Independent Progeny Research Articles

A Malaria Parasite Cross Reveals Genetic Determinants of Plasmodium falciparum Growth in Different Culture Media.

What genes determine in vitro growth and nutrient utilization in asexual blood-stage malaria parasites? Competition experiments between NF54, clone 3D7, a lab-adapted African parasite, and a recently isolated Asian parasite (NHP4026) reveal contrasting outcomes in different media: 3D7 outcompetes NHP4026 in media containing human serum, while NHP4026 outcompetes 3D7 in media containing AlbuMAX, a commercial lipid-rich bovine serum formulation. To determine the basis for this polymorphism, we conducted parasite genetic crosses using humanized mice and compared genome-wide allele frequency changes in three independent progeny populations cultured in media containing human serum or AlbuMAX. This bulk segregant analysis detected three quantitative trait loci (QTL) regions [on chromosome (chr) 2 containing aspartate transaminase AST; chr 13 containing EBA-140; and chr 14 containing cysteine protease ATG4] linked with differential growth in serum or AlbuMAX in each of the three independent progeny pools. Selection driving differential growth was strong (s = 0.10 – 0.23 per 48-hour lifecycle). We conducted validation experiments for the strongest QTL on chr 13: competition experiments between ΔEBA-140 and 3D7 wildtype parasites showed fitness reversals in the two medium types as seen in the parental parasites, validating this locus as the causative gene. These results (i) demonstrate the effectiveness of bulk segregant analysis for dissecting fitness traits in P. falciparum genetic crosses, and (ii) reveal intimate links between red blood cell invasion and nutrient composition of growth media. Use of parasite crosses combined with bulk segregant analysis will allow systematic dissection of key nutrient acquisition/metabolism and red blood cell invasion pathways in P. falciparum.

The messy merger of a large satellite and a Milky Way-like galaxy

Aims. About 10 billion years ago the Milky Way merged with a massive satellite, Gaia-Enceladus. To gain insight into the properties of its debris we analyse in detail a suite of simulations that includes an experiment that produces a good match to the kinematics of nearby halo stars inferred from Gaia data. Methods. We compare the kinematic distributions of stellar particles in the simulations and study the distribution of debris in orbital angular momentum, eccentricity, and energy, and its relation to the mass loss history of the simulated satellite. Results. We confirm that Gaia-Enceladus probably fell in on a retrograde, 30° inclination orbit. We find that while 75% of the debris in our preferred simulation has high eccentricity (> 0.8), roughly 9% has eccentricity lower than 0.6. Star particles lost early have large retrograde motions, and a subset of these have low eccentricity. Such stars would be expected to have lower metallicities as they stem from the outskirts of the satellite, and hence naively they could be confused with debris associated with a separate system. These considerations seem to apply to some of the stars from the postulated Sequoia galaxy. Conclusions. When a massive disc galaxy undergoes a merger event, it leaves behind debris with a complex phase-space structure, a wide range of orbital properties, and a range of chemical abundances. Observationally, this results in substructures with very different properties, which can be misinterpreted as implying independent progeny. Detailed chemical abundances of large samples of stars and tailored hydrodynamical simulations are critical to resolving such conundrums.

Genetic analysis of fiber-dimension traits and combined selection for simultaneous improvement of growth and stiffness in lodgepole pine (Pinus contorta)

Quantitative genetic variation of fiber-dimension traits and their relationship with diameter at breast height (DBH) and solid-wood traits (i.e., density and modulus of elasticity (MOE)) was investigated in lodgepole pine (Pinus contorta Dougl. ex Loud. var. latifolia Engelm.). A total of 823 increment cores were sampled from 207 half-sib families at two independent progeny trials, aged 34–35 years, located in northern Sweden. High-resolution pith-to-bark profiles were obtained for radial fiber width (RFW), tangential fiber width (TFW), fiber wall thickness (FWT), and fiber coarseness (FC) using SilviScan. Heritabilities ranged from 0.29 to 0.74, and inheritance increased with cambial maturity. Estimated age–age genetic correlations indicate that early selection between ages 5 and 8 years is highly efficient. Our results indicate that selection for a 1% increase in DBH or MOE incurs a negligible effect on fiber-dimension traits and maximum genetic gains are reached when DBH and MOE are considered jointly. Moreover, simultaneous improvement of growth and stiffness is achievable when a selection index with 7 to 10 economical weights for MOE relative to 1 for DBH is incorporated. However, the unfavorable relationship between solid-wood traits and pulp and paper related traits suggests that breeding strategies must be implemented to improve wood quality of lodgepole pine for multiple uses.

Genetic analysis of lodgepole pine (Pinus contorta) solid-wood quality traits

Potential improvement of lodgepole pine (Pinus contorta Dougl. ex Loud. var. latifolia Engelm.) solid-wood properties was examined by estimating age trends of inheritance, age–age genetic correlations, and the efficiency of early selection using 823 increment cores sampled from 207 half-sib families at two independent progeny trials, aged 34–35 years, located in northern Sweden. High-resolution radial variation of annual ring width, wood density, microfibril angle (MFA), and modulus of elasticity (clearwood stiffness; MOES) was measured using SilviScan. The dynamic stiffness (MOEtof) of standing trees was also obtained using Hitman ST300. Heritabilities ranged from 0.10 to 0.64 for growth and earlywood, transition-wood, and latewood proportions, from 0.29 to 0.77 for density traits, and from 0.13 to 0.33 for MFA and stiffness traits. Genetic correlations between early age and the reference age (26 years) suggested that early selection is efficient at age 4 years for MFA and between ages 5 to 8 years for density and MOES. Unfavorable diameter–stiffness genetic correlations and correlated responses indicate that breeding for a 1% increase in diameter would confer 5.5% and 2.3% decreases in lodgepole pine MOES and MOEtof, respectively. Index selection with appropriate economical weights for growth and wood stiffness is highly recommended for selective breeding.

First intergeneric hybrids within the tribe Anthemideae Cass. III. Chrysanthemum indicum L. Des Moul. × Opisthopappus taihangensis (Ling) Shih

Ovule rescue was used to obtain the intergeneric hybrid Chrysanthemum indicum L. Des Moul. x Opisthopappus taihangensis (Ling) Shih. The hybridity of the seven independent progeny was confirmed by morphology, chromosome counting and GISH analysis. Plant height, crown width, leaf length, leaf width, epidermal hair height and epidermal hair length of the hybrid were all intermediate between those of the parents. However, their petiole length and epidermal hair density exceeded those of either parent. GISH analysis of the hybrid was able to distinguish between the parental genomes.

Genetics of virulence in Ascochyta rabiei

In order to critically test the hypothesis that virulence variation in the Ascochyta rabiei/chickpea pathosystem is a discrete character under simple genetic control, a genetic cross was made between a highly virulent isolate of A. rabiei from Syria and a less virulent isolate from the USA. Two independent virulence assays conducted by inoculating susceptible and resistant chickpea cultivars under controlled conditions with 77 independent progeny isolates from this cross revealed a continuous distribution of disease phenotypes. Bimodality, as would be predicted for the segregation of virulence under simple genetic control, was not supported by statistical tests of the progeny phenotype distribution. anova revealed highly significant pathogen‐genotype × host‐genotype interactions demonstrating the segregation of genes controlling specialization on the two cultivars tested. These interactions could be localized to two isolates that changed virulence rank on the cultivars. It was concluded that variation in virulence to these two cultivars is under quantitative genetic control. If this conclusion applies to other cultivars, it can be speculated that the discrete categories of virulence variation identified in previous studies were probably the result of incomplete sampling of host resistance or pathogen virulence variation and/or of selection for increased virulence in contemporary A. rabiei populations.

Genetic exchange in Trypanosoma brucei: Evidence for mating prior to metacyclic stage development

It is well established that genetic exchange occurs between Trypanosoma brucei parasites when two stocks are used to infect tsetse flies under laboratory conditions and a number of such crosses have been undertaken [1,2]. Both cross and self-fertilisation can take place [3,4], with the products of mating being the equivalent of F1 progeny in a Mendelian system [2,3]. Recently, analysis of a large collection of independent progeny using a series of polymorphic micro and minisatellite markers, has formally demonstrated that the allelic segregation at loci on each of the 11-megabase chromosomes conforms to ratios predicted for a classical diploid genetic system involving meiosis as well as independent assortment of markers on different chromosomes [5]. Further extensive analysis of these F1 progeny, using a large panel of micro and minisatellite markers, has led to the construction of a genetic map of one parasite stock [6].

Allelic segregation and independent assortment in T. brucei crosses: Proof that the genetic system is Mendelian and involves meiosis

The genetic system on Trypanosoma brucei has been analysed by generating large numbers of independent progeny clones from two crosses, one between two cloned isolates of Trypanosoma brucei brucei and one between cloned isolates of T. b. brucei and Trypanosoma brucei gambiense, Type 2. Micro and minisatellite markers (located on each of the 11 megabase housekeeping chromosomes) were identified, that are heterozygous in one or more of the parental strains and the segregation of alleles at each locus was then determined in each of the progeny clones. The results unequivocally show that alleles segregate in the predicted ratios and that alleles at loci on different chromosomes segregate independently. These data provide statistically robust proof that the genetic system is Mendelian and that meiosis occurs. Segregation distortion is observed with the minisatellite locus located on chromosome I of T. b. gambiense Type 2 and neighboring markers, but analysis of markers further along this chromosome did not show distortion leading to the conclusion that this is due to selection acting on one part of this chromosome. The results obtained are discussed in relation to previously proposed models of mating and support the occurrence of meiosis to form haploid gametes that then fuse to form the diploid progeny in a single round of mating.

Origin and fate of a one-male savanna babbon group formed by fissioning

An adult male chacma baboon,Papio hamadryas ursinus, emigrated from his habituated natal group at age 100 months and joined a neighboring multimale-multifemale group. Subsequently this male, together with eight adult females, all with infants <1 year old, formed a one-male group (OMU). The male was the only adult male in this group for over 200 days. Circumstantial evidence shows that shared parentage of the infants with this male determined which females joined the new group. In addition, 10 juveniles younger than 4.5 years joined the new group. Based upon persistent close physical association, especially at sleeping sites, these juveniles were presumed to be the older independent progeny of the eight adult females. Thus, the founding elements of the new group were the adult male and partial matrilines—mothers, their infants, and their independent juvenile offspring. The OMU status of the group ended with the sequential intrusion of three males known to be low ranking in other groups. The first of these intruders was successful in joining the OMU in spite of the OMU male's earlier successes in rebuffing this and other potentially high-ranking males. Six additional males later entered the fission group during a 6-month interval. Two of them outranked the original male. All additional males present in the last 100 days of observations came from the same group, which was not the group of origin of the females.

Estimation of self-fertilization rate and allelic frequencies in diploidized tetraploids

A methodology for the estimation of the self-fertilization rate and allelic frequencies in diploidized tetraploids is developed. Data from both a population survey and independent progeny arrays from homozygous parents are generally necessary for maximum likelihood estimates. If progeny arrays from a random population sample are examined, then data from the resulting six phenotypic categories may be used. An example using the first approach is given using data from the Menzies’ wallflower, an endangered species in California.

A Test for Genetic Exchange in Mixed Infections ofLeishmania majorin the Sand FlyPhlebotomus papatasi

We tested if genetic exchange was observable between two strains of Leishmania major (Trypanosomatidae) during mixed infection of the sand fly Phlebotomus papatasi. Previous studies suggested that genetic exchange may occur in natural populations of Leishmania at a low frequency, but experimental crosses examining small numbers of progeny (less than 60) did not reveal hybrid parasites. Accordingly, a strategy was devised to increase the number of progeny that could be screened by 100-fold. Clonal derivatives from two strains that were infective to flies and contained numerous restriction fragment length polymorphisms were characterized and selected for resistance to methotrexate or tunicamycin by gene amplification. A successfully mixed infection of P. papatasi was obtained, and a method was developed for directly plating promastigotes from the gut contents of infected flies onto selective media. Twenty-five hundred independent progeny were scored for the presence of both drug resistance markers. No hybrid parasites were observed, indicating that the frequency of genetic exchange in this cross must be less than 4 x 10(-4). The lines and methods established in this work may prove useful in future studies of the mechanism and frequency of gene exchange in Leishmania.

Mutation rate of the hepadnavirus genome

An essential factor for charting the evolution of hepadnaviruses is an estimation of the mutation rate of the virus genome during replication in the host. In order to determine the mutation rate of the hepadnavirus genome under defined experimental conditions, we transfected 10 neonatal woodchucks with an infectious molecular clone of woodchuck hepatitis virus (WHV). By 4 months post-transfection, all 10 animals showed serological evidence for WHV infection. Subsequently, 1 animal became chronically infected and was used for further study. At 16 months post-transfection WHV DNA from serum virions was cloned and the nucleotide sequence of three independent progeny genomes compared directly with that of the input recombinant DNA. Although the consensus nucleotide sequence remained unchanged, we found three differences in individual progeny genomes when compared to the parental genome sequence. Thus, we estimate the mutation rate of the WHV genome to be ⩽ 2 x 10 −4 base substitutions/site/year. This figure is one to two orders of magnitude lower than the mutation rates previously calculated for the positive- and negative-strand RNA viruses, but is similar to the mutation rate of the gag gene which is the most slowly evolving gene of retroviruses. Therefore, we find that the hepadnavirus genome is relatively stable during replication in host tissues when compared to other viruses that lack polymerase-associated proofreading functions.

Individual BFU-E in polycythemia vera produce both erythropoietin dependent and independent progeny

Erythropoietic progenitors from peripheral blood of normal individuals or patients with polycythemia vera (PV) were cultured in methylcellulose medium containing 2.5 U/ml of erythropoietin (Ep). After 7–9 days, colonies considered to be early stage large bursts were individually removed, resuspended in a small volume of fresh methylcellulose medium, and then divided between 2 dishes. To one of these secondary cultures, sufficient Ep was added to bring the concentration of Ep up to approximately 3 U/ml. To the other was added an equal volume of medium but no Ep. The final concentration of Ep in these cultures was determined to be less than 0.01 U/ml. Nine days later, both types of secondary cultures were scored for the presence of colonies containing 8 or more hemoglobinized erythroblasts. Of 90 primary colonies from 3 normal individuals assessed in this way, 59 gave secondary erythroid colonies in the high Ep cultures, while none gave secondary erythroid colonies in the low Ep cultures. Additional control experiments in which primary colonies from normal individuals were divided into duplicate high Ep cultures showed that on average, the procedure used divided primary colonies equally. Of 109 primary colonies from 5 PV patients that yielded secondary erythroid colonies in the high Ep cultures, 21 yielded no secondary erythroid colonies in the low Ep cultures. The other 88 yielded erythroid colonies in both, but the secondary colonies in the low Ep cultures were consistently smaller in size and significantly fewer in number. Similar results were obtained when primary colonies were generated in cultures to which no Ep was added. These findings indicate that primitive BFU-E in patients with PV can be subdivided into 2 populations: a minor population restricted to the production of erythroid colony-forming cells (Ep- dependent progenitors) that require Ep for their detection, and a major population that is not restricted in this way. In addition, these experiments show that most of the primitive BFU-E that generate Ep- independent progenitors also produce significant numbers of cells that are Ep-dependent.

Marker rescue of the adsorption properties of bacteriophage T4 by bacteriophage T2.

Rescue of adsorption properties from UV-irradiated T4 by T2 as a helper phage, revealed progeny phage with intermediate properties. Fourteen independent progeny phages, plating onE. coli B/2, were plated on several indicator strains and their adsorption properties were also studied with specific T4 antibodies. Two of these, plating onE. coli KS/4, were not inactivated by the T4 antiserum, and were T2h without apparent T4 properties. The other 12 progeny phages did not plate on KS/4, and were inactivated, but at a slower rate than the parental T4. Their mean efficiency of plating onE. coli B/2 (0.83) was significantly lower than that of the parental T4. The efficiency of plating was positively correlated with the velocity of inactivation by T4 antiserum. The observations were explained by assuming that the progeny phages were recombinants of T4 and T2 loci for adsorption sites.