Raymond L. Tremblay

Host specificity and low reproductive success in the rare endemic Puerto Rican orchid Lepanthes caritensis

Tree host specificity is unusual in orchids. Here we show host specificity in a rare orchid and describe other potential causes of host specificity. Lepanthes caritensis occurs only in Carite State Forest, Puerto Rico. We found L. caritensis only on the largest individuals of one species of tree, Micropholis guyanensis, limiting the orchid to only 7% of the potential tree community > 5 cm dbh. Orchid occurrence was associated with high moss cover on M. guyanensis. The abundance of undescribed fungi cultured from tree bark was negatively associated with orchid presence. Low fruit production in orchids is not uncommon, but reproductive success in L. caritensis appeared to be unusually low; no fruits were produced by 68 adults during 16 months of observation. If this trend of low reproductive success is borne out over time, it would suggest an obvious explanation for the rarity of L. caritensis. Our results suggest ways to manage populations of L. caritensis, but also reveal aspects of tropical orchid biology (e.g. the importance of fungi and mosses for orchid establishment) which require more thorough investigation.

MICROBIAL QUALITY OF TROPICAL INLAND WATERS AND EFFECTS OF RAINFALL EVENTS

ABSTRACT Novel markers of fecal pollution in tropical waters are needed since conventional methods recommended for other geographical regions may not apply. To address this, the prevalence of thermotolerant coliforms, enterococci, coliphages, and enterophages was determined by culture methods across a watershed. Additionally, human-, chicken-, and cattle-specific PCR assays were used to identify potential fecal pollution sources in this watershed. An enterococcus quantitative PCR (qPCR) assay was tested and correlated with culture methods at three sites since water quality guidelines could incorporate this technique as a rapid detection method. Various rainfall events reported before sample collection at three sites were considered in the data analyses. Thermotolerant coliforms, enterococci, coliphages, and enterophages were detected across the watershed. Human-specific Bacteroides bacteria, unlike the cattle- and chicken-specific bacteria, were detected mostly at sites with the corresponding fecal impact. Enterococci were detected by qPCR as well, but positive correlations with the culture method were noted at two sites, suggesting that either technique could be used. However, no positive correlations were noted for an inland lake tested, suggesting that qPCR may not be suitable for all water bodies. Concentrations of thermotolerant coliforms and bacteriophages were consistently lower after rainfall events, pointing to a possible dilution effect. Rainfall positively correlated with enterococci detected by culturing and qPCR, but this was not the case for the inland lake. The toolbox of methods and correlations presented here could be potentially applied to assess the microbial quality of various water types.

Biology and natural history of Caladenia

The genus Caladenia comprises species that exhibit remarkable consistency in terms of growth form and phenological patterns. All taxa are herbaceous perennials, with the shoot arising annually from a single, usually spheroid, tuber and producing a single, hairy leaf. The tuber is annually replaced either side-by-side with the parent tuber or terminating a descending structure known as a dropper. The dropper is a depth-seeking mechanism that enables placement of the tuber at depth in the soil as a means to avoid predation by surface-foraging native mammals or away from the high temperatures and desiccating conditions during summer dormancy. The 3--5 attenuated roots produced at the junction between the tuber and shoot and produced late in the growing cycle and devoid of mycorrhiza suggest their functional significance may relate to water uptake. Mycorrhizal endophytes are confined to a hypertrophic stem region at the soil surface (collar) subtending the leaf that positions the collar directly in the organically rich zone at the soil surface. This morphology is a unique characteristic of several Australasian orchids in the tribe Diuridae. Mycorrhizal infection occurs rapidly, with maximum colonisation in concert with the onset of breaking rains. Pelotons are restricted to cortical cells, with fully developed pelotons throughout infected tissues within a week or so of soil wetting. Infection occurs as a ‘once-off’ event, with little evidence of secondary infection later in the growth cycle and no evidence of peloton digestion. Some taxa utilise vegetative propagation, often leading to localised clustering as for taxa in the ‘filamentosa’ complex or, extensive clonal mats as found in Caladenia flava and C. latifolia where daughter tubers are produced at the end of extending horizontal outgrowths. For the majority of taxa, plants remain dry-season (summer) dormant from a few months up to 7 months for arid-zone taxa, with shoot emergence from the tuber of temperate species thought to occur in response to a drop in the mean minimum temperature. Pollination biology of Caladenia is apparently through a process of deception, either as food or sexual mimics, with some taxa engaging in self-pollination. Here we review the natural history of Caladenia and acknowledge that much of our understanding is based on assumptions of the biology of terrestrial orchids in general and emphasise areas of research and biological enquiry that will be critical in the development of an effective conservation program for the genus.

When stable-stage equilibrium is unlikely: integrating transient population dynamics improves asymptotic methods

BACKGROUND AND AIMS Evaluation of population projection matrices (PPMs) that are focused on asymptotically based properties of populations is a commonly used approach to evaluate projected dynamics of managed populations. Recently, a set of tools for evaluating the properties of transient dynamics has been expanded to evaluate PPMs and to consider the dynamics of populations prior to attaining the stable-stage distribution, a state that may never be achieved in disturbed or otherwise ephemeral habitats or persistently small populations. This study re-evaluates data for a tropical orchid and examines the value of including such analyses in an integrative approach. METHODS Six small populations of Lepanthes rubripetala were used as a model system and the R software package popdemo was used to produce estimates of the indices for the asymptotic growth rate (lambda), sensitivities, reactivity, first-time step attenuation, maximum amplification, maximum attenuation, maximal inertia and maximal attenuation. The response in lambda to perturbations of demographic parameters using transfer functions and multiple perturbations on growth, stasis and fecundity were also determined. The results were compared with previously published asymptotic indices. KEY RESULTS It was found that combining asymptotic and transient dynamics expands the understanding of possible population changes. Comparison of the predicted density from reactivity and first-time step attenuation with the observed change in population size in two orchid populations showed that the observed density was within the predicted range. However, transfer function analysis suggests that the traditional approach of measuring perturbation of growth rates and persistence (inertia) may be misleading and is likely to result in erroneous management decisions. CONCLUSIONS Based on the results, an integrative approach is recommended using traditional PPMs (asymptotic processes) with an evaluation of the diversity of dynamics that may arise when populations are not at a stable-stage distribution (transient processes). This method is preferable for designing rapid and efficient interventions after disturbances, and for developing strategies to establish new populations.

Distribution of life cycle stages in a lithophytic and epiphytic orchid

Density-dependent processes may have multiple effects on populations, which among other things include the regulation of population abundance and of the relative distribution of life-cycle stages within populations. The epiphytic habitat is often characterized as highly ephemeral and therefore epiphytic orchid populations may never achieve density-dependent regulation. In this study, we investigated the potential for density-dependent regulation in epiphytic and lithophytic orchids by examining the association between seedlings, juvenile and adult life-history stages in the Caribbean endemic orchid,Lepanthes rupestris in a cross-sectional study of 179 populations surveyed in the Luquillo National Forest along a riparian area where it is locally abundant. Under density-dependent regulation we expected a negative association between the ratio of seedling/adults and juveniles/adults and total population density. Population density was in the range of 140 individuals per m2, however patch sizes were small and mostly limited to less than 0.5 m2 with a maximum of 3 m2. We found no evidence of reduction of the ratio of seedlings or juveniles to adults as population size increased in either tree or boulder populations suggesting negative density dependence for population regulation inL. rupestris is either rare or occurs at even higher densities than those measured here. Moreover, we found positive (although weak) relationship between the ratio of seedlings and juveniles to adults and population size, suggesting that facilitation may be occurring.

A new species of Lepanthes (Orchidaceae) from Puerto Rico

A new species,Lepanthes caritensis, is described and illustrated. There are now nine species ofLepanthes known to occur in Puerto Rico. Plant habitat and floral morphology ofL. caritensis is most similar to that ofL. sanguinea. A survey of the area where the new species is located suggests that it is rare and should be protected under the Federal Endangered Species Act.ResumenUna especic nueva,Lepanthes caritensis, es descrita e ilustrada para Puerto Rico. Con ésta hay ahora nueve especies deLepanthes en Puerto Rico. El hábito de la planta y la morfología de la flor deL. caritensis es más similar aL. sanguinea. Un muestreo del área donde se encuentra la nueva especie sugiere que ésta es rara y que debería ser protegida por el Federal Endangered Species Act.

Riding across the selection landscape: fitness consequences of annual variation in reproductive characteristics

Evolutionary models estimating phenotypic selection in character size usually assume that the character is invariant across reproductive bouts. We show that variation in the size of reproductive traits may be large over multiple events and can influence fitness in organisms where these traits are produced anew each season. With data from populations of two orchid species, Caladenia valida and Tolumnia variegata, we used Bayesian statistics to investigate the effect on the distribution in fitness of individuals when the fitness landscape is not flat and when characters vary across reproductive bouts. Inconsistency in character size across reproductive periods within an individual increases the uncertainty of mean fitness and, consequently, the uncertainty in individual fitness. The trajectory of selection is likely to be muddled as a consequence of variation in morphology of individuals across reproductive bouts. The frequency and amplitude of such changes will certainly affect the dynamics between selection and genetic drift.

Demographic response by a small epiphytic orchid

PREMISE OF THE STUDY Biotic changes are an inevitable consequence of climate change. Epiphytes may be more susceptible to changes in climate variation, but data regarding responses to climate variability under field conditions are limited. We evaluated whether the abundance of demographic stages in the epiphytic orchid Lepanthes rupestris at the Luquillo Experimental Forest in Puerto Rico was associated with short-term changes in climate variation over an 8-yr period. METHODS We used cross-correlation analyses to evaluate associations between the abundance of seedlings, juveniles, adults, and fruits per subpopulation, population growth, colonization and extinction rates in L. rupestris with variables related to precipitation and temperature, with and without lag- responses. KEY RESULTS We detected significant negative correlations between the average number of seedlings and the number of dry days, between the average number of fruits and minimum average temperature with a 6-mo response lag, and between the average number of adults and the maximum temperature with a 1-yr response lag. Neither population growth rate nor probability of colonization and extinction were directly related to climatic variation between 2000 and 2007. CONCLUSIONS Associations between climatic variables and demographic stages could have negative implications for this orchid within the context of expected drying trends in the Caribbean. Results argue for the establishment of long-term monitoring studies of orchid populations, because only long-term studies would provide the appropriate temporal scale to detect and predict climate change effects and adaptive management of orchid populations.

Immunomodulator-Based Enhancement of Anti Smallpox Immune Responses

Background The current live vaccinia virus vaccine used in the prevention of smallpox is contraindicated for millions of immune-compromised individuals. Although vaccination with the current smallpox vaccine produces protective immunity, it might result in mild to serious health complications for some vaccinees. Thus, there is a critical need for the production of a safe virus-free vaccine against smallpox that is available to everyone. For that reason, we investigated the impact of imiquimod and resiquimod (Toll-like receptors agonists), and the codon-usage optimization of the vaccinia virus A27L gene in the enhancement of the immune response, with intent of producing a safe, virus-free DNA vaccine coding for the A27 vaccinia virus protein. Methods We analyzed the cellular-immune response by measuring the IFN-γ production of splenocytes by ELISPOT, the humoral-immune responses measuring total IgG and IgG2a/IgG1 ratios by ELISA, and the TH1 and TH2 cytokine profiles by ELISA, in mice immunized with our vaccine formulation. Results The proposed vaccine formulation enhanced the A27L vaccine-mediated production of IFN-γ on mouse spleens, and increased the humoral immunity with a TH1-biased response. Also, our vaccine induced a TH1 cytokine milieu, which is important against viral infections. Conclusion These results support the efforts to find a new mechanism to enhance an immune response against smallpox, through the implementation of a safe, virus-free DNA vaccination platform.

Dormancy in Caladenia: a Bayesian approach to evaluating latency

Dormancy is common in many terrestrial orchids in southern Australia and other temperate environments. The difficulty for conservation and management when considering dormancy is ascertaining whether non-emergent plants aredormantordead.Hereweuseamulti-statecapture-recapturemethod,undertakenoverseveralseasons,todeterminethe likelihood of a plant becoming dormant or dying following its annual emergent period and evaluate the frequency of the lengthofdormancy.Weassessthetransitionprobabilitiesfromtimeseriesofvaryinglengthsforthefollowingnineterrestrial orchids in the genus Caladenia: C. amoena, C. argocalla, C. clavigera, C. elegans, C. graniticola, C. macroclavia, C. oenochila,C. rosella and C. valida from Victoria, South Australia and Western Australia. We used a Bayesian approach for estimating survivorship, dormancy and the likelihood of death from capture-recapture data. Considering all species together, the probability of surviving from one year to the next was ~86%, whereas the likelihood of observing an individual above ground in two consecutive years was ~74%. All species showed dormancy of predominantly 1 year, whereasdormancyofthreeormoreyearswasextremelyrare(<2%).Theresultshavepracticalimplicationsforconservation, in that (1) population sizes of Caladenia species are more easily estimated by being able to distinguish the likelihood of an unseen individual being dormant or dead, (2) population dynamics of individuals can be evaluated by using a 1-3-year dormancy period and (3) survey effort is not wasted on monitoring individuals that have not emerged for many years.