. After removal of the cast, her ankle was stable. Stress radiographs of both the affected and contralateral ankles were normal. One year after injury the patient demonstrated a full, pain-free, stable range of motion. The patient exhibited generalized ligamentous laxity, which may have contributed to her tendency to dislocate the talus within the mortise.

An in vivo and in vitro study of selenium deficiency and infection in rats.

Selenium deficiency in rats impairs the ability of neutrophils and peritoneal macrophages to kill Candida albicans organisms in vitro. In contrast, killing of Salmonella typhimurium and Staphylococcus aureus organisms is unaffected by the deficiency. Survival of rats after intraperitoneal injection of 8 X 10(7) S. aureus organisms was not affected by Se deficiency, but a 5-fold increase in the dose (4 X 10(8) S. aureus organisms) led to a significantly greater mortality in the Se deficient rats.

Mitochondrial abnormalities in muscle from vitamin B12-deficient sheep.

Skeletal and cardiac muscles from vitamin B12-deficient sheep were examined. Histochemical studies did not reveal any gross pathological changes in the muscle structure, but there was an abnormal distribution of the product of the NADH-diaphorase reaction. Electron microscopy revealed an abnormal distribution of mitochondria, changes in the number and arrangement of cristae within the mitochondria and the presence of inclusions. The possibility that alterations in mitochondrial morphology are early lesions attributable to metabolic changes in vitamin B12 deficiency is discussed.