Association between loci with deleterious alleles and distorted sex ratios in an inbred line of tilapia (Oreochromis aureus)

Three microsatellite markers (UNH159, UNH231, and UNH216) were examined for association with both deleterious genes and sex-ratio distortions in a full-sib family of 222 progeny from the fourth generation of a meiogynogenetic tilapia line (Oreochromis aureus). The three markers were mapped previously to different linkage groups and were shown to be associated with genes with deleterious alleles in this line. A restricted maximum likelihood model was used for analysis of major effects and their interactions on sex ratio and viability. This model was based on selective mortality of genders, ignoring effects of possible sex-determining genes. The results showed that deleterious genes linked to UNH216 and UNH231 exert higher lethality in females than in males (P < .0005 and P < .05, respectively). UNH159 was not associated directly with sex ratio distortion, but acts strongly as a modifier of sex ratio in combination with UNH216 and UNH231. Each of the three loci was found to have a significant effect on viability (P < .05) in the maximum likelihood analysis. The deleterious single-locus effects act strongly against females, while most of the epistatic interactions exert higher lethality in males. This contradiction results in a close to 1:1 sex ratio at maturity. The genetic mechanism and significance of such a balance between genders are still unknown. A detailed analysis of sex-specific lethality may be applied by screening in appropriate series of matings and fine mapping with additional markers. Our data suggest that UNH216 and UNH231 are linked to sex ratio distortion genes and that UNH159 may be linked to a modifier of these genes.     

Amh and Dmrta2 genes map to tilapia (Oreochromis spp.) linkage group 23 within quantitative trait locus regions for sex determination

Recent studies have revealed that the major genes of the mammalian sex determination pathway are also involved in sex determination of fish. Several studies have reported QTL in various species and strains of tilapia, regions contributing to sex determination have been identified on linkage groups 1, 3, and 23. Genes contributing to sex-specific mortality have been detected on linkage groups 2, 6, and 23. To test whether the same genes might control sex determination in mammals and fishes, we mapped 11 genes that are considered putative master key regulators of sex determination: Amh, Cyp19, Dax1, Dmrt2, Dmrta2, Fhl3l, Foxl2, Ixl, Lhx9, Sf1, and Sox8. We identified polymorphisms in noncoding regions of these genes and genotyped these sites for 90 individuals of an F2 mapping family. Mapping of Dax1 joined LG16 and LG21 into a single linkage group. The Amh and Dmrta2 genes were mapped to two distinct regions of LG23. The Amh gene was mapped 5 cM from UNH879 within a QTL region for sex determination and 2 cM from UNH216 within a QTL region for sex-specific mortality. Dmrta2 was mapped 4 cM from UNH848 within another QTL region for sex determination. Cyp19 was mapped to LG1 far from a previously reported QTL region for sex determination on this chromosome. Seven other candidate genes mapped to LG4, -11, -12, -14, and -17.     

Segregation of QTL for production traits in commercial meat-type chickens

This study investigated whether quantitative trait loci (QTL) identified in experimental crosses of chickens provide a short cut to the identification of QTL in commercial populations. A commercial population of broilers was targeted for chromosomal regions in which QTL for traits associated with meat production have previously been detected in extreme crosses. A three-generation design, consisting of 15 grandsires, 608 half-sib hens and over 15 000 third-generation offspring, was implemented within the existing breeding scheme of a broiler breeding company. The first two generations were typed for 52 microsatellite markers spanning regions of nine chicken chromosomes and covering a total of 730 cM, approximately one-fifth of the chicken genome. Using half-sib analyses with a multiple QTL model, linkage was studied between these regions and 17 growth and carcass traits. Out of 153 trait x region comparisons, 53 QTL exceeded the threshold for genome-wide significance while an additional 23 QTL were significant at the nominal 1% level. Many of the QTL affect the carcass proportions and feed intake, for which there are few published studies. Given intensive selection for efficient growth in broilers for more than 50 generations it is surprising that many QTL affecting these traits are still segregating. Future fine-mapping efforts could elucidate whether ancestral mutations are still segregating as a result of pleiotropic effects on fitness traits or whether this variation is due to new mutations.     

Genome-scan analysis for quantitative trait loci in an F<Subscript>2</Subscript> tilapia hybrid

We searched for genetic linkage between DNA markers and quantitative trait loci (QTLs) for innate immunity, response to stress, biochemical parameters of blood, and fish size in an F₂ population derived from an interspecific tilapia hybrid ( Oreochromis mossambicus × O. aureus). A family of 114 fish was scanned for 40 polymorphic microsatellite DNA markers and two polymorphic genes, covering ~80% of the tilapia genome. These fish had previously been phenotyped for seven immune-response traits and six blood parameters. Critical values for significance were P <0.05 with the false discovery rate (FDR) controlled at 40%. The genome-scan analysis resulted in 35 significant marker-trait associations, involving 26 markers in 16 linkage groups. In a second experiment, nine markers were re-sampled in a second family of 79 fish of the same species hybrid. Seven markers ( GM180, GM553 , MHC-I , UNH848 , UNH868 , UNH898 and UNH925) in five linkage groups (LG 1, 3, 4, 22 and 23) were associated with stress response traits. An additional six markers ( GM47, GM552 , UNH208 , UNH881 , UNH952 , UNH998) in five linkage groups (LG 4, 16, 19, 20 and 23) were verified for their associations with immune response traits, by linkage to several different traits. The portion of variance explained by each QTL was 11% on average, with a maximum of 29%. The average additive effect of QTLs was 0.2 standard deviation units of stress response traits and fish size, with a maximum of 0.33. In three linkage groups (LG 1, 3 and 23) markers were associated with stress response, body weight and sex determination, confirming the location of QTLs reported by several other studies.Communicated by G. Reuter     

Studies on mouse autoreactive cells: I. Role of H-2 antigens in mouse autologous rosette formation

A minority (1–2%) of normal mouse lymphoid cells bind autologous erythrocytes and form rosettes. In this study we examined the antigenic specificity involved in the formation of such rosettes. A significant difference in the incidence of rosettes formed, respectively, with autologous and allogeneic mouse erythrocytes is found. Moreover, preincubation of lymphoid cells with low concentrations of syngeneic erythrocytic ghosts causes significant competitive inhibition of subsequent rosette formation. Allogeneic ghosts obtained from nonrelated or from congenic resistant strains of mice do not display this inhibitory effect under the same conditions. It is thus suggested that mouse autologous rosette-forming cells bear receptors for syngeneic H-2 antigens that are involved in the binding of autologous erythrocytes. More precisely, compatibility between lymphocyte and erythrocyte restricted to K or D only is sufficient to ensure a level of rosettes similar to that obtained when complete identity occurs for K, I, and D regions.     

The genetic diversity of wild Oreochromis mossambicus populations from the Mozambique southern watersheds as evaluated by microsatellites

Tilapia is native to Africa, and one of the most cultivated fish in the world. The goal of this research was to evaluate the genetic diversity of tilapia Oreochromis mossambicus from the Limpopo, Incomati, Umbeluzi and Sabié rivers in Mozambique. Microsatellite markers were used to assess the genetic structure and to compare the genetic variability of wild populations of O. mossambicus. DNA samples from 200 specimens were analyzed. All five loci (UNH104, UNH129, UNH142, UNH222 and UNH231) used in this study were polymorphic, with observed heterozygosity ranging from 0.940 to 1.000 and the allelic richness average (Ar) ranging from 8.937 to 15.751. All of the stocks exhibited a remarkably significant excess of heterozygosity relative to the Hardy‐Weinberg Equilibrium. Evidence of a genetic bottleneck was found in the four populations evaluated herein. The genetic structure of the population was investigated using the analogues F_ST and D_EST. The most genetic variability occurred within populations. Differentiation among populations ranged from low to moderate levels. No significant correlation was found between geographic and genetic distances. Implications of these findings for management and conservation of O. mossambicus stocks are discussed.     

Screening and identification of a microsatellite marker associated with sex in Wami tilapia, <Emphasis Type="Italic">Oreochromis urolepis hornorum</Emphasis>

In this study, primer pairs of 15 microsatellite markers associated with sex determination of tilapia were selected and amplified in Wami tilapia, Oreochromis urolepis hornorum. While one marker, UNH168, on linkage group 3 (LG3) was associated (P < 0.001) with the phenotypic sex in the experimental population, nine genotypes were detected in both sexes. Only 99-bp allele was detected in the female samples, while 141, 149 and 157-bp alleles were present in both male and female samples. UNH168 was localized by fluorescence in situ hybridization (FISH) on the long arm of the largest tilapia chromosome pair (chromosome 1, equivalent to LG3). This sex-linked microsatellite marker could potentially be used for marker-assisted selection in tilapia breeding programmes to produce monosex male tilapia.     

Behavioral plasticity in rainbow trout (Oncorhynchus mykiss) with divergent coping styles: when doves become hawks

Consistent and heritable individual differences in reaction to challenges, often referred to as stress coping styles, have been extensively documented in vertebrates. In fish, selection for divergent post-stress plasma cortisol levels in rainbow trout (Oncorhynchus mykiss) has yielded a low (LR) and a high responsive (HR) strain. A suite of behavioural traits is associated with this physiological difference, with LR (proactive) fish feeding more rapidly after transfer to a new environment and being socially dominant over HR (reactive) fish. Following transport from the UK to Norway, a switch in behavioural profile occurred in trout from the 3rd generation; HR fish regained feeding sooner than LR fish in a novel environment and became dominant in size-matched HR–LR pairs. One year after transport, HR fish still fed sooner, but no difference in social dominance was found. Among offspring of transported fish, no differences in feeding were observed, but as in pre-transported 3rd generation fish, HR fish lost fights for social dominance against size-matched LR opponents. Transported fish and their offspring retained their distinctive physiological profile throughout the study; HR fish showed consistently higher post-stress cortisol levels at all sampling points. Altered risk-taking and social dominance immediately after transport may be explained by the fact that HR fish lost more body mass during transport than did LR fish. These data demonstrate that some behavioural components of stress coping styles can be modified by experience, whereas behavioural plasticity is limited by genetic effects determining social position early in life story.     

Regulation of in vitro PWM-induced IgG secretion in humans

PWM-induced differentiation of human PBMC into immunoglobulin (Ig) secreting cells is a widely used model of in vivo IgG secretion. We examined the relationship between the amount of IgG secreted in PBMC cultures obtained from individuals who consistently secrete either very high (HR) or very low amounts (LR) of IgG and various in vitro immune function assays. The PBMC obtained from LR contained a higher ratio of cells expressing the T-suppressor/inducer to T-helper/inducer phenotype. The autologous mixed lymphocyte response was lower and the amount of in vivo radiation sensitive suppression was higher in the LR than in the HR. LR E- cells secreted less IgG than the HR E- cells when both were mixed with heterologous HR E+ cells. Monocyte depletion reduced IgG secretion in both LR and HR. These results suggest that each of the subsets Ts, Th (Thi, Tsi), and B lymphocytes are involved in the regulation of PWM-induced Ig secretion and that the functions of each of these subsets differ in HR and LR individuals.     

Effects of serum on phagocytic activity and proteomic analysis of tilapia (Oreochromis mossambicus) serum after acute osmotic stress

The objective of the study was to analyze the effect of serum from freshwater (FW) exposed tilapia or from 25 ppt seawater (SW) exposed tilapia on the ability to mediate the phagocytic activity of tilapia phagocytes. To analyze the phagocytic activity, head kidney (HK) and spleen leukocytes were tested in 300 or 500 mOsm medium using three different treatment groups (a) control, (b) addition of 25% serum from freshwater (FW) exposed tilapia, and (c) addition of 25% of serum from 25 ppt seawater (SW) exposed tilapia. HK leukocytes cultured in 300 and 500 mOsm media for 4 h showed an increase of phagocytic ability in the control group as compared to the addition of serum from either FW or SW exposed tilapia. HK leukocytes exposed to 500 mOsm medium showed a higher phagocytic ability than those leukocytes exposed to 300 mOsm medium in each corresponding group. Concurrently, spleen leukocytes in the control group showed a higher phagocytic ability than those leukocytes with the addition of serum from FW or SW exposed tilapia. As compared to spleen leukocytes cultured in 300 mOsm medium, leukocytes cultured in 500 mOsm medium showed an increase of phagocytic ability within their respective group. To further investigate the observed phenomenon, 2D-gel electrophoresis was performed for analyzing the differentially expressed proteins in serum that was thought to influence the phagocytic ability. Up-regulated serum proteins in SW exposed tilapia contained complement C3 protein, NADH dehydrogenase (Ubiquinone) Fe–S protein 3, Mg²⁺-dependent neutral sphingomyelinase, Semaphorins, and Caspase 3. Taken together these results suggest that addition of serum decreased the phagocytic activity in HK and spleen leukocytes in vitro, furthermore, induced proteins semaphorin, complement C3, Mg²⁺-dependent neutral sphingomyelinase, and Caspase 3 are up-regulated in the serum, which might have decreased the phagocytic activity upon exposure to hyperosmotic solutions.     

Dendritic morphology of hippocampal and amygdalar neurons in adolescent mice is resilient to genetic differences in stress reactivity

Many studies have shown that chronic stress or corticosterone over-exposure in rodents leads to extensive dendritic remodeling, particularly of principal neurons in the CA3 hippocampal area and the basolateral amygdala. We here investigated to what extent genetic predisposition of mice to high versus low stress reactivity, achieved through selective breeding of CD-1 mice, is also associated with structural plasticity in Golgi-stained neurons. Earlier, it was shown that the highly stress reactive (HR) compared to the intermediate (IR) and low (LR) stress reactive mice line presents a phenotype, with respect to neuroendocrine parameters, sleep architecture, emotional behavior and cognition, that recapitulates some of the features observed in patients suffering from major depression. In late adolescent males of the HR, IR, and LR mouse lines, we observed no significant differences in total dendritic length, number of branch points and branch tips, summated tip order, number of primary dendrites or dendritic complexity of either CA3 pyramidal neurons (apical as well as basal dendrites) or principal neurons in the basolateral amygdala. Apical dendrites of CA1 pyramidal neurons were also unaffected by the differences in stress reactivity of the animals; marginally higher length and complexity of the basal dendrites were found in LR compared to IR but not HR mice. In the same CA1 pyramidal neurons, spine density of distal apical tertiary dendrites was significantly higher in LR compared to IR or HR animals. We tentatively conclude that the dendritic complexity of principal hippocampal and amygdala neurons is remarkably stable in the light of a genetic predisposition to high versus low stress reactivity, while spine density seems more plastic. The latter possibly contributes to the behavioral phenotype of LR versus HR animals.     

Fine‐mapping of a locus on linkage group 23 for sex determination in Nile tilapia (Oreochromis niloticus)

Genetic markers in tilapia species associated with loci affecting sex determination (SD), sex‐specific mortality or both were mapped to linkage groups (LG) 1, 2, 3, 6 and 23. The objective of this study was to use these markers to fine‐map the locus with the greatest effect on SD in Oreochromis niloticus. Our parental stock, full‐sibs of Nile tilapia (Swansea origin), were divided into three groups: (i) untreated, (ii) feminized by diethylstilbestrol and (iii) masculinized by 17α‐methyltestosterone. We analysed the first group for association of microsatellite markers representing these five LGs. The strongest association with gender was found on LG23 for marker UNH898 (χ²; P = 8.6 × 10^?5). Allele 276 was found almost exclusively in males, and we hypothesized that this allele is a male‐associated allele (MAA). Sex‐reversed individuals were used for mating experiments with and without the segregating MAA. Mating of individuals lacking the MAA resulted in all‐female progeny. Mating of two heterozygotes for MAA gave rise to 81 males and 30 females. Analysis of association between gender and genotypes identified the MAA in 98.6% of males as opposed to 8.0% of females (χ²; P = 2.5 × 10^?18). Eight markers that flank UNH898 were genotyped to map the locus on LG23 within a confidence interval of 16–21 cM. Mating of homozygous individuals for MAA is underway for production of all‐male populations.     

Heritable, diet-induced hyperlipidemia in California mice (Peromyscus californicus) is due to increased hepatic secretion of very low density lipoprotein triacylglycerol

California mice (Peromyscus californicus) develop type II diabetes mellitus when fed a high-fat diet. We undertook the current studies to determine whether hyperlipidemia precedes the development of insulin resistance and to establish breeding colonies of hyperlipidemic and normolipidemic mice. For 6 wk, mice (n = 24) received a diet containing 25.8% of energy from fat. Mice representing the upper and lower quartiles of serum triacylglycerol (TAG) response (mean, >1000 mg/dl versus <300 mg/dl, respectively; 6 mice per group) were designated as high (HR) and low (LR) responders, respectively, and were used for further study. After 12 wk of consuming the high-fat diet, HR mice remained hypertriglyceridemic and developed hyperinsulinemia (5.1 +/- 1.3 ng/ml), hypercholesterolemia (309.3 +/- 31.0 mg/dl), and hyperglycemia (205.9 +/- 30.3 mg/dl) compared with LR mice. HR mice were not hyperphagic or obese. Offspring of HR x HR mice had elevated serum TAG concentrations (mean, 1752.2 +/- 209.7 mg/dl), hypercholesterolemia, hyperinsulinemia, and mild hyperglycemia by 5.5 mo of age. Mating HR male and LR female mice produced HR, intermediate, and LR progeny. HR mice had elevated serum concentrations of cholesterol, and plasma concentrations of high density lipoprotein cholesterol, and the very low density lipoprotein TAG compared with LR mice. Lipoprotein lipase and hepatic lipase activities did not differ between HR and LR mice. Studies of in vivo hepatic TAG production indicated that the hyperlipidemia of HR mice is a consequence of TAG hypersecretion.     

Effects of maternal stress coping style on offspring characteristics in rainbow trout (Oncorhynchus mykiss)

Maternal size, age, and allostatic load influence offspring size, development, and survival. Some of these effects have been attributed to the release of glucocorticoids, and individual variation in these stress hormones is related to a number of traits. Correlated traits are often clustered and used to define the proactive and reactive stress coping styles. Although stress coping styles have been identified in a number of animal groups, little is known about the coupling between stress coping style and offspring characteristics. In the present study, plasma cortisol levels in ovulated mothers and cortisol levels in non-fertilized eggs from two rainbow trout (Oncorhynchus mykiss) strains selected for high (HR) and low (LR) post-stress plasma cortisol levels were compared. Offspring characteristics such as egg size, larval growth, and energy reserves also were compared between the two strains. Maternal plasma and egg cortisol levels were correlated, but no difference between the HR and LR strains was detected in either parameter. LR females produced larger eggs, and larvae with larger yolk sacs compared to HR females, however no differences in larval body size (excluding the yolk) was detected between strains. Considering that the HR and LR strains have a number of correlated behavioral and physiological traits that resemble the reactive and proactive stress coping styles, respectively, the results suggest that proactive mothers invest more energy into their offspring, producing larvae with larger energy reserves. It is possible that larger energy reserves in proactive larvae support the energy requirement for establishing and defending territory in salmonid fish. Furthermore, in the present study we found a positive relationship between mother plasma cortisol and egg cortisol; however neither mother plasma cortisol nor egg cortisol differed between strains. These results indicate that cortisol endowment from the mother to the offspring plays a minor role in the transfer of the behavioral and physiological traits which separates these strains.     

Identification of Quantitative Trait Loci Influencing Wood Specific Gravity in an Outbred Pedigree of Loblolly Pine

We report the identification of quantitative trait loci (QTL) influencing wood specific gravity (WSG) in an outbred pedigree of loblolly pine (Pinus taeda L.). QTL mapping in an outcrossing species is complicated by the presence of multiple alleles (>2) at QTL and marker loci. Multiple alleles at QTL allow the examination of interaction among alleles at QTL (deviation from additive gene action). Restriction fragment length polymorphism (RFLP) marker genotypes and wood specific gravity phenotypes were determined for 177 progeny. Two RFLP linkage maps were constructed, representing maternal and paternal parent gamete segregations as inferred from diploid progeny RFLP genotypes. RFLP loci segregating for multiple alleles were vital for aligning the two maps. Each RFLP locus was assayed for cosegregation with WSG QTL using analysis of variance (ANOVA). Five regions of the genome contained one or more RFLP loci showing differences in mean WSG at or below the P = 0.05 level for progeny as grouped by RFLP genotype. One region contained a marker locus (S6a) whose QTL-associated effects were highly significant (P > 0.0002). Marker S6a segregated for multiple alleles, a prerequisite for determining the number of alleles segregating at the linked QTL and analyzing the interactions among QTL alleles. The QTL associated with marker S6a appeared to be segregating for multiple alleles which interacted with each other and with environments. No evidence for digenic epistasis was found among the five QTL.     

Generation of selective microenvironment at the cell periphery through bulk-phase hydrophilic polymers

In order to understand the previously demonstrated effect of poly(ethylene glycol) on the stimulation of lymphocyte responses to syngeneic tumor cells (Ben-Sasson, S.A. and Henkart, P.A. (1977) J. Immunol. 119, 227–231), we have investigated the effects of addition of poly(ethylene glycol) to the medium in a number of cellular systems. The binding of trimeric IgG to tumor-lymphocyte Fc receptors was greatly enhanced by poly(ethylene glycol); a substantial increase in binding of trimeric IgG to non-Fc-receptor-bearing tumor cells was also observed. Similarly, the binding of labeled bovine serum albumin to lymphocyte surfaces was increased by poly(ethylene glycol), implying that nonspecific binding of proteins to cells was generally enhanced. The dose-response curve of concanavalin A mitogenesis was shifted to the right, as would be expected from a local increase in concanavalin A concentration. Antibody binding to erythrocytes as detected by complement lysis was similarly increased. It was found that in aqueous two-phase mixtures created by poly(ethylene glycol) and dextran, erythrocytes partition into the dextran phase through exclusion into dextran-rich microdroplets. It is proposed that addition of poly(ethylene glycol) to cell culture media creates a similar separate phase around the cell surface in which the local concentration of proteins is greater than that in the bulk medium. This concept explains many of the diverse effects of addition of poly(ethylene glycol) to the medium. It also can partially explain the requirement for serum to observe the poly(ethylene glycol) effect on the lymphocyte response to syngeneic tumor cells.     

An AFLP and RFLP linkage map and quantitative trait locus (QTL) analysis of growth traits in Salix

A genetic linkage map of Salix (2n = 38), composed of 325 AFLP and 38 RFLP markers has been constructed. The map was based on a population ( n = 87) derived from a cross between the male hybrid clone "Bj?rn" ( Salix viminalis x Salix schwerinii) and the female clone "78183" ( S. viminalis). Three hundred fifty seven AFLPs corresponding to DNA polymorphisms heterozygous in one parent and null in the other were scored. A total of 87 RFLP probes, most (83) derived from the Populus genome, yielded 39 and 11 polymorphic loci segregating in a 1:1 and 1:2:1 ratio respectively. Two maps, one for each parent, were constructed according to the "two-way pseudo-testcross" mapping strategy. The S. viminalis x S. schwerinii map (2,404 cM) included 217 markers and formed 26 major linkage groups while S. viminalis (1,844 cM) consisted of 146 markers placed on 18 major groups. In addition, eight and 14 additional minor linkage groups composed of less than four markers (doubles and triplets) were obtained in the S. viminalis x S. schwerinii and the S. viminalis maps, respectively. Both maps provided 70-80% genome coverage with an average density of markers of 14 cM. To investigate possible homologies between the parental maps, 20 AFLPs and 11 RFLPs segregating in 3:1 or 1:2:1 ratios were included in the linkage analysis. Eight linkage groups homologous between the two maps were detected. The present genetic map was used to identify quantitative trait loci (QTLs) affecting growth-related traits. Eleven QTLs were identified; seven QTLs for height growth, one QTL for stem diameter, one QTL for the height: diameter ratio, one QTL for the number of vegetative buds during flowering time and one QTL for the number of shoots. The estimated magnitude of the QTL effect ranged from 14 to 22% of the total phenotypic variance. One QTL associated with height growth and one affecting the height: diameter ratio were overlapping in the same marker interval with the QTL affecting stem diameter. QTL stability over years was estimated for traits measured in multiple years. Generally, QTLs were only significant in a single year although two QTLs for height growth were close to reaching the significance level in 2 consecutive years.     

Genomic regions determining resistance to leaf stripe (Pyrenophora graminea) in barley.

Leaf stripe is a seed-borne disease of barley (Hordeum vulgare) caused by Pyrenophora graminea. Little is known about the genetics of resistance to this pathogen. In the present work, QTL analysis was applied on two recombinant inbred line (RIL) populations derived from two- and six-rowed barley genotypes with different levels of partial resistance to barley leaf stripe. Quantitative trait loci for partial resistance were identified using the composite interval mapping (CIM) method of PLABQTL software, using the putative QTL markers as cofactors. In the L94 x 'Vada' mapping population, one QTL for resistance was detected on chromosome 2H; the same location as the leaf-stripe resistance gene Rdg1 mapped earlier in 'Alf', where it confers complete resistance to the pathogen. An additional minor-effect QTL was identified by further analyses in this segregating population on chromosome 7H. In L94 x C123, two QTLs for resistance were mapped, one each on chromosomes 7H and 2H.     

Ultrastructure of experimentally produced subcutaneous haematomas in the rabbit.

Haematomas were produced in rabbits by subcutaneous injection of autologous blood. Clotting and marked lysis of erythrocytes was noted in these haematomas, but there was no evidence of fragmentation of erythrocytes prior to or after ingestion by macrophages as has been reported in other sites such as the peritoneal cavity and the joint cavity. The phagocytosis of intact erythrocytes, lysed erythrocytes and haemoglobin led to the formation of three main types of lysosomal bodies; (1) myelinosomes containing whorled osmiophilic membranes, (2) siderosomes containing haemosiderin, and (3) myelinosiderosomes containing a mixture of osmiophilic membranes and haemosiderin.     

Whole blood leukocytes isolation with microfabricated filter for cell analysis

The flow cytometric analysis of leukocytes in whole blood usually requires isolation of leukocytes from other components of whole blood. Density gradient centrifugation and red blood cell lysis are the most commonly used methods to separate leukocytes but come with significant limitations. We report the results of the evaluation of a microfabricated filtration device for blood preparation that separates erythrocytes from leukocytes based on their size and mechanical properties. The microfabricated filter evaluated here requires a rapid and simple procedure and results in high leukocytes recovery without introducing bias among the leukocyte subpopulations. The filter removes erythrocytes, platelets, plasma proteins, and unbound staining reagent. This gentle filtration process produces very clean stained leukocytes for cytometric analysis without any apparent damage to leukocytes.