Seasonal Time Points Research Articles

Proteomic Identification of Seasonally Expressed Proteins Contributing to Heart Function and the Avoidance of Skeletal Muscle Disuse Atrophy in a Hibernating Mammal.

Hibernation in the thirteen-lined ground squirrel (Ictidomys tridecemlineatus) takes place over 4-6 months and is characterized by multiday bouts of hypothermic torpor (5-7 °C core body temperature) that are regularly interrupted every 1-2 weeks by brief (12-24 h) normothermic active periods called interbout arousals. Our goal was to gain insight into the molecular mechanisms that underlie the hibernator's ability to preserve heart function and avoid the deleterious effects of skeletal muscle disuse atrophy over prolonged periods of inactivity, starvation, and near-freezing body temperatures. To achieve this goal, we performed organelle enrichment of heart and skeletal muscle at five seasonal time points followed by LC-MS-based label-free quantitative proteomics. In both organs, we saw an increase in the levels of many proteins as ground squirrels transition from an active state to a prehibernation state in the fall. Interestingly, seasonal abundance patterns identified DHRS7C, SRL, TRIM72, RTN2, and MPZ as potential protein candidates for mitigating disuse atrophy in skeletal muscle, and ex vivo contractile mechanics analysis revealed no deleterious effects in the ground squirrel's muscles despite prolonged sedentary activity. Overall, an increased understanding of protein abundance in hibernators may enable novel therapeutic strategies to treat muscle disuse atrophy and heart disease in humans.

Seasonal variation in telomerase activity and telomere dynamics in a hibernating rodent, the garden dormouse (Eliomys quercinus).

Telomere dynamics in hibernating species are known to reflect seasonal changes in somatic maintenance. Throughout hibernation, the periodic states of rewarming, known as inter-bout euthermia or arousals, are associated with high metabolic costs including shortening of telomeres. In the active season, if high energetic resources are available, telomere length can be restored in preparation for the upcoming winter. The mechanism for telomere elongation has not been clearly demonstrated, although the action of the ribonucleoprotein complex, telomerase, has been implicated in many species. Here we tested for levels of telomerase activity in the garden dormouse (Eliomys quercinus) at different seasonal time points throughout the year and across ages from liver tissues of male juveniles to adults. We found that telomerase is active at high levels across seasons (during torpor and inter-bout euthermia, plus in the active season) but that there was a substantial decrease in activity in the month prior to hibernation. Telomerase levels were consistent across age groups and were independent of feeding regime and time of birth (early or late born). The changes in activity levels that we detected were broadly associated with changes in telomere lengths measured in the same tissues. We hypothesise that i) telomerase is the mechanism used by garden dormice for maintenance of telomeres and that ii) activity is kept at high levels throughout the year until pre-hibernation when resources are diverted to increasing fat reserves for overwintering. We found no evidence for a decrease in telomerase activity with age or a final increase in telomere length which has been detected in other hibernating rodents.

Effect of long-term conservation tillage management on microbial diversity under Mediterranean rainfed conditions

Conservation Agriculture has been the subject of numerous studies for decades. However, its impact on the soil microbial community has often been overlooked. Recent studies have shed light on the changes in soil bacterial composition resulting from modifications in tillage practices, particularly when compared to conventional tillage methods. However, there is a lack of soil microbial community analysis conducted under conservation tillage and rainfed conditions in the Mediterranean basin. A metataxonomic approach was employed to assess the influence of 19 years (2004–2022) of minimum tillage (MT) and no-till (NT) practices, along with crop rotation, on the soil bacterial community at upper layers (15 cm depth). This field study involved three different seasonal moments throughout the year. Results revealed that both sampling time and tillage management significantly influenced the structure of the bacterial community, whereas no effect was observed due to the crop. May showed a higher taxonomic diversity than July and December, showing differences in some o fthe most abundant phyla, such as Actinobacteria, Firmicutes and Proteobacteria. On the other hand, the long-term application of NT was found to lead to a significant decrease in bacterial diversity, with lower abundances of the phyla Chloroflexi, Gemmatimonadetes, and Armatimonadota, as well as an increase in k-strategist bacteria compared to MT. However, no significant effect on alpha diversity was observed as a result of tillage. no variations in phyla composition were observed attributable to crop, as the primary distinctions were identified among legume crops, which were associated with different sampling times. Therefore, our conclusion is that over this period, NT did not promote microbial diversity in the upper soil layers, despite notable shifts in bacterial composition occurring across seasonal time points in both tillage management systems.

Skinfold thickness in elite male professional football players: changes across 3 seasons including a COVID-19 lockdown period

ABSTRACT This study investigates variations in skinfold thickness in a European professional football team within and across multiple seasons, including before and after COVID-19 restrictions. Skinfold thickness was measured in 49 male players on 32 occasions across three seasons (2018/19, 2019/20 and 2020/21). Intra- and inter-seasonal trends were examined by comparing values for the sum of eight skinfolds (sum8) at key time points using linear mixed models. In each season, the largest differences in sum8 occurred after the preseason period. These differences ranged from a mean difference (Mdiff) of −11.0 mm (p < 0.001) in 2018/19 to −4.8 mm (p < 0.001) in 2020/21. Increases in sum8 occurred after each off-season; however, the increase after the off-season before the 2020/21 season (which included the COVID-19 lockdown period) was the lowest in magnitude and was not statistically significant (Mdiff = 2.9 mm, p = 0.06). Changes occurring in sum8 between the start and end of the competitive season did not follow a consistent pattern and were generally smaller in magnitude than the changes between other seasonal time points. Despite the notable heterogeneity in sum8 between players, changes within players generally followed a similar pattern between the different seasonal timepoints. In summary, changes in skinfold thickness were predominantly related to off-season and ‘preparation for competition’ periods. Increases in skinfold thickness can be mitigated during extended remote training periods with appropriate training and nutritional strategies.

Mass spectrometry of the white adipose metabolome in a hibernating mammal reveals seasonal changes in alternate fuels and carnitine derivatives.

Mammalian hibernators undergo substantial changes in metabolic function throughout the seasonal hibernation cycle. We report here the polar metabolomic profile of white adipose tissue isolated from active and hibernating thirteen-lined ground squirrels (Ictidomys tridecemlineatus). Polar compounds in white adipose tissue were extracted from five groups representing different timepoints throughout the seasonal activity-torpor cycle and analyzed using hydrophilic interaction liquid chromatography-mass spectrometry in both the positive and negative ion modes. A total of 224 compounds out of 660 features detected after curation were annotated. Unsupervised clustering using principal component analysis revealed discrete clusters representing the different seasonal timepoints throughout hibernation. One-way analysis of variance and feature intensity heatmaps revealed metabolites that varied in abundance between active and torpid timepoints. Pathway analysis compared against the KEGG database demonstrated enrichment of amino acid metabolism, purine metabolism, glycerophospholipid metabolism, and coenzyme A biosynthetic pathways among our identified compounds. Numerous carnitine derivatives and a ketone that serves as an alternate fuel source, beta-hydroxybutyrate (BHB), were among molecules found to be elevated during torpor. Elevated levels of the BHB-carnitine conjugate during torpor suggests the synthesis of beta-hydroxybutyrate in white adipose mitochondria, which may contribute directly to elevated levels of circulating BHB during hibernation.

IgE-binding monocytes upregulate the coagulation cascade in allergic horses

IgE-binding monocytes are a rare peripheral immune cell type involved in the allergic response through binding of IgE on their surface. IgE-binding monocytes are present in both healthy and allergic individuals. We performed RNA sequencing to ask how the function of IgE-binding monocytes differs in the context of allergy. Using a large animal model of allergy, equine Culicoides hypersensitivity, we compared the transcriptome of IgE-binding monocytes in allergic and non-allergic horses at two seasonal timepoints: (i) when allergic animals were clinical healthy, in the winter “Remission Phase”, and (ii) during chronic disease, in the summer “Clinical Phase”. Most transcriptional differences between allergic and non-allergic horses occurred only during the “Remission Phase”, suggesting principal differences in monocyte function even in the absence of allergen exposure. F13A1, a subunit of fibrinoligase, was significantly upregulated at both timepoints in allergic horses. This suggested a role for increased fibrin deposition in the coagulation cascade to promote allergic inflammation. IgE-binding monocytes also downregulated CCR10 expression in allergic horses during the “Clinical Phase”, suggesting a defect in maintenance of skin homeostasis, which further promotes allergic inflammation. Together, this transcriptional analysis provides valuable clues into the mechanisms used by IgE-binding monocytes in allergic individuals.

Dengue fever occurrence in India, Brazil, Paraguay, Philippines and Singapore using Google trends

Background. Dengue is a mosquito-borne disease and its occurrence is a worldwide problem. The health seekers may turn to the Internet based data for possible complications and various therapeutic approaches for the treatment. Therefore, our objective is to identify and characterize the seasonal transition of dengue fever using Google Trends. Methods. This was a cross-sectional study of dengue fever internet searches between 2004 and 2017. The Google was indexed the aggregated web data and ranked based on the search volume with respect to time. The countries selected for the study include India, Brazil, Paraguay, Philippines and Singapore based on the maximum number of cases across world. The selected regions were further classified in to seasonal time points and the relative search volume ranks were categorized to two season variables. Mann–Whitney U test was applied to identify the statistical significance. Results. The summer to monsoon transitions were showed highest number cases beginning from 2004 in the selected regions. Furthermore, dengue searches were peaked in both the seasons of summer and monsoon in selected regions. The analyses of data further revealed that 16.2 % of variation in dengue trends is associated with the time between 2004 and 2017.

Identifying climate‐resistant vernal pools: Hydrologic refugia for amphibian reproduction under droughts and climate change

AbstractVernal pools of the northeastern United States provide important breeding habitat for amphibians but may be sensitive to droughts and climate change. These seasonal wetlands typically fill by early spring and dry by mid‐to‐late summer. Because climate change may produce earlier and stronger growing‐season evapotranspiration combined with increasing droughts and shifts in precipitation timing, management concerns include the possibility that some pools will increasingly become dry earlier in the year, potentially interfering with amphibian life‐cycle completion. In this context, a subset of pools that continues to provide wetland habitat later into the year under relatively dry conditions might function as ecohydrologic refugia, potentially supporting species persistence even as summer conditions become warmer and droughts more frequent. We used approximately 3000 field observations of inundation from 449 pools to train machine‐learning models that predict the likelihood of pool inundation based on pool size, day of the year, climate conditions, short‐term weather patterns, and soil, geologic and landcover attributes. Models were then used to generate predictions of pool wetness across five seasonal time points, three short‐term weather scenarios and four sets of downscaled climate projections. Model outputs are available through a website allowing users to choose the inundation thresholds, time points, weather scenarios and future climate projections most relevant to their management needs. Together with long‐term monitoring of individual pools at the site scale, this regional‐scale study can support amphibian conservation by helping to identify which pools may be most likely to function as ecohydrologic refugia from droughts and climate change.

Trained immunity and tolerance in innate lymphoid cells, monocytes, and dendritic cells during allergen-specific immunotherapy

Despite the efficacy of allergen-specific immunotherapy (AIT), the role of trained immunity and tolerance in this process has not been elucidated. Here, we have performed a comprehensive longitudinal analysis of the systemic innate immune cell repertoire during the course of AIT. Patients with allergy received standard preseasonal subcutaneous AIT with allergoids to birch and/or grass. Healthy controls were monitored without any intervention. Flow cytometry of innate lymphoid cell (ILC), natural killer cell, monocyte cell, and dendritic cell (DC) subsets was performed at baseline, 3 months (birch season), 6 months (grass seasons), and 12 months after the therapy in patients or at similar seasonal time points in controls. Additional analyses were performed in the third-year birch and grass season. We observed a durable decrease in group 2 ILCs and an increase of group 1 ILCs after AIT, with dynamic changes in their composition. We found that an expansion of CD127+CD25++ clusters caused observed shifts in the heterogeneity of group 1 ILCs. In addition, we observed development of CD127+CD25++c-Kit+ group 3 ILC clusters. Moreover, we found an increase in the number of intermediate monocytes in parallel with a reduction in nonclassical monocytes during the first year after AIT. Classical and intermediate monocytes presented significant heterogeneity in patients with allergy, but AIT reduced the HLA-DR++ clusters. Finally, an increase in plasmacytoid DCs and CD141+ myeloid DCs was observed in individuals with allergy, whereas the number of CD1c+ myeloid DCs was reduced during the first year of AIT. AIT induces changes in the composition and heterogeneity of circulating innate immune cells and brings them to the level observed in healthy individuals. Monitoring of ILCs, monocytes, and DCs during AIT might serve as a novel biomarker strategy.

An initial industrial flora: A framework for botanical research in cooperation with industry for biodiversity conservation.

Humans have created an accelerating, increasingly connected, globalized economy, resulting in a more globalized, shared flora. The prevention of new, establishing species is less costly, both economically and ecologically, and is more manageable than eradicating nonnative invasive species once they are widespread and negatively impactful. We ask if international trade hubs and points-of-entry with high-volume trade, constant disturbance, and propagule rain have a higher number of nonnative species compared to surrounding areas and if they may serve as initial establishment sites and refugia of nonnative, invasive populations. Therefore, we partnered with various federal, state, and private interests to evaluate the floristic composition at the Garden City Terminal of the Port of Savannah, Georgia, USA. We conducted the following study to demonstrate the collaborative relationship-building between researchers and industry and to develop a framework for biodiversity conservation. In our study, we collected all reproductive vascular plants in the secured areas of the Garden City Terminal during four major seasonal time points over two years. The percent of nonnative species and number of nonnative plant species per hectare at this industrial location exceeded all other comparison floras. The mean coefficient of conservatism was lowest among the comparison floras, indicating a highly disturbed habitat with nonnative, weedy native, and other native species tolerant of disturbance. Our study represents one of the first inventories of an Industrialized Flora and indicates that such areas are hot-spots of nonnative plant diversity and possible sources of emergent plant invasions. We posit that industrial sites and international points-of-entry should be considered laboratories for research on species transport and introduction, adaptability, and taxonomic delineation to better understand the mechanisms and consequences of biotic homogenization due to the volume and frequency of anthropogenic activities.

Longitudinal Comparison of Bacterial Diversity and Antibiotic Resistance Genes in New York City Sewage.

Bacterial resistance to antibiotics is a pressing health issue around the world, not only in health care settings but also in the community and environment, particularly in crowded urban populations. The aim of our work was to characterize the microbial populations in sewage and the spread of antibiotic resistance within New York City (NYC). Here, we investigated the structure of the microbiome and the prevalence of antibiotic resistance genes in raw sewage samples collected from the fourteen NYC Department of Environmental Protection wastewater treatment plants, distributed across the five NYC boroughs. Sewage, a direct output of anthropogenic activity and a reservoir of microbes, provides an ecological niche to examine the spread of antibiotic resistance. Taxonomic diversity analysis revealed a largely similar and stable bacterial population structure across all the samples, which was found to be similar over three time points in an annual cycle, as well as in the five NYC boroughs. All samples were positive for the presence of the seven antibiotic resistance genes tested, based on real-time quantitative PCR assays, with higher levels observed for tetracycline resistance genes at all time points. For five of the seven genes, abundances were significantly higher in May than in February and August. This study provides characteristics of the NYC sewage resistome in the context of the overall bacterial populations.IMPORTANCE Urban sewage or wastewater is a diverse source of bacterial growth, as well as a hot spot for the development of environmental antibiotic resistance, which can in turn influence the health of the residents of the city. As part of a larger study to characterize the urban New York City microbial metagenome, we collected raw sewage samples representing three seasonal time points spanning the five boroughs of NYC and went on to characterize the microbiome and the presence of a range of antibiotic resistance genes. Through this study, we have established a baseline microbial population and antibiotic resistance abundance in NYC sewage which can prove to be very useful in studying the load of antibiotic usage, as well as for developing effective measures in antibiotic stewardship.

Season structures prokaryotic partners but not algal symbionts in subtropical hard corals.

Coral reef ecosystems have great economic, social, and ecological value. The ecological success of coral reef ecosystems critically depends on coral-algal symbiosis and coral-prokaryotic partnership. However, seasonal changes underlying these relationships in subtropical hard corals of Hong Kong are poorly studied. Here, we compared the community changes of algal symbionts and prokaryotic partners in Platygyra carnosa and Galaxea fascicularis from Hong Kong collected at two seasonal time points of winter and summer via massively parallel sequencing of genetic markers and multivariate analysis. The results indicated that algal symbionts showed no significant changes between the two seasonal time points but prokaryotic partners changed substantially. Prokaryotic partners putatively involved in photosynthesis, nitrogen fixation, and sulfur oxidation increased significantly from winter to summer, while prokaryotic partners potentially associated with chemosynthesis, ammonia oxidation, and nitrite oxidation decreased significantly from winter to summer. Dissolved oxygen and pH served as the main contributors influencing prokaryotic partners in winter, while temperature, turbidity, and salinity played a dominant role in shaping prokaryotic partners in summer. The findings of the present study suggest that season structures prokaryotic partners but not algal symbionts in subtropical hard corals.

Season, but not symbiont state, drives microbiome structure in the temperate coral Astrangia poculata

BackgroundUnderstanding the associations among corals, their photosynthetic zooxanthella symbionts (Symbiodinium), and coral-associated prokaryotic microbiomes is critical for predicting the fidelity and strength of coral symbioses in the face of growing environmental threats. Most coral-microbiome associations are beneficial, yet the mechanisms that determine the composition of the coral microbiome remain largely unknown. Here, we characterized microbiome diversity in the temperate, facultatively symbiotic coral Astrangia poculata at four seasonal time points near the northernmost limit of the species range. The facultative nature of this system allowed us to test seasonal influence and symbiotic state (Symbiodinium density in the coral) on microbiome community composition.ResultsChange in season had a strong effect on A. poculata microbiome composition. The seasonal shift was greatest upon the winter to spring transition, during which time A. poculata microbiome composition became more similar among host individuals. Within each of the four seasons, microbiome composition differed significantly from that of surrounding seawater but was surprisingly uniform between symbiotic and aposymbiotic corals, even in summer, when differences in Symbiodinium density between brown and white colonies are the highest, indicating that the observed seasonal shifts are not likely due to fluctuations in Symbiodinium density.ConclusionsOur results suggest that symbiotic state may not be a primary driver of coral microbial community organization in A. poculata, which is a surprise given the long-held assumption that excess photosynthate is of importance to coral-associated microbes. Rather, other environmental or host factors, in this case, seasonal changes in host physiology associated with winter quiescence, may drive microbiome diversity. Additional studies of A. poculata and other facultatively symbiotic corals will provide important comparisons to studies of reef-building tropical corals and therefore help to identify basic principles of coral microbiome assembly, as well as functional relationships among holobiont members.

Genetic evidence that the Makira region in northeastern Madagascar is a hotspot of malaria transmission.

BackgroundEncouraging advances in the control of Plasmodium falciparum malaria have been observed across much of Africa in the past decade. However, regions of high relative prevalence and transmission that remain unaddressed or unrecognized provide a threat to this progress. Difficulties in identifying such localized hotspots include inadequate surveillance, especially in remote regions, and the cost and labor needed to produce direct estimates of transmission. Genetic data can provide a much-needed alternative to such empirical estimates, as the pattern of genetic variation within malaria parasite populations is indicative of the level of local transmission. Here, genetic data were used to provide the first empirical estimates of P. falciparum malaria prevalence and transmission dynamics for the rural, remote Makira region of northeastern Madagascar.MethodsLongitudinal surveys of a cohort of 698 total individuals (both sexes, 0–74 years of age) were performed in two communities bordering the Makira Natural Park protected area. Rapid diagnostic tests, with confirmation by molecular methods, were used to estimate P. falciparum prevalence at three seasonal time points separated by 4-month intervals. Genomic loci in a panel of polymorphic, putatively neutral markers were genotyped for 94 P. falciparum infections and used to characterize genetic parameters known to correlate with transmission levels.ResultsOverall, 27.8% of individuals tested positive for P. falciparum over the 10-month course of the study, a rate approximately sevenfold higher than the countrywide average for Madagascar. Among those P. falciparum infections, a high level of genotypic diversity and a high frequency of polygenomic infections (68.1%) were observed, providing a pattern consistent with high and stable transmission.ConclusionsPrevalence and genetic diversity data indicate that the Makira region is a hotspot of P. falciparum transmission in Madagascar. This suggests that the area should be highlighted for future interventions and that additional areas of high transmission may be present in ecologically similar regions nearby.Electronic supplementary materialThe online version of this article (doi:10.1186/s12936-016-1644-4) contains supplementary material, which is available to authorized users.

In-situ measurement of metabolic status in three coral species from the Florida Reef Tract

The goal of this study was to gain an understanding of intra- and inter-specific variation in calcification rate, lipid content, symbiont density, and chlorophyll a of corals in the Florida Reef Tract to improve our insight of in-situ variation and resilience capacity in coral physiology. The Florida Keys are an excellent place to assess this question regarding resilience because coral cover has declined dramatically since the late 1970s, yet has remained relatively high on some inshore patch reefs. Coral lipid content has been shown to be an accurate predictor of resilience under stress, however much of the current lipid data in the literature comes from laboratory-based studies, and previous in-situ lipid work has been highly variable. The calcification rates of three species were monitored over a seven-month period at three sites and lipid content was quantified at two seasonal time points at each of the three sites. Montastraea cavernosa had the highest mean calcification rate (4.7 mg cm−2 day−1) and lowest mean lipid content (1.6 mg cm−2) across sites and seasons. In contrast, Orbicella faveolata and Porites astreoides had lower mean calcification rates (2.8 mg cm−2 day−1 and 2.4 mg cm−2 day−1, respectively) and higher mean lipid contents (3.5 mg cm−2 and 2.3 mg cm−2, respectively) across sites and seasons. Given the recent Endangered Species Act (ESA) listing of O. faveolata and the relative persistence of M. cavernosa and P. astreoides on a population-scale, this study suggests that the hypothesis that coral lipids are good indicators of resilience may be species-specific, or more complex and interrelated with other environmental factors than previously understood. Additionally, coral lipid storage under benign thermal conditions may differ from lipid storage before, during, and after thermal stress events.

Neural transcriptome reveals molecular mechanisms for temporal control of vocalization across multiple timescales.

BackgroundVocalization is a prominent social behavior among vertebrates, including in the midshipman fish, an established model for elucidating the neural basis of acoustic communication. Courtship vocalizations produced by territorial males are essential for reproductive success, vary over daily and seasonal cycles, and last up to hours per call. Vocalizations rely upon extreme synchrony and millisecond precision in the firing of a homogeneous population of motoneurons, the vocal motor nucleus (VMN). Although studies have identified neural mechanisms driving rapid, precise, and stable neuronal firing over long periods of calling, little is known about underlying genetic/molecular mechanisms.ResultsWe used RNA sequencing-based transcriptome analyses to compare patterns of gene expression in VMN to the surrounding hindbrain across three daily and seasonal time points of high and low sound production to identify candidate genes that underlie VMN’s intrinsic and network neuronal properties. Results from gene ontology enrichment, enzyme pathway mapping, and gene category-wide expression levels highlighted the importance of cellular respiration in VMN function, consistent with the high energetic demands of sustained vocal behavior. Functionally important candidate genes upregulated in the VMN, including at time points corresponding to high natural vocal activity, encode ion channels and neurotransmitter receptors, hormone receptors and biosynthetic enzymes, neuromodulators, aerobic respiration enzymes, and antioxidants. Quantitative PCR and RNA-seq expression levels for 28 genes were significantly correlated. Many candidate gene products regulate mechanisms of neuronal excitability, including those previously identified in VMN motoneurons, as well as novel ones that remain to be investigated. Supporting evidence from previous studies in midshipman strongly validate the value of transcriptomic analyses for linking genes to neural characters that drive behavior.ConclusionsTranscriptome analyses highlighted a suite of molecular mechanisms that regulate vocalization over behaviorally relevant timescales, spanning milliseconds to hours and seasons. To our knowledge, this is the first comprehensive characterization of gene expression in a dedicated vocal motor nucleus. Candidate genes identified here may belong to a conserved genetic toolkit for vocal motoneurons facing similar energetic and neurophysiological demands.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-1577-2) contains supplementary material, which is available to authorized users.

Characterization of wastewater treatment plant microbial communities and the effects of carbon sources on diversity in laboratory models.

We are developing a laboratory-scale model to improve our understanding and capacity to assess the biological risks of genetically engineered bacteria and their genetic elements in the natural environment. Our hypothetical scenario concerns an industrial bioreactor failure resulting in the introduction of genetically engineered bacteria to a downstream municipal wastewater treatment plant (MWWTP). As the first step towards developing a model for this scenario, we sampled microbial communities from the aeration basin of a MWWTP at three seasonal time points. Having established a baseline for community composition, we investigated how the community changed when propagated in the laboratory, including cell culture media conditions that could provide selective pressure in future studies. Specifically, using PhyloChip 16S-rRNA-gene targeting microarrays, we compared the compositions of sampled communities to those of inocula propagated in the laboratory in simulated wastewater conditionally amended with various carbon sources (glucose, chloroacetate, D-threonine) or the ionic liquid 1-ethyl-3-methylimidazolium chloride ([C2mim]Cl). Proteobacteria, Bacteroidetes, and Actinobacteria were predominant in both aeration basin and laboratory-cultured communities. Laboratory-cultured communities were enriched in γ-Proteobacteria. Enterobacteriaceae, and Aeromonadaceae were enriched by glucose, Pseudomonadaceae by chloroacetate and D-threonine, and Burkholderiacea by high (50 mM) concentrations of chloroacetate. Microbial communities cultured with chloroacetate and D-threonine were more similar to sampled field communities than those cultured with glucose or [C2mim]Cl. Although observed relative richness in operational taxonomic units (OTUs) was lower for laboratory cultures than for field communities, both flask and reactor systems supported phylogenetically diverse communities. These results importantly provide a foundation for laboratory models of industrial bioreactor failure scenarios.

Investigating patterns of symbiotic nitrogen fixation during vegetation change from grassland to woodland using fine scale δ(15) N measurements.

Biological nitrogen fixation (BNF) in woody plants is often investigated using foliar measurements of δ(15) N and is of particular interest in ecosystems experiencing increases in BNF due to woody plant encroachment. We sampled δ(15) N along the entire N uptake pathway including soil solution, xylem sap and foliage to (1) test assumptions inherent to the use of foliar δ(15) N as a proxy for BNF; (2) determine whether seasonal divergences occur between δ(15) Nxylem sap and δ(15) Nsoil inorganic N that could be used to infer variation in BNF; and (3) assess patterns of δ(15) N with tree age as indicators of shifting BNF or N cycling. Measurements of woody N-fixing Prosopis glandulosa and paired reference non-fixing Zanthoxylum fagara at three seasonal time points showed that δ(15) Nsoil inorganic N varied temporally and spatially between species. Fractionation between xylem and foliar δ(15) N was consistently opposite in direction between species and varied on average by 2.4‰. Accounting for these sources of variation caused percent nitrogen derived from fixation values for Prosopis to vary by up to ∼70%. Soil-xylem δ(15) N separation varied temporally and increased with Prosopis age, suggesting seasonal variation in N cycling and BNF and potential long-term increases in BNF not apparent through foliar sampling alone.

Comparison of fertility and seasonal effects on grassland microbial communities

The activity of saprotrophic fungi and bacteria, and the balance between them, can affect decomposition. Arbuscular mycorrhizal (AM) fungi are also important for the nutrient and energy transfer in soil. Microbial community composition and activity are believed to have seasonal patterns, and are known to be highly influenced by environmental factors such as pH and nutrient conditions. To evaluate the importance of season for the variation in microbial decomposer community in a context of well-known environmental factor variation, we studied microbial growth, biomass and community structure along a fertility gradient (pH 5.9–8.1; NH4-N 3–19 μg g−1 soil, f.w.) in a sandy grassland during one year. The microbial community structure (phospholipid fatty acid (PLFA) composition) and biomass (PLFA and neutral lipid fatty acid (NLFA) signatures) as well as fungal (acetate incorporation in ergosterol) and bacterial (leucine incorporation) growth rates were investigated at eight seasonal time points during one year. The environmental factors pH and NH4 concentrations explained a larger share of the variation in the microbial community structure. Together they explained 37% of the variation, while season (proxied by temperature) only explained 6% of the variation in PLFA composition. Bacterial and fungal biomass were both highest in early spring, while AM fungal biomass peaked in early summer. Bacterial growth rate, on the other hand, was highest during the autumn, while fungal growth rate showed no clear seasonal pattern. In conclusion, the influence of seasonal variation on microbial communities proved to be relatively small compared to that which could be assigned to pH and NH4 in the studied ranges.

Daily temporal structure in African savanna flower visitation networks and consequences for network sampling

Ecological interaction networks are a valuable approach to understanding plant-pollinator interactions at the community level. Highly structured daily activity patterns are a feature of the biology of many flower visitors, particularly provisioning female bees, which often visit different floral sources at different times. Such temporal structure implies that presence/absence and relative abundance of specific flower-visitor interactions (links) in interaction networks may be highly sensitive to the daily timing of data collection. Further, relative timing of interactions is central to their possible role in competition or facilitation of seed set among coflowering plants sharing pollinators. To date, however, no study has examined the network impacts of daily temporal variation in visitor activity at a community scale. Here we use temporally structured sampling to examine the consequences of daily activity patterns upon network properties using fully quantified flower-visitor interaction data for a Kenyan savanna habitat. Interactions were sampled at four sequential three-hour time intervals between 06:00 and 18:00, across multiple seasonal time points for two sampling sites. In all data sets the richness and relative abundance of links depended critically on when during the day visitation was observed. Permutation-based null modeling revealed significant temporal structure across daily time intervals at three of the four seasonal time points, driven primarily by patterns in bee activity. This sensitivity of network structure shows the need to consider daily time in network sampling design, both to maximize the probability of sampling links relevant to plant reproductive success and to facilitate appropriate interpretation of interspecific relationships. Our data also suggest that daily structuring at a community level could reduce indirect competitive interactions when coflowering plants share pollinators, as is commonly observed during flowering in highly seasonal habitats.