Vegetation dynamics and rainfall sensitivity of the Amazon

We show that the vegetation canopy of the Amazon rainforest is highly sensitive to changes in precipitation patterns and that reduction in rainfall since 2000 has diminished vegetation greenness across large parts of Amazonia. Large-scale directional declines in vegetation greenness may indicate decreases in carbon uptake and substantial changes in the energy balance of the Amazon. We use improved estimates of surface reflectance from satellite data to show a close link between reductions in annual precipitation, El Niño southern oscillation events, and photosynthetic activity across tropical and subtropical Amazonia. We report that, since the year 2000, precipitation has declined across 69% of the tropical evergreen forest (5.4 million km²) and across 80% of the subtropical grasslands (3.3 million km²). These reductions, which coincided with a decline in terrestrial water storage, account for about 55% of a satellite-observed widespread decline in the normalized difference vegetation index (NDVI). During El Niño events, NDVI was reduced about 16.6% across an area of up to 1.6 million km² compared with average conditions. Several global circulation models suggest that a rise in equatorial sea surface temperature and related displacement of the intertropical convergence zone could lead to considerable drying of tropical forests in the 21st century. Our results provide evidence that persistent drying could degrade Amazonian forest canopies, which would have cascading effects on global carbon and climate dynamics.Rise in equatorial sea surface temperature has led to concerns that intensified El Niño southern oscillation (ENSO) events and a displacement of the intertropical convergence zone (1) could alter precipitation patterns in Amazonia (2, 3), resulting in increased length of the dry season (4) and more frequent severe droughts (5, 6). The feedbacks of such drying on global climate change could be substantial; the Amazon rainforest stores an estimated 120 billion tons of carbon (7, 8). Loss of forest productivity across Amazonia would clearly exacerbate atmospheric CO₂ levels (9, 10); however, the extent to which drying affects terrestrial vegetation is currently unknown (11). Satellite remote sensing is the only practical way to observe the potential impacts that these changes may have on vegetation at useful spatial and temporal scales (12), but in recent years, conflicting results have been reported of whether productivity of tropical forests is limited by sunlight or precipitation (7, 11, 13–16). Several studies have indicated that gross primary productivity increases initially during drought as a result of an increase in photosynthetically active radiation (PAR) (17, 18), but sensitivity to prolonged drought events and thresholds of forest dieback remain unclear. For instance, as a result of the severe Amazon drought in 2005, Phillips et al. (8) estimated an accumulated carbon loss of 1.2–1.6 petagram (Pg) based on records from 55 long-term monitoring plots. In contrast, Saleska et al. (13) reported greening of the Amazon forest based on remotely sensed estimates of the enhanced vegetation index acquired from the Moderate Resolution Imaging Spectroradiometer (MODIS) from the National Aeronautics and Space Administration (NASA). Saleska et al. (13) concluded that tropical forests were more drought resistant than previously thought and remained a strong carbon sink even during drought. However, these assertions were subsequently questioned (7, 11), and after a second drought in 2010, Xu et al. (19) documented widespread decline in tropical vegetation.Similar to interannual changes related to drought, there have also been contradictory findings related to seasonal changes between dry and wet seasons. A substantial body of literature (15, 17, 18, 20–22) supports the view that photosynthetic activity increases during the dry season in response to an increase in incident PAR, whereas water supply is maintained through deep root systems of tropical forests (23). In contrast, Morton et al. (14) argued that MODIS-derived observations of seasonal greening of tropical vegetation are an artifact of the sun-sensor geometry, concluding that tropical forests maintain consistent greenness throughout the dry and wet seasons.Resolving the discussion about drought tolerance of tropical vegetation is critical to reduce uncertainties in carbon balance models (16, 24, 25) and establish possible thresholds beyond which forest dieback may occur (15). Recent work suggests a substantial uncertainty of MODIS surface reflectance across the Amazon basin as a likely cause of these discrepancies in interpretation (26–28). Surface reflectance is routinely derived from top of atmosphere measurements using pixel-based atmospheric correction and cloud screening (29). Poor estimation of atmospheric aerosol loadings (11, 26) and deficiencies in cloud screening (30) can, therefore, introduce errors in vegetation indices (27). We take advantage of a new multiangle implementation of atmospheric correction algorithm (MAIAC) (31) to refine the analysis of the sensitivity of tropical and subtropical vegetation to variation in precipitation using daily observations of MODIS surface reflectance acquired between 2000 and 2012. MAIAC improves the accuracy of satellite-based surface reflectance over tropical vegetation by 3- to 10-fold compared with current MODIS products (30). This improvement is accomplished, in part, through a more accurate and less conservative cloud mask, which increases the number of clear-sky observations by a factor of 2–5 compared with standard procedures (30). A higher number of clear-sky observations is particularly important for analysis of tropical regions, where average cloud cover may be as high as 70% during the dry season and 95–99% during the wet season (30). This improvement along with the removal of calibration errors in the latest MODIS Terra Collection 6 (C6) data provide us with more confidence in interpreting the state and changes in the Amazon forests.     

From the Cover: Thermal-safety margins and the necessity of thermoregulatory behavior across latitude and elevation

Physiological thermal-tolerance limits of terrestrial ectotherms often exceed local air temperatures, implying a high degree of thermal safety (an excess of warm or cold thermal tolerance). However, air temperatures can be very different from the equilibrium body temperature of an individual ectotherm. Here, we compile thermal-tolerance limits of ectotherms across a wide range of latitudes and elevations and compare these thermal limits both to air and to operative body temperatures (theoretically equilibrated body temperatures) of small ectothermic animals during the warmest and coldest times of the year. We show that extreme operative body temperatures in exposed habitats match or exceed the physiological thermal limits of most ectotherms. Therefore, contrary to previous findings using air temperatures, most ectotherms do not have a physiological thermal-safety margin. They must therefore rely on behavior to avoid overheating during the warmest times, especially in the lowland tropics. Likewise, species living at temperate latitudes and in alpine habitats must retreat to avoid lethal cold exposure. Behavioral plasticity of habitat use and the energetic consequences of thermal retreats are therefore critical aspects of species’ vulnerability to climate warming and extreme events.Predicting the organismal responses to climate change—a global priority—requires an understanding of the physiological, behavioral, ecological, and evolutionary factors that constrain where species can live (1, 2). Macrophysiological analyses that predict large-scale patterns in the vulnerability of ectotherms to climate warming often invoke the concept of the “thermal-safety margin” (3–6), which measures the difference between a species’ maximum tolerance to heat and the warm air temperatures it regularly experiences. Such heat safety margins often increase markedly with latitude, implying that tropical species might be relatively more vulnerable to climate warming than are species living at higher latitudes (refs. 3–6; but see ref. 7), even though the rate of climate warming is lower in the tropics (8). Indeed, many temperate ectotherms appear to have maximum thermal tolerances that are 10–20 °C higher than required to withstand the average summer air temperatures where they live (3).Comparative physiology offers three reasons to be skeptical about such high thermal-safety margins. First, to index environmental temperatures, prior studies often used mean annual or seasonal air temperatures—measures that may have little ecological relevance in more variable and seasonal environments (5, 9, 10). Indeed, rare extreme temperatures—not average ones—may be more important for long-term species persistence (11, 12).Second, studies generally use air temperatures (T_a, taken in shade at 1- to 2-m height) to index thermal environments: These temperatures are readily available but poorly characterize the thermal environment from an ectotherm''s perspective. An ectotherm''s body temperature can differ strikingly from local T_a because heat exchange is affected not merely by convection, but also by radiation, conduction, evaporation, and metabolism (7, 13–15). For example, an Andean lizard basking at 4,450 m had a body temperature of 31 °C even though air temperature was only ∼0 °C (16). Operative temperatures (T_e), which estimate an ectotherm''s steady-state body temperature, are more biophysically accurate indices of microclimates experienced by ectotherms. T_e can be estimated either with physical models placed in the environment (14) or by mathematical models (17, 18) (Methods).Third, most terrestrial ectotherms are mobile and can behaviorally exploit local heterogeneity in T_e to regulate body temperatures somewhat independently of local environmental temperatures (“Bogert effect”) (19). For example, merely by shifting time (e.g., day/night) or place of activity (e.g., open habitat, shade, or burrows), many ectotherms can have a body temperature that is markedly different from air temperature (20, 21). In addition, wet-skinned ectotherms such as amphibians cool their bodies evaporatively and thus have a lower T_e than otherwise-comparable dry-skinned ectotherms (13).Because T_a and T_e are thus fundamentally different metrics of environmental heat loads (with T_e varying according to microhabitat), thermal-safety margins based on these alternative metrics must also differ. Here, we estimate global patterns of thermal-safety margins based on maximum and minimum T_e, instead of T_a.Global patterns in thermal-safety margins are useful for understanding not only species’ vulnerabilities to climate warming, but also the historical role of physiology and behavior in protecting species from temperature extremes; we therefore also consider cold thermal-safety margins, which represent the offset between cold temperature tolerance and minimum T_e. We also explore variation in thermal tolerance and thermal-safety margins with elevation, to compare patterns along elevational and latitudinal gradients.We start by expanding a global, empirical dataset on physiological heat and cold tolerance limits in amphibians, reptiles, and insects from diverse latitudes and elevations (22). We next use a global climate database and a biophysical model to calculate maximum and minimum air (T_a) and operative temperatures (T_e, as described in ref. 21) at the location of collection for both the dry and the wet-skinned ectotherms for which we have thermal-tolerance estimates. We then calculate and compare thermal-safety margins based on maximum and minimum T_e for individual species versus “traditional” margins based on T_a (3–5) and evaluate patterns across latitude and elevation. We find that safety margins based on T_e are much more likely to be negative than those based on T_a. This result implies that terrestrial ectotherms do not have sufficient physiological tolerance to protect them from dangerously extreme operative temperatures. Consequently, terrestrial ectotherms in almost all localities must rely on behavioral adjustments to survive the warmest times of year (17, 23–25). Finally, we estimate T_e in a range of microhabitats (e.g., full sun, shade, burrows) and show that behavioral shifts in microhabitat use can provide the refugia necessary (24).Our findings force a reevaluation of latitudinal and elevational patterns of thermal danger, revealing that exposure to extreme heat can occur even at high elevations and latitudes (7) and giving insight into why heat-tolerance limits are relatively invariant in comparison with cold limits (22, 26, 27). Moreover, we uncover taxon-specific patterns in biophysically based thermal-safety margins and behavioral options necessary and sufficient to evade dangerous thermal environments.     

N terminus of ASPP2 binds to Ras and enhances Ras/Raf/MEK/ERK activation to promote oncogene-induced senescence

The ASPP2 (also known as 53BP2L) tumor suppressor is a proapoptotic member of a family of p53 binding proteins that functions in part by enhancing p53-dependent apoptosis via its C-terminal p53-binding domain. Mounting evidence also suggests that ASPP2 harbors important nonapoptotic p53-independent functions. Structural studies identify a small G protein Ras-association domain in the ASPP2 N terminus. Because Ras-induced senescence is a barrier to tumor formation in normal cells, we investigated whether ASPP2 could bind Ras and stimulate the protein kinase Raf/MEK/ERK signaling cascade. We now show that ASPP2 binds to Ras–GTP at the plasma membrane and stimulates Ras-induced signaling and pERK1/2 levels via promoting Ras–GTP loading, B-Raf/C-Raf dimerization, and C-Raf phosphorylation. These functions require the ASPP2 N terminus because BBP (also known as 53BP2S), an alternatively spliced ASPP2 isoform lacking the N terminus, was defective in binding Ras–GTP and stimulating Raf/MEK/ERK signaling. Decreased ASPP2 levels attenuated H-RasV12–induced senescence in normal human fibroblasts and neonatal human epidermal keratinocytes. Together, our results reveal a mechanism for ASPP2 tumor suppressor function via direct interaction with Ras–GTP to stimulate Ras-induced senescence in nontransformed human cells.ASPP2, also known as 53BP2L, is a tumor suppressor whose expression is altered in human cancers (1). Importantly, targeting of the ASPP2 allele in two different mouse models reveals that ASPP2 heterozygous mice are prone to spontaneous and γ-irradiation–induced tumors, which rigorously demonstrates the role of ASPP2 as a tumor suppressor (2, 3). ASPP2 binds p53 via the C-terminal ankyrin-repeat and SH3 domain (4–6), is damage-inducible, and can enhance damage-induced apoptosis in part through a p53-mediated pathway (1, 2, 7–10). However, it remains unclear what biologic pathways and mechanisms mediate ASPP2 tumor suppressor function (1). Indeed, accumulating evidence demonstrates that ASPP2 also mediates nonapoptotic p53-independent pathways (1, 3, 11–15).The induction of cellular senescence forms an important barrier to tumorigenesis in vivo (16–21). It is well known that oncogenic Ras signaling induces senescence in normal nontransformed cells to prevent tumor initiation and maintain complex growth arrest pathways (16, 18, 21–24). The level of oncogenic Ras activation influences its capacity to activate senescence; high levels of oncogenic H-RasV12 signaling leads to low grade tumors with senescence markers, which progress to invasive cancers upon senescence inactivation (25). Thus, tight control of Ras signaling is critical to ensure the proper biologic outcome in the correct cellular context (26–28).The ASPP2 C terminus is important for promoting p53-dependent apoptosis (7). The ASPP2 N terminus may also suppress cell growth (1, 7, 29–33). Alternative splicing can generate the ASPP2 N-terminal truncated protein BBP (also known as 53BP2S) that is less potent in suppressing cell growth (7, 34, 35). Although the ASPP2 C terminus mediates nuclear localization, full-length ASPP2 also localizes to the cytoplasm and plasma membrane to mediate extranuclear functions (7, 11, 12, 36). Structural studies of the ASPP2 N terminus reveal a β–Grasp ubiquitin-like fold as well as a potential Ras-binding (RB)/Ras-association (RA) domain (32). Moreover, ASPP2 can promote H-RasV12–induced senescence (13, 15). However, the molecular mechanism(s) of how ASPP2 directly promotes Ras signaling are complex and remain to be completely elucidated.Here, we explore the molecular mechanisms of how Ras-signaling is enhanced by ASPP2. We demonstrate that ASPP2: (i) binds Ras-GTP and stimulates Ras-induced ERK signaling via its N-terminal domain at the plasma membrane; (ii) enhances Ras-GTP loading and B-Raf/C-Raf dimerization and forms a ASPP2/Raf complex; (iii) stimulates Ras-induced C-Raf phosphorylation and activation; and (iv) potentiates H-RasV12–induced senescence in both primary human fibroblasts and neonatal human epidermal keratinocytes. These data provide mechanistic insight into ASPP2 function(s) and opens important avenues for investigation into its role as a tumor suppressor in human cancer.     

Self-assembly of alkynylplatinum(II) terpyridine amphiphiles into nanostructures via steric control and metal–metal interactions

A series of mono- and dinuclear alkynylplatinum(II) terpyridine complexes containing the hydrophilic oligo(para-phenylene ethynylene) with two 3,6,9-trioxadec-1-yloxy chains was designed and synthesized. The mononuclear alkynylplatinum(II) terpyridine complex was found to display a very strong tendency toward the formation of supramolecular structures. Interestingly, additional end-capping with another platinum(II) terpyridine moiety of various steric bulk at the terminal alkyne would lead to the formation of nanotubes or helical ribbons. These desirable nanostructures were found to be governed by the steric bulk on the platinum(II) terpyridine moieties, which modulates the directional metal−metal interactions and controls the formation of nanotubes or helical ribbons. Detailed analysis of temperature-dependent UV-visible absorption spectra of the nanostructured tubular aggregates also provided insights into the assembly mechanism and showed the role of metal−metal interactions in the cooperative supramolecular polymerization of the amphiphilic platinum(II) complexes.Square-planar d⁸ platinum(II) polypyridine complexes have long been known to exhibit intriguing spectroscopic and luminescence properties (1–54) as well as interesting solid-state polymorphism associated with metal−metal and π−π stacking interactions (1–14, 25). Earlier work by our group showed the first example, to our knowledge, of an alkynylplatinum(II) terpyridine system [Pt(tpy)(C ≡ CR)]⁺ that incorporates σ-donating and solubilizing alkynyl ligands together with the formation of Pt···Pt interactions to exhibit notable color changes and luminescence enhancements on solvent composition change (25) and polyelectrolyte addition (26). This approach has provided access to the alkynylplatinum(II) terpyridine and other related cyclometalated platinum(II) complexes, with functionalities that can self-assemble into metallogels (27–31), liquid crystals (32, 33), and other different molecular architectures, such as hairpin conformation (34), helices (35–38), nanostructures (39–45), and molecular tweezers (46, 47), as well as having a wide range of applications in molecular recognition (48–52), biomolecular labeling (48–52), and materials science (53, 54). Recently, metal-containing amphiphiles have also emerged as a building block for supramolecular architectures (42–44, 55–59). Their self-assembly has always been found to yield different molecular architectures with unprecedented complexity through the multiple noncovalent interactions on the introduction of external stimuli (42–44, 55–59).Helical architecture is one of the most exciting self-assembled morphologies because of the uniqueness for the functional and topological properties (60–69). Helical ribbons composed of amphiphiles, such as diacetylenic lipids, glutamates, and peptide-based amphiphiles, are often precursors for the growth of tubular structures on an increase in the width or the merging of the edges of ribbons (64, 65). Recently, the optimization of nanotube formation vs. helical nanostructures has aroused considerable interests and can be achieved through a fine interplay of the influence on the amphiphilic property of molecules (66), choice of counteranions (67, 68), or pH values of the media (69), which would govern the self-assembly of molecules into desirable aggregates of helical ribbons or nanotube scaffolds. However, a precise control of supramolecular morphology between helical ribbons and nanotubes remains challenging, particularly for the polycyclic aromatics in the field of molecular assembly (64–69). Oligo(para-phenylene ethynylene)s (OPEs) with solely π−π stacking interactions are well-recognized to self-assemble into supramolecular system of various nanostructures but rarely result in the formation of tubular scaffolds (70–73). In view of the rich photophysical properties of square-planar d⁸ platinum(II) systems and their propensity toward formation of directional Pt···Pt interactions in distinctive morphologies (27–31, 39–45), it is anticipated that such directional and noncovalent metal−metal interactions might be capable of directing or dictating molecular ordering and alignment to give desirable nanostructures of helical ribbons or nanotubes in a precise and controllable manner.Herein, we report the design and synthesis of mono- and dinuclear alkynylplatinum(II) terpyridine complexes containing hydrophilic OPEs with two 3,6,9-trioxadec-1-yloxy chains. The mononuclear alkynylplatinum(II) terpyridine complex with amphiphilic property is found to show a strong tendency toward the formation of supramolecular structures on diffusion of diethyl ether in dichloromethane or dimethyl sulfoxide (DMSO) solution. Interestingly, additional end-capping with another platinum(II) terpyridine moiety of various steric bulk at the terminal alkyne would result in nanotubes or helical ribbons in the self-assembly process. To the best of our knowledge, this finding represents the first example of the utilization of the steric bulk of the moieties, which modulates the formation of directional metal−metal interactions to precisely control the formation of nanotubes or helical ribbons in the self-assembly process. Application of the nucleation–elongation model into this assembly process by UV-visible (UV-vis) absorption spectroscopic studies has elucidated the nature of the molecular self-assembly, and more importantly, it has revealed the role of metal−metal interactions in the formation of these two types of nanostructures.     

Exchange protein directly activated by cAMP plays a critical role in bacterial invasion during fatal rickettsioses

Rickettsiae are responsible for some of the most devastating human infections. A high infectivity and severe illness after inhalation make some rickettsiae bioterrorism threats. We report that deletion of the exchange protein directly activated by cAMP (Epac) gene, Epac1, in mice protects them from an ordinarily lethal dose of rickettsiae. Inhibition of Epac1 suppresses bacterial adhesion and invasion. Most importantly, pharmacological inhibition of Epac1 in vivo using an Epac-specific small-molecule inhibitor, ESI-09, completely recapitulates the Epac1 knockout phenotype. ESI-09 treatment dramatically decreases the morbidity and mortality associated with fatal spotted fever rickettsiosis. Our results demonstrate that Epac1-mediated signaling represents a mechanism for host–pathogen interactions and that Epac1 is a potential target for the prevention and treatment of fatal rickettsioses.Rickettsiae are responsible for some of the most devastating human infections (1–4). It has been forecasted that temperature increases attributable to global climate change will lead to more widespread distribution of rickettsioses (5). These tick-borne diseases are caused by obligately intracellular bacteria of the genus Rickettsia, including Rickettsia rickettsii, the causative agent of Rocky Mountain spotted fever (RMSF) in the United States and Latin America (2, 3), and Rickettsia conorii, the causative agent of Mediterranean spotted fever endemic to southern Europe, North Africa, and India (6). A high infectivity and severe illness after inhalation make some rickettsiae (including Rickettsia prowazekii, R. rickettsii, Rickettsia typhi, and R. conorii) bioterrorism threats (7). Although the majority of rickettsial infections can be controlled by appropriate broad-spectrum antibiotic therapy if diagnosed early, up to 20% of misdiagnosed or untreated (1, 3) and 5% of treated RMSF cases (8) result in a fatal outcome caused by acute disseminated vascular endothelial infection and damage (9). Fatality rates as high as 32% have been reported in hospitalized patients diagnosed with Mediterranean spotted fever (10). In addition, strains of R. prowazekii resistant to tetracycline and chloramphenicol have been developed in laboratories (11). Disseminated endothelial infection and endothelial barrier disruption with increased microvascular permeability are the central features of SFG rickettsioses (1, 2, 9). The molecular mechanisms involved in rickettsial infection remain incompletely elucidated (9, 12). A comprehensive understanding of rickettsial pathogenesis and the development of novel mechanism-based treatment are urgently needed.Living organisms use intricate signaling networks for sensing and responding to changes in the external environment. cAMP, a ubiquitous second messenger, is an important molecular switch that translates environmental signals into regulatory effects in cells (13). As such, a number of microbial pathogens have evolved a set of diverse virulence-enhancing strategies that exploit the cAMP-signaling pathways of their hosts (14). The intracellular functions of cAMP are predominantly mediated by the classic cAMP receptor, protein kinase A (PKA), and the more recently discovered exchange protein directly activated by cAMP (Epac) (15). Thus, far, two isoforms, Epac1 and Epac2, have been identified in humans (16, 17). Epac proteins function by responding to increased intracellular cAMP levels and activating the Ras superfamily small GTPases Ras-proximate 1 and 2 (Rap1 and Rap2). Accumulating evidence demonstrates that the cAMP/Epac1 signaling axis plays key regulatory roles in controlling various cellular functions in endothelial cells in vitro, including cell adhesion (18–21), exocytosis (22), tissue plasminogen activator expression (23), suppressor of cytokine signaling 3 (SOCS-3) induction (24–27), microtubule dynamics (28, 29), cell–cell junctions, and permeability and barrier functions (30–37). Considering the critical importance of endothelial cells in rickettsioses, we examined the functional roles of Epac1 in rickettsial pathogenesis in vivo, taking advantage of the recently generated Epac1 knockout mouse (38) and Epac-specific inhibitors (39, 40) generated from our laboratory. Our studies demonstrate that Epac1 plays a key role in rickettsial infection and represents a therapeutic target for fatal rickettsioses.     

Rapid upslope shifts in New Guinean birds illustrate strong distributional responses of tropical montane species to global warming

Temperate-zone species have responded to warming temperatures by shifting their distributions poleward and upslope. Thermal tolerance data suggests that tropical species may respond to warming temperatures even more strongly than temperate-zone species, but this prediction has yet to be tested. We addressed this data gap by conducting resurveys to measure distributional responses to temperature increases in the elevational limits of the avifaunas of two geographically and faunally independent New Guinean mountains, Mt. Karimui and Karkar Island, 47 and 44 y after they were originally surveyed. Although species richness is roughly five times greater on mainland Mt. Karimui than oceanic Karkar Island, distributional shifts at both sites were similar: upslope shifts averaged 113 m (Mt. Karimui) and 152 m (Karkar Island) for upper limits and 95 m (Mt. Karimui) and 123 m (Karkar Island) for lower limits. We incorporated these results into a metaanalysis to compare distributional responses of tropical species with those of temperate-zone species, finding that average upslope shifts in tropical montane species match local temperature increases significantly more closely than in temperate-zone montane species. That tropical species appear to be strong responders has global conservation implications and provides empirical support to hitherto untested models that predict widespread extinctions in upper-elevation tropical endemics with small ranges.Temperate species are responding to anthropogenic temperature increases by rapidly shifting geographic distributions to track their climatic niche (1–3). These shifts appear to be increasing in pace—a recent metaanalysis concluded that species are shifting their distributions poleward and upslope much faster than previously estimated (1, 2). Range shifts are less studied in tropical regions however (1, 4, 5), despite being home to the vast majority of biodiversity (6). Notwithstanding strong latitudinal bias in empirical studies, climate change-driven range shifts are predicted to cause widespread extinctions in both temperate and tropical species within the next century (7–10).With scarce empirical data, models of tropical species’ response to temperature increases predict a wide range of responses (11). At one extreme, tropical species may be relatively unaffected, as the magnitude of temperature increases is relatively low in the tropics (12). Alternately, vulnerability to warming temperatures could be highest in the tropics if tropical species are physiologically specialized to narrow thermal niches (13–18). Such thermal specialization has been documented in tropical ectotherms (16, 17), but it is unclear whether similar patterns may apply to tropical endotherms, whose distributional shifts in response to warming may result from indirect rather than direct impacts of temperature increases (5).We resurveyed geographically and faunally independent elevational gradients in New Guinea nearly a half-century after they were first surveyed. The original transect surveys were conducted by J. Diamond to determine bird species’ elevational limits on Mt. Karimui (July–August 1965) (19) and Karkar Island (May 1969) (20). These environments differ significantly: Mt. Karimui is located in New Guinea’s biodiverse Central Ranges and harbors a diverse resident avifauna of ca. 250 resident landbirds (19), whereas Karkar Island is a small oceanic island off New Guinea’s north coast with a depauperate flora and fauna (ca. 50 resident landbirds) dominated by highly dispersive taxa (20) (Fig. 1).Open in a separate windowFig. 1.Map of resurvey sites in Papua New Guinea. The elevational transects recently revisited by the authors are marked by dashed lines (Mt. Karimui: 1,130–2,520 m; Karkar Island: 800–1,600 m). Mt. Karimui is an extinct volcano in the southern Central Ranges of New Guinea, whereas Karkar Island is an oceanic island located 10 miles from the New Guinean mainland. These elevational gradients were originally surveyed by Diamond in the 1960s [Mt. Karimui: 1965 (19); Karkar Island: 1969 (20)], and remain covered in primary forest.We used elevational limits measured during historical transects and modern resurveys to investigate New Guinean montane birds’ response to warming temperatures. We predicted that species have moved upslope relative to historical range limits. Given that tropical species are hypothesized to be especially sensitive to temperature increases (either directly or via indirect ecological interactions), we additionally predicted that the magnitude of upslope shifts would closely match predicted shifts based on local temperature increases. We simultaneously tested two additional hypotheses, investigating whether upslope shifts at the leading range margin outpaced upslope shifts at the trailing range edge (21), and whether species’ dietary preferences influenced upslope shifts (22, 23). We then used our data in conjunction with recent tropical resurveys to test the tropical-species-are-strong-responders hypothesis, predicting that upslope shifts measured in tropical resurveys match predicted upslope shifts significantly more closely than for temperate-zone resurveys.     

Joint control of terrestrial gross primary productivity by plant phenology and physiology

Terrestrial gross primary productivity (GPP) varies greatly over time and space. A better understanding of this variability is necessary for more accurate predictions of the future climate–carbon cycle feedback. Recent studies have suggested that variability in GPP is driven by a broad range of biotic and abiotic factors operating mainly through changes in vegetation phenology and physiological processes. However, it is still unclear how plant phenology and physiology can be integrated to explain the spatiotemporal variability of terrestrial GPP. Based on analyses of eddy–covariance and satellite-derived data, we decomposed annual terrestrial GPP into the length of the CO₂ uptake period (CUP) and the seasonal maximal capacity of CO₂ uptake (GPP_max). The product of CUP and GPP_max explained >90% of the temporal GPP variability in most areas of North America during 2000–2010 and the spatial GPP variation among globally distributed eddy flux tower sites. It also explained GPP response to the European heatwave in 2003 (r² = 0.90) and GPP recovery after a fire disturbance in South Dakota (r² = 0.88). Additional analysis of the eddy–covariance flux data shows that the interbiome variation in annual GPP is better explained by that in GPP_max than CUP. These findings indicate that terrestrial GPP is jointly controlled by ecosystem-level plant phenology and photosynthetic capacity, and greater understanding of GPP_max and CUP responses to environmental and biological variations will, thus, improve predictions of GPP over time and space.Large variability exists among estimates of terrestrial carbon sequestration, resulting in substantial uncertainty in modeled dynamics of atmospheric CO₂ concentration and predicted future climate change (1). The variability in carbon sequestration is partially caused by variation in terrestrial gross primary productivity (GPP) (2), which is the cumulative rate over time of gross plant photosynthesis at the ecosystem level. Plant photosynthesis has been successfully modeled at the biochemical level (3, 4). When leaf-level biochemical models of photosynthesis are scaled up to estimate annual GPP over a region and the globe, however, great uncertainty arises from both vegetation properties, such as biome-dependent leaf parameters (5, 6), and environmental factors, such as climate variability (7–9) and episodic disturbances (10–12). As a consequence, estimated present day global GPP varies from 105 to 177 Pg C y⁻¹ in the fifth phase of the Coupled Model Intercomparison Project (13). Additionally, spatiotemporal patterns of GPP (2, 14), their responses to extreme climate events (12) and disturbances (10), and the underlying mechanisms are still not well-understood. Previous studies have indicated that vegetation properties and environmental factors shape annual GPP of an ecosystem directly or indirectly through affecting plant physiological activities (15) and/or phenology (16–21). Thus, integrating plant physiological and phenological properties may provide a unified approach to explain the variability of GPP over time and space and in response to disturbance.In this study, we show that annual GPP in grams C meter⁻² year⁻¹, the rate at which terrestrial ecosystems take up CO₂ from the atmosphere in a given year, can be quantitatively decomposed intoGPP = α?CUP?GPP_max, [1]where the carbon dioxide uptake period (CUP; number of days per year) is a phenological indicator of the duration of ecosystem CO₂ assimilation within a given year. GPP_max (grams C meter⁻² day⁻¹) is the maximal daily rate of gross photosynthesis during the CUP and represents a property of plant canopy physiology. The ratio between annual GPP and the product of CUP and GPP_max is represented by α. We estimated α, CUP, and GPP_max for 213 globally distributed terrestrial sites with daily GPP from the global network of micrometeorological tower sites (FLUXNET; La Thuile Database) (22) (SI Appendix, section S1.1.1 and Table S1) and all 0.1° × 0.1° land grid cells in North America during 2000–2010 with an 8-d GPP product from the Moderate Resolution Imaging Spectroradiometer (MODIS) onboard the National Aeronautics and Space Administration Terra satellite (23) (Materials and Methods). Here, we show how CUP and GPP_max jointly control the spatiotemporal variability of GPP and its response to and recovery from disturbances in different terrestrial ecosystems.     

CACTA-like transposable element in ZmCCT attenuated photoperiod sensitivity and accelerated the postdomestication spread of maize

The postdomestication adaptation of maize to longer days required reduced photoperiod sensitivity to optimize flowering time. We performed a genome-wide association study and confirmed that ZmCCT, encoding a CCT domain-containing protein, is associated with the photoperiod response. In early-flowering maize we detected a CACTA-like transposable element (TE) within the ZmCCT promoter that dramatically reduced flowering time. TE insertion likely occurred after domestication and was selected as maize adapted to temperate zones. This process resulted in a strong selective sweep within the TE-related block of linkage disequilibrium. Functional validations indicated that the TE represses ZmCCT expression to reduce photoperiod sensitivity, thus accelerating maize spread to long-day environments.Maize (Zea mays L.) was domesticated in Southern Mexico roughly 9,000 y ago from Balsas teosinte (Zea mays ssp. parviglumis) (1), which requires short-day conditions to flower (2). Therefore the spread of maize from tropical to temperate regions required the postdomestication adaptation of maize to longer days (1, 3, 4). As such, temperate maize is largely day-length insensitive, whereas tropical maize lines are generally sensitive to longer day lengths.To modulate the timing of flowering, plants integrate signals from the environment and from endogenous regulatory pathways (5). Most genes known to regulate maize flowering (6–12) are part of the autonomous pathway, such as id1 (6, 7), ZCN8 (8), dlf1 (9), zfl1 (10), conz1 (11), and Vgt1 (12). Flowering time in maize is extremely variable (ranging from 35–120 d) (13) and is controlled primarily by a large number of quantitative trait loci (QTLs), each with a small effect (14). Relatively few of these flowering-time QTLs affect the photoperiod response, although ZmCCT, encoding a CCT domain-containing protein, appears to be the most important locus in these contexts (15–18). As such, molecular details concerning the photoperiodic control of maize flowering remain unclear.Transposable elements (TEs) played a key role in adaptive plant evolution and phenotypic variation by altering gene expression and function (19–23). In fact, TEs often served as targets of selection during evolution (24). Insertion of the Rider retrotransposon into the tomato genome increased expression of the gene SUN, which led to an elongated fruit shape (25). Similarly, insertion of a miniature inverted-repeat TE (MITE) into Vgt1, which is a cis-regulatory element located ∼70 kb upstream of the flowering-time repressor ZmRap2.7, is tightly associated with flowering-time variation in maize (12). Finally, insertion of a Hopscotch retrotransposon upstream of the maize-domestication gene tb1 increased apical dominance in maize (26, 27).Here we performed a genome-wide association study (GWAS) using a diverse panel of maize lines (28, 29) to identify genetic variants near ZmCCT that associate with flowering time. Using an overlapping PCR approach, we detected a CACTA-like TE within the ZmCCT regulatory region. Genetic effects of this TE on flowering time were investigated by ZmCCT-based association mapping and biparental linkage analysis. The CACTA-like TE appeared to be a causative factor in reducing photoperiod sensitivity under long-day conditions and was the target of a strong selective sweep during the postdomestication spread of maize. Functional validations demonstrate that ZmCCT is involved in the photoperiod response and that the CACTA-like TE within ZmCCT represses gene expression, rendering maize insensitive to long days.     

Reconstitution of long and short patch mismatch repair reactions using Saccharomyces cerevisiae proteins

A problem in understanding eukaryotic DNA mismatch repair (MMR) mechanisms is linking insights into MMR mechanisms from genetics and cell-biology studies with those from biochemical studies of MMR proteins and reconstituted MMR reactions. This type of analysis has proven difficult because reconstitution approaches have been most successful for human MMR whereas analysis of MMR in vivo has been most advanced in the yeast Saccharomyces cerevisiae. Here, we describe the reconstitution of MMR reactions using purified S. cerevisiae proteins and mispair-containing DNA substrates. A mixture of MutS homolog 2 (Msh2)–MutS homolog 6, Exonuclease 1, replication protein A, replication factor C-Δ1N, proliferating cell nuclear antigen and DNA polymerase δ was found to repair substrates containing TG, CC, +1 (+T), +2 (+GC), and +4 (+ACGA) mispairs and either a 5′ or 3′ strand interruption with different efficiencies. The Msh2–MutS homolog 3 mispair recognition protein could substitute for the Msh2–Msh6 mispair recognition protein and showed a different specificity of repair of the different mispairs whereas addition of MutL homolog 1–postmeiotic segregation 1 had no affect on MMR. Repair was catalytic, with as many as 11 substrates repaired per molecule of Exo1. Repair of the substrates containing either a 5′ or 3′ strand interruption occurred by mispair binding-dependent 5′ excision and subsequent resynthesis with excision tracts of up to ∼2.9 kb occurring during the repair of the substrate with a 3′ strand interruption. The availability of this reconstituted MMR reaction now makes possible detailed biochemical studies of the wealth of mutations identified that affect S. cerevisiae MMR.DNA mismatch repair (MMR) is a critical DNA repair pathway that is coupled to DNA replication in eukaryotes where it corrects misincorporation errors made during DNA replication (1–9). This pathway prevents mutations and acts to prevent the development of cancer (10, 11). MMR also contributes to gene conversion by repairing mispaired bases that occur during the formation of recombination intermediates (3, 4, 12). Finally, MMR acts to suppress recombination between divergent but homologous DNA sequences, thereby preventing the formation of genome rearrangements that can result from nonallelic homologous recombination (4, 13–15).Our knowledge of the mechanism of eukaryotic MMR comes from several general lines of investigation (3–9). Studies of bacterial MMR have provided a basic mechanistic framework for comparative studies (5). Genetic and cell-biology studies, primarily in Saccharomyces cerevisiae, have identified eukaryotic MMR genes, provided models for how their gene products define MMR pathways, and elucidated some of the details of how MMR pathways interact with replication (1–4). Reconstitution studies, primarily in human systems, have identified some of the catalytic features of eukaryotic MMR (7–9, 16, 17). Biochemical and structural studies of S. cerevisiae and human MMR proteins have provided information about the function of individual MMR proteins (6–9).In eukaryotic MMR, mispairs are bound by MutS homolog 2 (Msh2)–MutS homolog 6 (Msh6) and Msh2–MutS homolog 3 (Msh3), two partially redundant complexes of MutS-related proteins (3, 4, 18, 19). These complexes recruit a MutL-related complex, called MutL homoloh 1 (Mlh1)–postmeiotic segregation 1 (Pms1) in S. cerevisiae and Mlh1–postmeiotic segregation 2 (Pms2) in human and mouse (3, 4, 20–23). The Mlh1–Pms1/Pms2 complex has an endonuclease activity suggested to play a role in the initiation of the excision step of MMR (24, 25). Downstream of mismatch recognition is a mispair excision step that can be catalyzed by Exonuclease 1 (Exo1) (26–28); however, defects in both S. cerevisiae and mouse Exo1 result in only a partial MMR deficiency, suggesting the existence of additional excision mechanisms (26, 27, 29). DNA polymerase δ, the single-strand DNA binding protein replication protein A (RPA), the sliding clamp proliferating cell nuclear antigen (PCNA), and the clamp loader replication factor C (RFC) are also required for MMR at different steps, including activation of Mlh1–Pms1/Pms2, stimulation of Exo1, potentially in Exo1-independent mispair excision, and in the gap-filling resynthesis steps of MMR (3, 16, 17, 24, 27, 30–36). Although much is known about these core MMR proteins, it is not well understood how eukaryotic MMR is coupled to DNA replication (1, 2), how excision is targeted to the newly replicated strand (1, 25, 37–39), or how different MMR mechanisms such as Exo1-dependent and -independent subpathways are selected or how many such subpathways exist (1, 24, 27, 29).S. cerevisiae has provided a number of tools for studying MMR, including forward genetic screens for mutations affecting MMR, including dominant and separation-of-function mutations, the ability to evaluate structure-based mutations in vivo, cell biological tools for visualizing and analyzing MMR proteins in vivo, and overproduction of individual MMR proteins for biochemical analysis. However, linking these tools with biochemical systems that catalyze MMR reactions in vitro for mechanistic studies has not yet been possible. Here, we describe the development of MMR reactions reconstituted using purified proteins for the analysis of MMR mechanisms.     

Complex-type N-glycan recognition by potent broadly neutralizing HIV antibodies

Broadly neutralizing HIV antibodies (bNAbs) can recognize carbohydrate-dependent epitopes on gp120. In contrast to previously characterized glycan-dependent bNAbs that recognize high-mannose N-glycans, PGT121 binds complex-type N-glycans in glycan microarrays. We isolated the B-cell clone encoding PGT121, which segregates into PGT121-like and 10-1074–like groups distinguished by sequence, binding affinity, carbohydrate recognition, and neutralizing activity. Group 10-1074 exhibits remarkable potency and breadth but no detectable binding to protein-free glycans. Crystal structures of unliganded PGT121, 10-1074, and their likely germ-line precursor reveal that differential carbohydrate recognition maps to a cleft between complementarity determining region (CDR)H2 and CDRH3. This cleft was occupied by a complex-type N-glycan in a “liganded” PGT121 structure. Swapping glycan contact residues between PGT121 and 10-1074 confirmed their importance for neutralization. Although PGT121 binds complex-type N-glycans, PGT121 recognized high-mannose-only HIV envelopes in isolation and on virions. As HIV envelopes exhibit varying proportions of high-mannose- and complex-type N-glycans, these results suggest promiscuous carbohydrate interactions, an advantageous adaptation ensuring neutralization of all viruses within a given strain.Antibodies are essential for the success of most vaccines (1), and antibodies against HIV appear to be the only correlate of protection in the recent RV144 anti-HIV vaccine trial (2). Some HIV-1–infected patients develop broadly neutralizing serologic activity against the gp160 viral spike 2–4 y after infection (3–10), but these antibodies do not generally protect infected humans because autologous viruses escape through mutation (11–13). Nevertheless, broadly neutralizing activity puts selective pressure on the virus (13) and passive transfer of broadly neutralizing antibodies (bNAbs) to macaques protects against simian/human immunodeficiency virus (SHIV) infection (14–24). It has therefore been proposed that vaccines that elicit such antibodies may be protective against HIV infection in humans (10, 25–28).The development of single-cell antibody cloning techniques revealed that bNAbs target several different epitopes on the HIV-1 gp160 spike (29–35). The most potent HIV-1 bNAbs recognize the CD4 binding site (CD4bs) (31, 34, 36) and carbohydrate-dependent epitopes associated with the variable loops (32, 33, 37, 38), including the V1/V2 (antibodies PG9/PG16) (33) and V3 loops (PGTs) (32). Less is known about carbohydrate-dependent epitopes because the antibodies studied to date are either unique examples or members of small clonal families.To better understand the neutralizing antibody response to HIV-1 and the epitope targeted by PGT antibodies, we isolated members of a large clonal family dominating the gp160-specific IgG memory response from the clade A-infected patient who produced PGT121. We report that PGT121 antibodies segregate into two groups, a PGT121-like and a 10-1074–like group, according to sequence, binding affinity, neutralizing activity, and recognition of carbohydrates and the V3 loop. The 10-1074 antibody and related family members exhibit unusual potent neutralization, including broad reactivity against newly transmitted viruses. Unlike previously characterized carbohydrate-dependent bNAbs, PGT121 binds to complex-type, rather than high-mannose, N-glycans in glycan microarray experiments. Crystal structures of PGT121 and 10-1074 compared with structures of their germ-line precursor and a structure of PGT121 bound to a complex-type N-glycan rationalize their distinct properties.     

Long-term exposure to elevated CO2 enhances plant community stability by suppressing dominant plant species in a mixed-grass prairie

Climate controls vegetation distribution across the globe, and some vegetation types are more vulnerable to climate change, whereas others are more resistant. Because resistance and resilience can influence ecosystem stability and determine how communities and ecosystems respond to climate change, we need to evaluate the potential for resistance as we predict future ecosystem function. In a mixed-grass prairie in the northern Great Plains, we used a large field experiment to test the effects of elevated CO₂, warming, and summer irrigation on plant community structure and productivity, linking changes in both to stability in plant community composition and biomass production. We show that the independent effects of CO₂ and warming on community composition and productivity depend on interannual variation in precipitation and that the effects of elevated CO₂ are not limited to water saving because they differ from those of irrigation. We also show that production in this mixed-grass prairie ecosystem is not only relatively resistant to interannual variation in precipitation, but also rendered more stable under elevated CO₂ conditions. This increase in production stability is the result of altered community dominance patterns: Community evenness increases as dominant species decrease in biomass under elevated CO₂. In many grasslands that serve as rangelands, the economic value of the ecosystem is largely dependent on plant community composition and the relative abundance of key forage species. Thus, our results have implications for how we manage native grasslands in the face of changing climate.Ecologists have long recognized the importance of climate in shaping plant communities across spatial and temporal scales (1). Together, precipitation and temperature characterize the distribution of terrestrial biomes across the globe. As climate changes, some biomes will be more vulnerable to temperature increase (2) or altered precipitation (3), whereas others will be more resistant (4–6). Ecological stability, the maintenance of community structure and function despite climatic fluctuation or disturbance (7–9), includes two components: resistance [lack of change despite perturbation (9)] and resilience [return to a previous state following a perturbation (10–13)]. Diversity (14) and productivity (11, 15) can both influence community stability (16) and dampen responses to environmental perturbation (5, 9, 17, 18). What remains unclear is how stability and resistance respond to predicted changes in climate.Multiple climate change factors simultaneously impact plant performance, community structure, and productivity (4, 19, 20). For example, elevated CO₂ can improve water use efficiency and increase plant productivity (21–23), but warming can reduce it, counteracting the positive water-saving effects of elevated CO₂ (24). In addition, plant species and functional groups that differ in photosynthetic pathway often have contrasting responses to elevated CO₂, warming, and altered precipitation. Furthermore, the effects of individual climate change factors may be additive (25, 26), subadditive (4, 24, 27), or antagonistic (27, 28). As a result, the performance of a given species or functional group depends on interactions among CO₂, temperature, and soil characteristics that influence plant water availability at the community level.Globally, both elevated CO₂ and warming are expected to lead to pronounced changes in vegetation distribution and structure (25, 29, 30). In North American grasslands, warming is expected to promote C₄ dominance, dampening the ability of these areas to show large responses to elevated CO₂ (25). Because responses to climate change differ among individual plant species and depend on community context (31–33), the resultant community dynamics are difficult to predict. In addition, plant responses to climate manipulations can shift over time. Our earlier work in a mixed-grass prairie shows that in the first 3 y of the Prairie Heating and CO₂ Enrichment (PHACE) experiment, both C₃ and C₄ grass production benefited from elevated CO₂ conditions (34). However, long-term studies of CO₂ enrichment show that plant responses can diminish over time (22, 35), including the responses of dominant grass species in our mixed-grass prairie (36). To accurately characterize the trajectory of species responses and predict the interacting impacts of global climate change on plant community structure and function, long-term experiments are necessary.Grasslands in the northern Great Plains are experiencing rapid climate change, with average annual temperatures increasing by 2.6 °C over the last century and winter and spring temperatures increasing more rapidly than summer temperatures (37). Grasslands are extensively grazed, and moisture availability (timing and amount of rainfall) affects grassland productivity to support domestic and native herbivores (3, 38). Compared with other regions, precipitation change is expected to be relatively modest, but there is a general consensus that even if annual precipitation change is small, precipitation timing will become increasingly variable (37) and the number of extreme precipitation events will also increase (39–41). When coupled with rising temperatures, water limitation will increase (42), potentially reducing rangeland productivity (43). Because the timing of water availability regulates grassland productivity and community dynamics (3, 44), variation in background climate may promote or reduce the resistance of grasslands to climate change. The economic value of the ecosystem is largely dependent on the plant community and the relative abundance of key forage grass species (45). Thus, changes in grassland productivity can have clear economic impacts for ranching and managing wildlife (46).To understand how climate change influences plant community dynamics and stability (namely, resistance to interannual shifts in precipitation), we quantified the impacts of experimentally imposed elevated CO₂, warming, and summer irrigation on plant community composition and aboveground biomass production over 8 y in a northern mixed-grass prairie in southeastern Wyoming. Species that dominate biomass production are expected to respond to changes in climate most directly (47), whereas subdominant species may respond to climate change directly and indirectly through their interactions with the dominant species (6, 48, 49). Thus, we quantified climate change effects on the entire community and on dominant and subdominant community members separately. We addressed three questions: (i) Do the effects of climate change on plant community composition and productivity depend on temperature and precipitation variation? (ii) Do dominant and subdominant components of the plant community respond differently to climate change? and (iii) What is the influence of climate change on community composition and biomass stability?     

Multiannual forecasting of seasonal influenza dynamics reveals climatic and evolutionary drivers

Human influenza occurs annually in most temperate climatic zones of the world, with epidemics peaking in the cold winter months. Considerable debate surrounds the relative role of epidemic dynamics, viral evolution, and climatic drivers in driving year-to-year variability of outbreaks. The ultimate test of understanding is prediction; however, existing influenza models rarely forecast beyond a single year at best. Here, we use a simple epidemiological model to reveal multiannual predictability based on high-quality influenza surveillance data for Israel; the model fit is corroborated by simple metapopulation comparisons within Israel. Successful forecasts are driven by temperature, humidity, antigenic drift, and immunity loss. Essentially, influenza dynamics are a balance between large perturbations following significant antigenic jumps, interspersed with nonlinear epidemic dynamics tuned by climatic forcing.Influenza outbreaks have been documented in the scientific literature in records that extend back to at least 1650 (1), making it an exceptional example of a persisting, recurrent disease. Being a respiratory infection, influenza spreads rapidly from person to person through a population in the form of virus particles airborne as respiratory droplets or aerosols. Depending on the circumstances, influenza typically infects between 10% and 50% of a given population and has become a source of considerable human morbidity and mortality (2). There is much controversy in identifying the seasonal drivers that generate annual influenza epidemics and the processes that give rise to their large variability (3–12). This is an outstanding problem of influenza research today. Using long-term modeling, a recent study (9) gave support to the possibility that absolute humidity is the predominant determinant of influenza seasonality in temperate zones, driving disease transmission and controlling the timing of individual wintertime outbreaks. Another study investigated the physical properties of absolute humidity on influenza virus transmission and influenza virus survival (3). However, a general understanding of the mechanisms underlying influenza seasonal variation remains quite limited (8). Here, we use a simple mathematical model to unravel the interplay between climate and evolution to predict long-term influenza dynamics correctly for the years since June 2010.A requirement for the generation of recurrent epidemics is a sufficient and continuous source of new susceptible individuals arising in the population, enough to fuel each new outbreak (13). In the case of influenza, infected individuals recover with immunity but eventually become susceptible again because of the rapidly evolving nature of the influenza virus (7, 14). Positive selection exerted by the host immune system leads to a continual antigenic drift of the influenza virus’s glycoproteins, particularly the main antigen, hemagglutinin, thus allowing the virus to eventually evade the immune system (15). The process of antigenic drift thereby creates an important renewed source of susceptible individuals. Hence, evolutionary forces are considered tremendously important in shaping complex recurrent patterns of infectious diseases and explain why influenza is regarded as “an invariable disease caused by a variable virus” (1).The changing rate of antigenic drift also has a significant impact on the timing and amplitude of influenza outbreaks (16). Recent studies reveal that the evolution of influenza A H3N2’s main antigen is punctuated in character such that the drift occurs within discrete antigenic clusters (neutral periods), but with jumps to newly arising clusters after irregular periods (17, 18). A significant jump for the A H3N2 lineage last occurred during the 2003–4 season with the appearance of the A/Fujian virus strain coinciding with a sharp influenza outbreak approximately 2 mo earlier than usual, with a normal attack rate. Nevertheless, it is difficult to demonstrate a consistent and conclusive direct link between the size of antigenic jumps and changes in influenza dynamics at the population level (6, 19–21).     

Novel serologic biomarkers provide accurate estimates of recent Plasmodium falciparum exposure for individuals and communities

Tools to reliably measure Plasmodium falciparum (Pf) exposure in individuals and communities are needed to guide and evaluate malaria control interventions. Serologic assays can potentially produce precise exposure estimates at low cost; however, current approaches based on responses to a few characterized antigens are not designed to estimate exposure in individuals. Pf-specific antibody responses differ by antigen, suggesting that selection of antigens with defined kinetic profiles will improve estimates of Pf exposure. To identify novel serologic biomarkers of malaria exposure, we evaluated responses to 856 Pf antigens by protein microarray in 186 Ugandan children, for whom detailed Pf exposure data were available. Using data-adaptive statistical methods, we identified combinations of antibody responses that maximized information on an individual’s recent exposure. Responses to three novel Pf antigens accurately classified whether an individual had been infected within the last 30, 90, or 365 d (cross-validated area under the curve = 0.86–0.93), whereas responses to six antigens accurately estimated an individual’s malaria incidence in the prior year. Cross-validated incidence predictions for individuals in different communities provided accurate stratification of exposure between populations and suggest that precise estimates of community exposure can be obtained from sampling a small subset of that community. In addition, serologic incidence predictions from cross-sectional samples characterized heterogeneity within a community similarly to 1 y of continuous passive surveillance. Development of simple ELISA-based assays derived from the successful selection strategy outlined here offers the potential to generate rich epidemiologic surveillance data that will be widely accessible to malaria control programs.Many countries have extensive programs to reduce the burden of Plasmodium falciparum (Pf), the parasite responsible for most malaria morbidity and mortality (1). Effectively using limited resources for malaria control or elimination and evaluating interventions require accurate measurements of the risk of being infected with Pf (2–15). To reflect the rate at which individuals are infected with Pf in a useful way, metrics used to estimate exposure in a community need to account for dynamic changes over space and time, especially in response to control interventions (16–18).A variety of metrics can be used to estimate Pf exposure, but tools that are more precise and low cost are needed for population surveillance. Existing metrics have varying intrinsic levels of precision and accuracy and are subject to a variety of extrinsic factors, such as cost, time, and availability of trained personnel (19). For example, entomological measurements provide information on mosquito to human transmission for a community but are expensive, require specially trained staff, and lack standardized procedures, all of which reduce precision and/or make interpretation difficult (19–22). Parasite prevalence can be measured by detecting parasites in the blood of individuals from a cross-sectional sample of a community and is, therefore, relatively simple and inexpensive to perform, but results may be imprecise, especially in areas of low transmission (19, 23), and biased by a number of factors, including immunity and access to antimalarial treatment (5, 6, 19, 23–25). The burden of symptomatic disease in a community can be estimated from routine health systems data; however, such data are frequently unreliable (5, 26–28) and generally underestimate the prevalence of Pf infection in areas of intense transmission. Precise and quantitative information about exposure at an individual level can be reliably obtained from cohort studies by measuring the incidence of asymptomatic and/or symptomatic Pf infection (i.e., by measuring the molecular force of infection) (29–35). Unfortunately, the expense of cohort studies limits their use to research settings. The end result is that most malaria-endemic regions lack reliable, timely data on Pf exposure, limiting the capabilities of malaria control programs to guide and evaluate interventions.Serologic assays offer the potential to provide incidence estimates for symptomatic and asymptomatic Pf infection, which are currently obtained from cohort studies, at the cost of cross-sectional studies (36–38). Although Pf infections are transient, a record of infection remains detectable in an individual’s antibody profile. Thus, appropriately chosen antibody measurements integrated with age can provide information about an individual’s exposure history. Antibodies can be measured by simple ELISAs and obtained from dried blood spots, which are easy to collect, transport, and store (39–41). Serologic responses to Pf antigens have been explored as potential epidemiological tools (42–45), and estimated rates of seroconversion to well-characterized Pf antigens accurately reflect stable rates of exposure in a community, whereas distinct changes in these rates are obtained from successful interventions (22, 39, 41, 46–53). However, current serologic assays are not designed to detect short-term or gradual changes in Pf exposure or measure exposure to infection at an individual level. The ability to calibrate antibody responses to estimates of exposure in individuals could allow for more flexible sampling of a population (e.g., not requiring age stratification), improve accuracy of exposure estimates from small sample sizes, and better characterize heterogeneity in exposure within a community.Different Pf antigens elicit antibody responses with different magnitudes and kinetics, providing a large and diverse set of potential biomarkers for exposure (38, 54–58). We hypothesized that new and more highly informative serologic biomarkers better able to characterize an individual’s recent exposure history could be identified by analyzing antibody responses to a large number of candidate Pf antigens in participants with well-characterized exposure histories. To test this hypothesis, we probed plasma from participants in two cohort studies in Uganda against a protein microarray containing 856 Pf antigens. The primary aim of this analysis was to identify responses to select antigens that were most informative of recent exposure using robust, data-adaptive statistical methods. Each participant’s responses to these selected antigens were used as predictors for two primary outcomes of their recent exposure to Pf: (i) days since last Pf infection and (ii) the incidence of symptomatic malaria in the last year. These individual-level estimates were then aggregated across a population to assess community-level malaria exposure. The selection strategy presented here identified accurate biomarkers of exposure for children living in areas of moderate to high Pf exposure and illustrates the utility of this flexible and broadly applicable approach.