Clonal diversity and structure of the invasive aquatic plant Eichhornia crassipes in China

The information on diversity and spatial distribution of clones of an invasive clonal plant is crucial for the understanding of its clonal structure and invasive history. In this paper, random amplified polymorphic DNA (RAPD) markers were used to explore the clonal diversity and clonal structure of Eichhornia crassipes (Mart.) Solms in natural populations, and their possible effects on the plant success as an invader are also discussed. Five populations covering the entire distribution area in China were studied, sampling 43 individuals per population at an interval of 1 m in a sampling plot. Twelve RAPD primers produced 69 reproducible bands, with 22 being polymorphic. Only five RAPD phenotypes (clones) were detected in these five populations, but each population consisted of at least three clones, contrary to the traditional expectations that E. crassipes populations should be monoclonal. The diversity of clones within populations is thought to be mainly resulted from multiple introductions by humans. The evenness of distribution of clones varied slightly and most clones were widespread, suggesting clonal growth is the predominant mode of regeneration in all the populations. A single clone dominated each population and this clone might be the first one introduced into China or the genotype with a higher phenotypic plasticity, which could survive and reproduce via clonal growth in various habitats. The clones in each population were highly intermixed, especially in river populations, suggesting this species has a guerilla clonal structure which can be facilitated by water current.     

Spatial genetic structure and clonal diversity in an alpine population of Salix herbacea (Salicaceae)

BACKGROUND AND AIMS: Many alpine plant species combine clonal and sexual reproduction to minimize the risks of flowering and seed production in high mountain regions. The spatial genetic structure and diversity of these alpine species is strongly affected by different clonal strategies (phalanx or guerrilla) and the proportion of generative and vegetative reproduction. METHODS: The clonal structure of the alpine plant species Salix herbacea was investigated in a 3 x 3 m plot of an alpine meadow using microsatellite (simple sequence repeat; SSR) analysis. The data obtained were compared with the results of a random amplified polymorphic DNA (RAPD) analysis. KEY RESULTS: SSR analysis, based on three loci and 16 alleles, revealed 24 different genotypes and a proportion of distinguishable genotypes of 0.18. Six SSR clones were found consisting of at least five samples, 17 clones consisting of more than two samples and seven single genotypes. Mean clone size comprising at least five samples was 0.96 m(2), and spatial autocorrelation analysis showed strong similarity of samples up to 130 cm. RAPD analysis revealed a higher level of clonal diversity but a comparable number of larger clones and a similar spatial structure. CONCLUSIONS: The spatial genetic structure as well as the occurrence of single genotypes revealed in this study suggests both clonal and sexual propagation and repeated seedling recruitment in established populations of S. herbacea and is thus suggestive of a relaxed phalanx strategy.     

Spatial models of foraging in clonal plant species

Ramets of some clonal plant species alter their internode lengths or their frequency of lateral branching in response to their immediate microenvironment. Such “plant foraging” responses are thought to allow clones to concentrate in favorable portions of their environment. Despite widespread interest among ecologists in plant foraging, few realistic models have been developed to examine conditions under which plant foraging responses are likely to provide clones with ecological benefit. In this paper, we develop spatially explicit, stochastic simulation models to examine consequences of both empirical and hypothetical plant foraging responses. We construct a hierarchical series of models in which we incorporate effects of resource heterogeneity on spacer lengths, angles of growth, and lateral branch production. We also vary the number, size, and arrangement of patches, and the presence or absence of ramet mortality. Simulations based on hypothetical data demonstrated the potential importance of shortening spacer lengths in favorable habitat. In these simulations, ramet crowding increased significantly, implying a potential cost to plant foraging responses whose magnitude is large enough to cause ramets to concentrate in favorable patches. Models calibrated with empirical data suggest that when clonal plants were able to concentrate in favorable habitat, this was usually caused by increased daughter ramet production in the favorable habitat. Variation in clonal growth angles had little impact on the ability of ramets or clones to locate favorable patches, but did increase the ability of clones to remain in favorable patches once found. Alterations in the number and size of patches strongly influenced the effectiveness of the foraging response. The spatial arrangement of patches also was important: clumped distributions of patches decreased the success with which plants located favorable patches, especially at the genet level and when the number of patches was low. Finally, when ramet mortality varied with patch quality, there was an increase in the percentage of ramets located in favorable patches; differential ramet mortality also lessened the impact of other effects, such as the decreased success of clones when patches are clumped. Overall, our models indicate that the effectiveness of plant foraging responses is variable and is likely to depend on a suite of environmental conditions.     

Clonal diversity of populations of Calamagrostis epigejos in relation to environmental stress and habitat heterogeneity

The clonal diversity of four populations of the vegetatively propagating grass Calamagrostis epigejos was studied with isozyme markers From each population, 100 individuals were sampled within a 10 × 10 m gnd In two populations of unpolluted sites 5 and 10 clones were distinguished, respectively Higher levels of clonal diversity were found m the populations at an abandoned sewage farm (59 clones) and around a copper smelter (92 clones) The level of clonal diversity observed suggests that sexual recruitment is important for the genetic structure in this species On the unpolluted, presumably more suitable habitats, the clonal diversity was lower than on the habitats polluted with heavy metals The results are discussed in relation to environmental stress and habitat heterogeneity, with the conclusion that in the unpolluted habitats intraspecific competition results in low clonal diversity On the other hand, stressful conditions decrease the importance of competition and facilitate the coexistence of a great number of clones     

Effects of fragmentation of clones compound over vegetative generations in the floating plant Pistia stratiotes

Background and AimsClonal plants dominate many plant communities, especially in aquatic systems, and clonality appears to promote invasiveness and to affect how diversity changes in response to disturbance and resource availability. Understanding how the special physiological and morphological properties of clonal growth lead to these ecological effects depends upon studying the long-term consequences of clonal growth properties across vegetative generations, but this has rarely been done. This study aimed to show how a key clonal property, physiological integration between connected ramets within clones, affects the response of clones to disturbance and resources in an aquatic, invasive, dominant species across multiple generations.MethodsSingle, parental ramets of the floating stoloniferous plant Pistia stratiotes were grown for 3 weeks, during which they produced two or three generations of offspring; connections between new ramets were cut or left intact. Individual offspring were then used as parents in a second 3-week iteration that crossed fragmentation with previous fragmentation in the first iteration. A third iteration yielded eight treatment combinations, zero to three rounds of fragmentation at different times in the past. The experiment was run once at a high and once at a low level of nutrients.ResultsIn each iteration, fragmentation increased biomass of the parental ramet, decreased biomass of the offspring and increased number of offspring. These effects persisted and compounded from one iteration to another, though more recent fragmentation had stronger effects, and were stronger at the low than at the high nutrient level. Fragmentation did not affect net accumulation of mass by groups after one iteration but increased it after two iterations at low nutrients, and after three iterations at both nutrient levels.ConclusionsBoth the positive and negative effects of fragmentation on clonal performance can compound and persist over time and can be stronger when resource levels are lower. Even when fragmentation has no short-term net effect on clonal performance, it can have a longer-term effect. In some cases, fragmentation may increase total accumulation of mass by a clone. The results provide the first demonstration of how physiological integration in clonal plants can affect fitness across generations and suggest that increased disturbance may promote invasion of introduced clonal species via effects on integration, perhaps especially at lower nutrient levels.     

Effects of Spatial Scale of Soil Heterogeneity on the Growth of a Clonal Plant Producing Both Spreading and Clumping Ramets

Soil nutrients are commonly heterogeneously distributed at different spatial scales. Although numerous studies have tested the effects of soil nutrient heterogeneity on growth of clonal plants producing either spreading ramets or clumping ramets, no study has examined the effects on the growth of clonal plants producing both spreading and clumping ramets and how spatial scale affects such effects. To test these effects, clones of Buchloe dactyloides, a stoloniferous clonal plant that produces both clumping and spreading ramets, were grown in six heterogeneous environments with different patch sizes and one homogeneous environment containing the same quantity of nutrients. Total biomass, total number of ramets, number of clumping ramets, number of spreading ramets, spacer length, or root:shoot ratio of the whole plants did not differ significantly among the seven treatments. However, at the patch level there were significant effects of patch size by nutrient level on biomass, number of ramets, number of spreading ramets, and number of clumping ramets, and these four variables were significantly larger in the nutrient-rich patches than in the nutrient-poor patches in the heterogeneous treatment with the largest patch size, but not in the other five heterogeneous treatments with smaller patch sizes. Neither nutrient level nor patch size significantly affected spacer length or root:shoot ratio. Based on our results, we propose that B. dactyloides can efficiently exploit nutrient-rich patches by a plastic response of clumping ramets and spreading ramets at larger spatial scales of soil heterogeneity but not at smaller ones.     

Use of random amplified polymorphic DNA (RAPD) markers in the discrimination and verification of genotypes in Eucalyptus

We carried out four separate studies using random amplified polymorphic DNA (RAPD) markers to analyse samples of Eucalyptus supplied by several different organisations. The objective was to examine the reproducibility of the RAPD technique and its ability to discriminate between individual genotypes for verification of clonal identities. We found that RAPD profiles that are unique to a genotype can be generated reliably and simply and that even closely related genotypes can be distinguished. In addition, in each of the four studies, we detected cases where the plant material studied had been mis-sampled or mis-labelled (i.e. the RAPD profiles were not consistent with the identification numbers): (1) ramets of a Eucalyptus grandis clone were found to be derived from 2 different clones; (2) ramets labelled as 2 different Eucalyptus hybrid clones were found to be the same clone, owing to a mis-planted clonal hedge; (3) samples supplied as a single progeny of a controlled E. nitens cross were derived from two crosses involving different pairs of parents; (4) mis-labelling was detected for ramets of 4 of a set of 10 clones of E. grandis and E. camaldulensis. For three of the four studies, the detection of genotype mis-identifications was unexpected, suggesting that labelling or sampling errors during the handling of plant material are a frequent occurrence, with potentially serious economic consequences.     

Clonal distribution,fertility and sex ratios of the moss Plagiomnium affine (Bland.) T.Kop. in forests of contrasting age

Abstract

Six populations of the clonal forest floor moss Plagiomnium affine from forests of different age were screened for genetic variation at 23 allozyme loci, of which nine were polymorphic. Samples consisting of two adjacent unconnected shoots were taken at regular intervals along one transect from each population. A total of 602 shoots was analysed. Almost 80% of the shoots were sterile (i.e. not expressing male or female gender). Sex remained unknown for only 10% of shoots after identification of genets based on electrophoretic data. We identified a mean number of 3.7 fertile clones per population. The mean length of clones along transects in each population ranged between 2 and 3 m. The size distribution within populations was bimodal, with a few dominant clones and a varying number of much smaller clones. The overall sex ratio was slightly female biased at the ramet level, but balanced at the genet level. Forest age was negatively correlated with percentage of sterile shoots and positively correlated with frequency of sporophytes. In both cases correlations were significant only if population 1, which was subject to extreme soil disturbance by badgers, was excluded. We conclude that the effective population size is larger, and the susceptibility to genetic drift is lower, in old forests.     

Clonal variation in response to adrenocorticotropin in cultured bovine adrenocortical cells: relationship to senescence

During cellular senescence, non-clonal cultures of bovine adrenocortical cells show a continuous decline in the rate of production of cyclic AMP (cAMP) stimulated by adrenocorticotropin (ACTH), without changes in the rate of forskolin- or prostaglandin E1-stimulated cAMP production. We investigated the possible mechanisms for loss of response to ACTH by examining the properties of clones of bovine adrenocortical cells. ACTH-stimulated cAMP production rates were measured in clones immediately after isolation, during long-term growth following isolation, and after subcloning. ACTH-stimulated rates were compared with cAMP production in response to forskolin, which acts directly on the catalytic subunit of adenylate cyclase. The results show that cloning is not necessarily associated with a loss of response to ACTH, but that clones with high ACTH response can give rise to subclones with low response. Clones of adrenocortical cells, at the same approximate population doubling level (PDL), showed ACTH response levels that ranged from 12 to 135 pmol cAMP/10(6) cells/min, whereas mass cultures at this PDL showed approximately 50 pmol/10(6) cells/min. Forskolin-stimulated cAMP production rates in clones varied only over the range of 59-119 pmol/10(6) cells/min and showed no correlation with the ACTH-stimulated rates. All clones were adrenocortical cells, as shown by mitogenic response to angiotensin II and cAMP-inducible 17 alpha-hydroxylase activity. The replicative potential of clones varied widely, and there was no apparent correlation between ACTH response levels and growth potential. The level of ACTH response in each clone was stable during proliferation through at least 25 PD beyond the stage at which the clone was isolated. When clones were subcloned, a clone with a high ACTH response level produced sister subclones that had ACTH response levels ranging from 3% of that of the parent clone to a level slightly greater than that of the parent clone. The growth potential of sister subclones varied widely, as for the parent clones, and there was no obvious correlation between growth potential and ACTH response. Two subclones were cloned; in sub-subclones, levels of ACTH response were again different from each other and also from the parent subclone; in one case, the level of ACTH response was approximately eight-fold higher than that of the parent subclone. These experiments show that clonal variation in the extent of expression of a differentiated property may occur in a normal differentiated cell in culture. The loss of ACTH response and the loss of proliferative potential appear to be independent stochastic events.     

Differential regulation of activation, clonal expansion, and antibody secretion in human B cells

Limiting dilution analysis, hemolytic plaque assay, and ELISA procedures were used to study the recruitment, clonal expansion, and antibody secretion in human TNP-specific B cells activated in the presence of TNP-ovalbumin (TNP-OA), pokeweed mitogen (PWM), or regulatory T cells. TNP-OA-responsive, hapten-specific PFC precursor cells occupy approximately 0.5% of all sIgM+/sIgD+ B cells in cord blood, bone marrow, peripheral blood, and tonsil. The PWM-responsive, hapten-specific PFC precursor pool is 70 to 90% smaller and does not express sIgD. Antigen-reactive B cells go through a minimum of three divisions in culture (six to nine PFC per clone), and antibody secretory rates of about 10(4) molecules IgM/cell/hr are achieved. In contrast, PWM-induced clone sizes were at least 60 PFC per clone, with antibody secretory rates of approximately 6 to 7 X 10(4) molecules IgM/cell/hr. Addition of high-dose carrier-primed suppressor T cells to limit dilution cultures reduced PFC precursor cell recruitment by up to 99%. However, in the few clones escaping from suppression, both clonal expansion and antibody secretory rates were much higher than in suppressor cell-free cultures, generating 30 to 60% of the antibody secreted in controls but with consequently much more restricted clonal diversity. When limiting dilution cultures were compared with standard microcultures of 2 X 10(5) cells, both clonal expansion and antibody secretory rates were much lower than expected, with a culture efficiency calculated to be 10 to 20% of that in low-density cultures. Our data suggest that the B cell subsets activated by antigen and by mitogen differ in their abilities for clonal expansion and antibody secretion. The hapten-specific and -responsive B cell family is expressed early in ontogeny, and in adults it is distributed evenly throughout the body. These limiting dilution experiments revealed that the primary effect of regulatory T cells is a drastic reduction in clonal diversity, and much less a mere reduction in overall response magnitude.     

Isolation of paraquat-tolerant mutants from tomato cell cultures

Summary Tomato callus clones selected for the ability to grow at paraquat concentrations lethal to wild-type cells were found at an approximate frequency of 5×10^–8 per viable cell. Diploid plants were regenerated from nine of the nineteen paraquat-tolerant callus clones isolated. Although some of these plants appeared normal, others had altered morphology and reduced vigor and fertility. New callus cultures initiated from these regenerated plants typically had at least a 30-fold increase over the wild type in tolerance to paraquat. Tests on callus from sexual progeny showed that the paraquat-tolerant phenotypes of clones PQT⁴, PQT⁶, and probably also PQT¹³ resulted from dominant nuclear mutations, but the number of loci involved is not yet known. Paraquat spray experiments indicated that slight paraquat-tolerance was expressed at the plant level in PQT¹³, but not in any of the other clones tested.     

A clonal analysis of glial lineages in neonatal forebrain development in vitro

Retrovirus-mediated gene transfer combined with triple immunostaining for astro- and oligodendroglial markers (antibodies to glial fibrillary acidic protein, GD3 ganglioside, and galactocerebroside, and the O4 antibody) was used to study clonal aspects of glial lineage in primary cultures of the neonatal rat striatum. We found two major clonal populations: astrocyte clones containing GFAP+, but GD3-, O4-, and GC- cells, and oligodendrocyte clones containing cells expressing various combinations of GD3, O4, and GC, with rare GFAP+ cells. These results indicate that astrocytes and oligodendrocytes belong to separate lineages in forebrain postnatal development.     

Differentiation potential of conditionally immortalized mesenchymal progenitor cells from adult marrow of a H-2Kb-tsA58 transgenic mouse

Primary cultures were initiated from marrow, spleen, and bone explants of an adult H-2K^b-tsA58 transgenic mouse (immortomouse). All cultures were initiated in immortalizing conditions, and an additional marrow culture was first incubated for 1 week in standard conditions and then switched to immortalizing conditions. Marrow cells immediately immortalized were designated the marrow immediate population (MIP); those immortalized after 1 week were termed the marrow delayed population (MDP). MIP and MDP cells both contained a mixture of fibroblastic or flattened cells, and the MIP cells contained an additional subpopulation of adipocytic (Oil Red-O positive) cells. Alkaline phosphatase expression was induced by dexamethasone (10⁻⁷ M) in MDP cells while MIP, spleen, and bone explant cells had only a low level of expression. MDP and MIP cells differentiated into bone when combined with porous calcium phosphate ceramics and implanted subcutaneously into nude mice while bone- and spleen-derived cells did not. Clones were isolated from the MDP and MIP cell populations and tested for differentiated phenotypes. Some MIP-derived clones exhibited adipocytic characteristics while MDP-derived subclones were negative. Histologic examination of porous ceramic implanted clones showed that all of the clones had osteogenic potential. Clones exposed to either dexamethasone, human recombinant bone morphogenetic protein-2, or horse serum plus hydrocortisone showed differences in expression of adipocytic or osteogenic markers. These immortalized cultures have retained both adipocytic and osteogenic potential even after 1 year of continuous culture, and provide a model system for clonal analysis of the developmental potential of marrow-derived mesenchymal precursor cells. © 1996 Wiley-Liss, Inc.     

Latitudinal variation in life-cycle characteristics of Potamogeton pectinatus L.: vegetative growth and asexual reproduction

Across latitudinal gradients, environmental conditions that influenceplant growth and reproduction largely change. Here we study clonal variation inlife-cycle characteristics of the cosmopolitan water plantPotamogetonpectinatus L. across a broad latitudinal range.Two consecutive experiments were performed under standardised laboratoryconditions (photoperiod, irradiance and temperature). In the first experimentweinvestigated asexual reproduction among fifteen clones, obtained from latitudesranging from 24 to 68° N. After 90 days of growth, high-latitude clonesproduced more but smaller tubers, while the aboveground biomass was lower ascompared to the clones obtained from low latitudes.In a second experiment we studied inherent differences in early growth,morphology and photosynthesis for eleven clones (obtained from the samelatitudinal range as in experiment 1). We found high among clonal variation formost measured variables, but the number of latitude-correlated traits waslimited. The only trait that correlated with latitude was the number of leavesper plant, which increased in clones from higher latitudes.Our results agree with the hypothesis of a latitude-correlated switch inlife-cycle strategy for this species. For northern clones this results in ashort life-cycle, with an early and high investment in tuber biomass, while forlow latitude clones the length of the life-cycle is prolonged, with a delayedreproduction and increased total plant biomass.     

EFFECTS OF FIRE ON CLONAL GROWTH AND DYNAMICS OF PITYOPSIS GRAMINIFOLIA (ASTERACEAE)

The rhizomatous perennial Pityopsis graminifolia was studied in a Florida sandhill community in an annually burned site, a periodically burned site, and a site that has been protected from fire since 1965. These different fire regimes significantly affected the demography and life histories of both plants and plant parts in this clonal species. Fires resulted in reductions in ramet biomass and height, and an increase in the (root + rhizome)/shoot biomass ratio. Burning also decreased the total number of flower heads and new rhizomes produced per ramet. However, the survivorship of initiated rhizomes was greater in burned sites and resulted in a larger number of established daughter ramets per clone. As a result, in burned sites there was a shift in clone structure toward larger numbers of smaller ramets, but there were no significant reductions in seed or rhizome production on a per genet basis. The results showed that the responses to fire in P. graminifolia are different when measured at the genet vs. ramet level and that the effects of fire on clones can be explained by demographic responses of plant parts. Population regeneration in the study sites was dependent on successful clonal ramet production because no seedling recruitment was observed. This suggests that disturbances other than fire are important for new genet recruitment in these clonal populations.     

QUANTITATIVE GENETIC ANALYSIS OF MORPHOLOGICAL VARIATION IN AN ANTARCTIC DIATOM GROWN AT TWO LIGHT INTENSITIES1

Ten clonal isolates of Thalassiosira tumida (Janisch) Hasle were grown in duplicate semi-continuous batch cultures at 116 and 11.6 μE.m⁻².s⁻¹; acclimated cells were harvested during exponential growth and cleaned for examination by light microscopy (LM) and scanning electron microscopy (SEM). Number of strutted processes surrounding the central annulus (SP) and average number of satellite pores per process (AVSAT) were counted using SEM on 20 valves from each culture grown in high light, for a total of 400 valves examined; number of marginal labiate processes (LP) and overall diameter (DIAM) were measured using LM on 20 valves from each culture grown in both high and low light for a total of 800 valves examined. Univariate analysis of variance showed that bottle effects resulting from microenvironmental differences between replicates were a small but significant source of variation in DIAM, LP, and SP but not AVSAT. Significant differences among clones were observed for all characters. Decreased irradiance resulted in a significant decrease in valve diameter but no significant effect on LP; no light x clone interaction was obsered. Significant covariance between characters among clones was also observed; since valve diameter is known to decrease during asexual growth, the correlation coefficients for SP, AVSAT, and LP with DIAM were used to correct the data for this source of nongenetic differences between clones. Analysis of the size-corrected data showed that the proportion of total phenotypic variance in SP, LP, and AVSAT caused by genetic differences among clones was 0.14, 0.14, and 0.30, respectively. This indicates that the majority of total phenotypic variance was due to environmental or developmental causes, but that sufficient genetic variability exists to support rapid phenotypic evolution in SP, LP, and AVSAT under continued directional selection. Finally, the results of the genetic analysis revealed a high (0.82) genetic correlation between SP and LP.     

Adolescence and the Reversibility of Maturity in Euplotes crassus

ABSTRACT. The clonal life history of ciliated protists is characterized by a sequence of phenotypes; sexual immaturity, maturity, and senescence. The distinctiveness of immaturity and maturity has been investigated. Standard assays of the onset of maturity of progeny clones from a cross between stocks EC1 and EC2 of Euplotes crassus demonstrated significant differences among clones and among testers within clones. They also revealed that the first positive test(s) of a progeny subclone were typically followed by at least one negative test. Special protocols were devised to investigate if maturity was reversible at the cellular level. In these experiments, the first mating pair of a progeny subclone was split before the consummation of mating. From these two cells as well as from control progeny and tester cells, subclones were established and every leftover cell was tested for maturity after each transfer. Both standard and split-pair progeny subclones had immature and slow- to-mate cells. The number of fissions before progeny exhibited sexual behavior indistinguishable from the testers was more than twice that to the first mating reaction of a subclone. At the first sign of maturity, progeny lines are a heterogeneous population of cells able and not able to mate, but remarkably, clonal descendants of those able to mate may become unable to mate. The development of maturity is progressive, quantitative and non-monotonic rather than an instantaneous switch.     

Clonal dynamics in western North American aspen (Populus tremuloides)

Clonality is a common phenomenon in plants, allowing genets to persist asexually for much longer periods of time than ramets. The relative frequency of sexual vs. asexual reproduction determines long‐term dominance and persistence of clonal plants at the landscape scale. One of the most familiar and valued clonal plants in North America is aspen (Populus tremuloides). Previous researchers have suggested that aspen in xeric landscapes of the intermountain west represent genets of great chronological age, maintained via clonal expansion in the near absence of sexual reproduction. We synthesized microsatellite data from 1371 ramets in two large sampling grids in Utah. We found a surprisingly large number of distinct genets, some covering large spatial areas, but most represented by only one to a few individual ramets at a sampling scale of 50 m. In general, multi‐ramet genets were spatially cohesive, although some genets appear to be fragmented remnants of much larger clones. We conclude that recent sexual reproduction in these landscapes is a stronger contributor to standing genetic variation at the population level than the accumulation of somatic mutations, and that even some of the spatially large clones may not be as ancient as previously supposed. Further, a striking majority of the largest genets in both study areas had three alleles at one or more loci, suggesting triploidy or aneuploidy. These genets tended to be spatially clustered but not closely related. Together, these findings substantially advance our understanding of clonal dynamics in western North American aspen, and set the stage for a broad range of future studies.     

Clonal diversity within infections and the virulence of a malaria parasite, Plasmodium mexicanum

Both verbal and mathematical models of parasite virulence predict that genetic diversity of microparasite infections will influence the level of costs suffered by the host. We tested this idea by manipulating the number of co-existing clones of Plasmodium mexicanum in its natural vertebrate host, the fence lizard Sceloporus occidentalis. We established replicate infections of P. mexicanum made up of 1, 2, 3, or >3 clones (scored using 3 microsatellite loci) to observe the influence of clone number on several measures of parasite virulence. Clonal diversity did not affect body growth or production of immature erythrocytes. Blood haemoglobin concentration was highest for the most genetically complex infections (equal to that of non-infected lizards), and blood glucose levels and rate of blood clotting was highest for the most diverse infections (with greater glucose and more rapid clotting than non-infected animals). Neither specific clones nor parasitaemia were associated with virulence. In this first experiment that manipulated the clonal diversity of a natural Plasmodium-host system, the cost of infection with 1 or 2 clones of P. mexicanum was similar to that previously reported for infected lizards, but the most complex infections had either no cost or could be beneficial for the host.     

Inhibition of T cell-mediated cytolysis by monoclonal antibodies directed against Lyt-2: heterogeneity of inhibition at the clonal level

The inhibitory effect of monoclonal anti-Lyt-2 antibodies on T cell-mediated cytolysis has been investigated at the clonal level. In agreement with previous reports from several laboratories, populations of cytolytic T lymphocytes (CTL) generated in vitro in mixed leukocyte cultures (MLC) were reversibly inhibited by monoclonal anti-Lyt-2 antibodies in a dose-dependent fashion. However, when alloimmune peritoneal exudate lymphocytes (PEL) were used as a source of CTL, little or no inhibitory effect of anti-Lyt-2 antibodies on cytolysis was observed. A series of CTL clones derived from MLC or PEL populations was also tested for inhibition of cytolysis by anti-Lyt-2 antibodies. In agreement with results obtained at the population level, most MLC-derived clones (81%) were strongly inhibited by the reagent, whereas few PEL clones (15%) were inhibited. Several of these clones were expanded and maintained in culture without loss of their "inhibition phenotype." Flow cytofluorometric analysis using the same monoclonal anti-Lyt-2 antibodies further revealed that both inhibited and uninhibited clones expressed comparable amounts of Lyt-2 antigen. These results provide direct evidence that inhibition of CTL by anti-Lyt-2 antibodies is heterogeneous at the clonal level. The possibility that this heterogeneity may be related to avidity of antigen receptors is discussed.