Severe Karenia brevis red tides influence juvenile bottlenose dolphin (Tursiops truncatus) behavior in Sarasota Bay,Florida

Harmful algal blooms (HABs) are natural stressors in the coastal environment that may be increasing in frequency and severity. This study investigates whether severe red tide blooms, caused by Karenia brevis, affect the behavior of resident coastal bottlenose dolphins in Sarasota Bay, Florida through changes to juvenile dolphin activity budgets, ranging patterns, and social associations. Behavioral observations were conducted on free‐ranging juvenile dolphins during the summer months of 2005–2007, and behavior during red tide blooms was compared to periods of background K. brevis abundance. We also utilized dolphin group sighting data from 2004 to 2007 to obtain comparison information from before the most severe recent red tide of 2005 and incorporate social association information from adults in the study area. We found that coastal dolphins displayed a suite of behavioral changes associated with red tide blooms, including significantly altered activity budgets, increased sociality, and expanded ranging behavior. At present, we do not fully understand the mechanism behind these red tide‐associated behavioral effects, but they are most likely linked to underlying changes in resource availability and distribution. These behavioral changes have implications for more widespread population impacts, including increased susceptibility to disease outbreaks, which may contribute to unusual mortality events during HABs.     

Concurrent exposure of bottlenose dolphins (Tursiops truncatus) to multiple algal toxins in Sarasota Bay, Florida, USA

Sentinel species such as bottlenose dolphins (Tursiops truncatus) can be impacted by large-scale mortality events due to exposure to marine algal toxins. In the Sarasota Bay region (Gulf of Mexico, Florida, USA), the bottlenose dolphin population is frequently exposed to harmful algal blooms (HABs) of Karenia brevis and the neurotoxic brevetoxins (PbTx; BTX) produced by this dinoflagellate. Live dolphins sampled during capture-release health assessments performed in this region tested positive for two HAB toxins; brevetoxin and domoic acid (DA). Over a ten-year study period (2000–2009) we have determined that bottlenose dolphins are exposed to brevetoxin and/or DA on a nearly annual basis (i.e., DA: 2004, 2005, 2006, 2008, 2009; brevetoxin: 2000, 2004, 2005, 2008, 2009) with 36% of all animals testing positive for brevetoxin (n = 118) and 53% positive for DA (n = 83) with several individuals (14%) testing positive for both neurotoxins in at least one tissue/fluid. To date there have been no previously published reports of DA in southwestern Florida marine mammals, however the May 2008 health assessment coincided with a Pseudo-nitzschia pseudodelicatissima bloom that was the likely source of DA observed in seawater and live dolphin samples. Concurrently, both DA and brevetoxin were observed in common prey fish. Although no Pseudo-nitzschia bloom was identified the following year, DA was identified in seawater, fish, sediment, snails, and dolphins. DA concentrations in feces were positively correlated with hematologic parameters including an increase in total white blood cell (p = 0.001) and eosinophil (p<0.001) counts. Our findings demonstrate that dolphins within Sarasota Bay are commonly exposed to two algal toxins, and provide the impetus to further explore the potential long-term impacts on bottlenose dolphin health.     

象山港硅藻赤潮过程中真菌群落演替及互作特征

[背景] 近岸海域赤潮的发生能显著改变包括真菌在内的微生物群落组成,进而影响海洋中碳氮元素循环。真菌是海洋中重要的分解者,但赤潮过程对真菌群落的影响少有报道。[目的] 探明赤潮过程对真菌群落的影响,对进一步阐释赤潮对生态系统中物质循环的影响具有重要意义。[方法] 针对2017年春季象山港硅藻赤潮,对真菌内源转录间隔区1（Internally Transcribed Spacer 1,ITS1）序列进行高通量测序,研究赤潮过程中真菌群落响应及共现性特征。[结果] 真菌群落的α多样性指数在赤潮暴发的各阶段差异显著,而且均与磷酸盐、硝酸盐和亚硝酸盐有显著相关性（P<0.05）。子囊菌门（Ascomycota,90.2%）和担子菌门（Basidiomycota,8.27%）是本次硅藻赤潮的优势菌门,其中子囊菌门遍布赤潮暴发的4个阶段;锤舌菌纲（Leotiomycetes,16.1%）和散囊菌纲（Eurotiomycetes,9.3%）是纲水平上的主要优势类群。赤潮暴发过程中硝酸盐、亚硝酸盐、磷酸盐和温度等环境因素的改变,驱动真菌群落组成及结构发生显著变化。Helotiales、Eurotiales、Xylariales、Sacc haromycetales和Agaricostilbales构成了真菌群落共现网络的主体。[结论] 真菌群落在赤潮的各阶段具有显著的演替特征,赤潮藻生消是引起真菌群落演替的主要因素,但是环境因子也扮演着重要角色。赤潮过程中,真菌群落同类群内的互作要强于不同类群间的互作,这对维持群落的稳定性具有重要意义。     

Metabolic rate estimates for a eutrophic lake from diel dissolved oxygen signals

Estimates of net primary production, community respiration (R'), and gross primary production (P _g) are developed and presented for the productive layers of eutrophic Onondaga Lake, NY, U.S.A., for time scales ranging from diel to several months, based on 4 months of robotic diel profiles of dissolved oxygen (DO) and temperature. Metabolic rate calculations are made through application of a DO mass balance framework that also accommodates inputs and losses of DO mediated by exchange across the air–water interface and across the lower boundary of the productive layers. It is demonstrated that the dynamics of the flux across the air–water interface are important to the metabolic rate estimates, while vertical mixing-based losses to the underlying layers can be ignored. Study average estimates of R' (1.49 g O₂ m^–3 d^–1) and P _g (1.60 g O₂ m^–3 d^–1) obtained by this non-isolated community approach are consistent with levels reported in the literature for similar chlorophyll a concentrations, based on isolated community (bottle experiment) protocols to measure these metabolic rates. The non-isolated community approach is shown to have limited utility for quantifying day-to-day changes in these rates in this lake, apparently because of horizontal exchange with waters of different DO concentrations. However, this approach may support reliable estimates of metabolic rates at intermediate time scales; e.g., several days to a week. The DO mass balance framework is demonstrated to be valuable in resolving the relative roles of various physical and biological processes in regulating the DO pool of the productive layers.     

Physical maturity in common bottlenose dolphins (Tursiops truncatus) from Sarasota Bay,Florida

Cetacean physical maturity is defined by growth cessation and complete fusion of epiphyses to vertebral bodies indicated by invisible sutures. Many studies have shown epiphyseal fusion is highly variable among individuals. In-depth examinations into fusion variability are lacking. We analyzed vertebrae of 37 (n = 21 female, n = 16 male) stranded common bottlenose dolphins (Tursiops truncatus) from the well-studied Gulf of Mexico, Sarasota Bay community. For each specimen, vertebrae were examined by vertebral region for degree of fusion anteriorly and posteriorly of each centrum and categorized from unfused to fused in five degrees. An ordinal logistic regression was used to estimate degree of fusion probability for each epiphysis. The model had fixed effects for age, number of offspring, sex, sexual maturity, and a random effect for epiphysis. Results show that age/reproductive status significantly explains an individual's degree of fusion. Adult females with fewer calves had more fusion than those with more reproductive experience across multiple ages. Access to long-term observational and sample data on the dolphins residing in the area served by Mote Marine Laboratory's Stranding Investigations Program offers a unique opportunity to examine the relationship between energetic demands of reproduction (calcium production/reproductive output) versus preconceived definitions of physical maturity (skeletal fusion) more closely.     

Recreational fishing depredation and associated behaviors involving common bottlenose dolphins (Tursiops truncatus) in Sarasota Bay,Florida

Odontocete depredation involves stealing or damaging bait or prey already captured by fishing gear. The increase in depredation is of concern for small stocks of cetaceans because interactions with fishing gear can lead to serious injury or mortality through entanglement or ingestion. Using long‐term data sets available for the bottlenose dolphin (Tursiops truncatus) community in Sarasota Bay, Florida, we investigated recreational fishing gear interactions by (1) examining temporal patterns in depredation and associated behaviors from 2000 to 2007; (2) quantifying the behavior of dolphins that depredate or engage in associated behaviors; and (3) identifying factors associated with the rise in depredation locally. The number of incidents of dolphins (primarily adult males) interacting with recreational anglers and boaters increased following 2004. Depredation and associated behaviors increased during red tide lags and tourist seasons during times of prey depletion and heightened angler and boater activity. Dolphins with a history of fishing gear interactions shifted away from natural activity patterns and were more likely to be within 50 m of fishing lines. Recreational fishing gear interactions were attributed to a two percent population decline in Sarasota Bay in 2006 and need to be considered along with other cumulative human impacts in the development of conservation measures for dolphins.     

Anthropogenic injuries disrupt social associations of common bottlenose dolphins (Tursiops truncatus) in Sarasota Bay,Florida

Social connectivity is important for measuring the fitness of common bottlenose dolphins (Tursiops truncatus). While interactions in fission-fusion societies vary between individuals, studies show that repeated interactions enhance reproduction and foraging success. Injuries that potentially remove an individual from its association network may disrupt these interactions. Using data from the long-term resident dolphin community in Sarasota Bay, Florida, we investigated how anthropogenic injuries affect the dolphins' social associations by examining the differences before and after injury to individuals. We examined group size, strength, eigenvector centrality, clustering coefficient, and number of triangles and analyzed whether the animal's sex, age class, type of injury, or human intervention affected these values. We found that while group size did not change, injured dolphins had fewer preferred associates (HWI > 0.14) and were found in more fluid groups immediately after injury, but started returning to normal association levels after 2 years. This initial decrease in connectivity was not related to the age, sex, type of injury, or intervention. Despite the fluidity in individual associations, the strongest bonds remained stable, those between mothers and calves and those between male alliance partners. These findings provide some of the first information relating injuries and social networks for animals.     

Importance of phytoplankton carbon to heterotrophic bacteria in the Ohio,Cumberland, and Tennessee rivers,USA

The flux of carbon and nutrients through aquatic systems is largely dependent upon interactions between autotrophic and heterotrophic processes. As a means of assessing the relative importance of autotrophy and heterotrophy in large rivers, we compared phytoplankton production, heterotrophic bacterial production and community respiration in three regulated rivers of the Midwestern USA. Samples were collected monthly (March to December 1999) from impoundments of the Ohio (McAlpine Pool), Cumberland (Lake Barkley), and Tennessee (Kentucky Lake) Rivers. Bacterial production was tightly coupled to phytoplankton production at each site (r ² = 0.63–0.85). Ratios of phytoplankton production to bacterial production ranged from <1 to 15 in the Tennessee and Cumberland Rivers and 2 to 90 in the Ohio River. The ratio of primary production to community respiration (P:R) ranged from 0.03 to 2.76 across all sites, with average P:R values lower in the Ohio River (0.14) than the Tennessee River (0.39) and the Cumberland River (1.10). P:R values above unity (P > R) were observed only in the Tennessee and Cumberland Rivers during seasonal (April–July) spikes in primary production. We estimate that 3, 6, and 20% of annual bacterial carbon requirements were met by exudates from in situ phytoplankton in the Ohio River, Tennessee River, and Cumberland River, respectively. Our findings indicate that heterotrophic bacteria were largely dependant upon allochthonous carbon. Autochthonous sources provided supplemental organic matter (up to 40% of bacterial carbon demand) during summer low flow. Handling editor: J. Padisak     

Differences in oxygen consumption and diel activity as adaptations related to microhabitat in Neotropical freshwater decapods (Crustacea)

This study evaluated oxygen consumption (MO₂) and diel activity in Aegla longirostri, Trichodactylus panoplus and Parastacus brasiliensis (three species of freshwater decapods that occur in sympatry), under two different conditions of O₂ availability, limited and constant; and searched for the existence of a relationship between these two variables. The Kruskal–Wallis test showed that in all the species, MO₂ was higher under constant O₂ availability; T. panoplus and P. brasiliensis showed an oxygen-dependent pattern, while A. longirostri showed higher MO₂ values and less variation in the values between the treatments, indicating a higher and more oxygen-independent metabolism. P. brasiliensis was more active in constant O₂. A. longirostri was more active in limited O₂ and did not show a clear diel activity in any case, showing behavioral changes when in unfavorable conditions. The Spearman's rank correlation analysis did not indicate any relationship between MO₂ and activity. These results indicate a higher metabolism in A. longirostri. The less demanding metabolisms of P. brasiliensis and T. panoplus allow these species to occupy environments that are unavailable to A. longirostri due to differences in dissolved-oxygen concentrations.     

Annual and diel oxygen regime in two polder ditches

The oxygen regime of two polder ditches and two enclosures within these ditches was studied. Continous oxygen temperature and light measurements were performed for 24-hour periods each month during two and a half year in the ditches and one year in the enclosures. Oxygen concentrations between 0 and 23 ppm were found, with diurnal ranges as large as 18 ppm. Steep gradients between bottom and surface could develop, but mostly disappeared during nightly turnover. The 10-percentile of the surface water measured between 9 and 17 hours was above 3 ppm, fullfilling the Dutch standards for this type of ecosystems. The oxygen concentrations near the bottom, however, could drop to zero and during the night surface concentrations below 1 ppm were measured. Based on average oxygen saturation values it is concluded that in the open water of the ditches oxygen consumption prevailed while in the enclosures oxygen production was most important. Based on the mass balance equation gross primary production and respiration were calculated. Annual average respiration varied between 2.5 and 6.6 g O₂.m^–2.d^–1 and average gross primary production between 3.2 and 4.8 g O₂.m^–2.d^–1. Maximum daily production and respiration were 15.9 and 22.3 g O₂.m^–2.d^–1. These figures classify the polder ditches as highly productive aquatic ecosystems.     

Sulfate reduction and thiosulfate transformations in a cyanobacterial mat during a diel oxygen cycle

Abstract Bacterial sulfate reduction and transformations of thiosulfate were studied with radiotracers in a Microcoleus chthonoplastes -dominated microbial mat growing in a hypersaline pond at the Red Sea. The study showed how a diel cycle of oxygen evolution affected respiration by sulfate-reducing bacteria and the metabolism of thiosulfate through oxidative and reductive pathways. Sulfate reduction occurred in both oxic and anoxic layers of the mat and varied diurnally, apparently according to temperature rather than to oxygen. Time course experiments showed that the radiotracer method underestimated sulfate reduction in the oxic zone due to rapid reoxidation of the produced sulfide. Extremely high reduction rates of up to 10 μmol cm⁻³ d⁻¹ were measured just below the euphotic zone. Although thiosulfate was simultaneously oxidized, reduced and disproportionated by bacteria in all layers of the mat, there was a shift from predominant oxidation in the oxic zone to predominant reduction below. Concurrent disproportionation of thiosulfate to sulfate and sulfide occurred in all zones and was an important pathway of the sulfur cycle in the mat.     

Bacterial dynamics and distribution during a spring diatom bloom in the Hudson River plume, USA

Bacterial abundance and [³H]thymidine incorporation rates wereused to characterize bacterial distributions and dynamics duringa spring diatom bloom in the coastal plume of the Hudson Riverduring March, 1981. Bacterial abundance did not decline significantlyaway from the plume or across the continental shelf. However,a pronounced gradient was observed for [³H]thymidine incorporation. Following a northeast gale, a bloom of Skeletonema costatumdeveloped in response to coastal upwelling. As the plume becamesaltier, warmer and larger, bacterial abundance averaged 1.3x 10⁹ cells 1^–1. Bacterial incorporation of [³H]thymidineinto cold 5% TCA insoluble materials, bacterial production andspecific growth rates averaged 80–120 pmol 1^–1 d^–1,1.4–2.0 x 10⁹ cells 1^–1 and 1.2–1.5 d^–1.respectively. The mean density of bacteria in the plume didnot change even though growth rates were higher than expectedlosses from sinking, mixing and export. The abundance and production levels of attached bacteria inthe plume were significantly higher than they were in coastalwater outside the plume. In contrast, free bacterial levelswere similar in all regions of the shelf and plume. Bacterialparameters in the plume were not correlated with phytoplanktonpigment concentrations, while significant positive correlationswere found in shelf waters >33 salinity. Thus bacteria werecoupled to phytoplankton in shelf waters but not in the plumewhere allochthonous dissolved matter was probably supportingthe bacteria. ^*Lamont-Doherty Geological Observatory Contribution No. 3459 ²Present address: Department of Microbiology, University ofGeorgia, Athens, GA 30602, USA.     

Annual, seasonal and individual variation in hematology and clinical blood chemistry profiles in bottlenose dolphins (Tursiops truncatus) from Sarasota Bay, Florida

Hematology and clinical blood chemistry (HCBC) profiles in free-living bottlenose dolphins from Sarasota Bay, Florida have been monitored over a 14-year period. This long-term dataset includes samples from recaptured dolphins, enabling individual variation to be accounted for when investigating seasonal and annual variability. Four different laboratories carried out the assays and inter-laboratory comparisons found significant differences in 31 of 39 parameters measured. However, variability in comparable HCBCs by sex, age, condition, season and year could be investigated. Significant relationships with the independent variables were found for the majority of the HCBCs. Notable consistent seasonal differences included significantly elevated glucose and significantly lower creatinine concentrations in winter compared to summer. These differences may be due to energetic or thermoregulatory fluctuations in the animals by season and do not necessarily have any clinical significance. Erythrocyte counts were significantly lower in the winter, possibly also due to nutritional differences. Albumin and calcium levels in this population have increased significantly over the years of monitoring and consistently across seasons, being higher in the winter than the summer. Again, nutritional and thermal constraints seem to be the most likely environmental factors influencing these patterns.     

Three-dimensional structure of a Karenia brevis bloom: Observations from gliders,satellites, and field measurements

Autonomous underwater gliders with customized sensors were deployed in October 2011 on the central West Florida Shelf to measure a Karenia brevis bloom, which was captured in satellite imagery since late September 2011. Combined with in situ taxonomy data, satellite measurements, and numerical circulation models, the glider measurements provided information on the three-dimensional structure of the bloom. Temperature, salinity, fluorescence of colored dissolved organic matter (CDOM) and chlorophyll-a, particulate backscattering coefficient, and K. brevis-specific chlorophyll-a concentrations were measured by the gliders over >250 km from the surface to about 30-m water depth on the shallow shelf. At the time of sampling the bloom was characterized by uniform vertical structures, with relatively high chlorophyll-a and CDOM fluorescence, low temperature, and high salinity. Satellite data extracted along the glider tracks demonstrated coherent spatial variations as observed by the gliders. Further, the synoptic satellite observations revealed the bloom evolution during the 7 months between late September 2011 and mid April 2012, and showed the maximum bloom size of ∼3000 km² around 23 November. The combined satellite and in situ data also confirmed that the ratio of satellite-derived fluorescence line height (FLH) to particulate backscattering coefficient at 547 nm (b_bp(547)) could be used as a better index than FLH alone to detect K. brevis blooms. Numerical circulation models further suggested that the bloom could have been initiated offshore and advected onshore via the bottom Ekman layer. The case study here demonstrates the unique value of an integrated coastal ocean observing system in studying harmful algal blooms (HABs).     

Influence of teleost abundance on the distribution and abundance of sharks in Florida Bay, USA

Understanding the factors that influence the distribution and abundance of predators, including sharks, is important for predicting the impacts of human changes to the environment. Such studies are particularly important in Florida Bay, USA where there are planned large-scale changes to patterns of freshwater input from the Everglades ecosystem. Studies of many marine predators suggest that links between predator and prey habitat use may vary with spatial scale, but there have been few studies of the role of prey distribution in shaping habitat use and abundance of sharks. We used longline catches of sharks and trawls for potential teleost prey to determine the influence of teleost abundance on shark abundance at the scale of regions and habitats in Florida Bay. We found that shark catch per unit effort (CPUE) was not linked to CPUE ofteleosts at the scale of sampling sites, but shark CPUE was positively correlated with the mean CPUE for teleosts within a region. Although there does not appear to be a strong match between the abundance of teleosts and sharks at small spatial scales, regional shark abundance is likely driven, at least partially, by the availability of prey. Management strategies that influence teleost abundance will have cascading effects to higher trophic levels in Florida Bay. Electronic Supplementary Material is available for this article at     

藻华发生过程中胶州湾水体颗粒有机物脂肪酸的组成与动态

胶州湾在2005年夏季发生了中肋骨条藻(Skeletonema costatum ( Grev.) Cleve)藻华,该优势种占细胞总量的比例在两个研究站位(C3站及A5站)分别达到59%和86%.对藻华发生过程中(7月,8月及9月份)的颗粒有机物进行了采样分析,测定了脂肪酸、叶绿素(Chl-a)和颗粒有机碳(POC)的浓度,分析了藻华发生过程中浮游植物和脂肪酸的组成特征,并通过标志脂肪酸对藻华发生过程中颗粒有机物组成的动态变化进行了分析.总脂肪酸浓度同叶绿素和颗粒有机碳浓度一样,在藻华发生期达到最高,两个采样站位(C3站及A5站)分别为29.0μg·L-1及185.5μg·L-1,比藻华发生前和消退后高2～3倍及20倍.胶州湾悬浮颗粒脂肪酸主要包括16:0,14:0,18:0等直链饱和脂肪酸(SSFA),16:1ω7、16:1ω5+ω9、18:1ω9,18:1ω7等单不饱和脂肪酸(MUFA)及以 20:5ω3(DHA)和22:6ω3(EPA)为主的多不饱和脂肪酸(PUFA)和以i-15:0及ai-15:0为主的支链饱和脂肪酸(BSFA).藻华期与发生前相比,脂肪酸中的不饱和组分(MUFA,PUFA)所占比例在两个取样站位均有提高.随着藻华的消退,藻华区域中心的A5 站的SSFA及BSFA比例有大幅提高,而在非藻华中心区域的C3站,不饱和脂肪酸尤其是ω3系列多不饱和脂肪酸(DHA, EPA等)比例的上升非常显著.藻华发生时浮游植物在颗粒有机物中的比重提高.藻华消退后,A5站颗粒有机物中碎屑有机物及细菌所占比重提高,而C3站颗粒有机物中浮游植物的比重仍然很高.浮游植物大量死亡是A5站藻华消退的主要形式,而C3站藻华的消退则可能与浮游动物的摄食有关.     

Mathematical modelling of primary production in Green Bay (Lake Michigan,USA), a phosphorus-and light-limited system

Application of mathematical models in the design and evaluation of lake restoration programmes must include due consideration of three basic concepts of model development; 1) that the model framework is appropriately matched to the intended management use, 2) that selection of the proper degree of model complexity is fundamental to the achievement of model credibility and 3) that field and laboratory studies must be designed and interpreted with the aid of the model to insure development of a comprehensive, integrated tool.These concepts are demonstrated for the case of lake restoration efforts in Green Bay (Lake Michigan, USA). Striking gradients in water quality (transparency, algal standing crop, hypolimnetic oxygen depletion) and trophic state occur along the major axis of the bay in response to phosphorus loaded from the Fox River. A simple model for gross primary production is developed to permit calculation of the relative importance of internal carbon production to the total organic carbon budget of the bay. Primary production varies from high rates over a limited photic depth in the turbid, phosphorus-rich waters of the eutrophic portions of the bay to low rates over an extensive photic depth in the transparent, phosphoruspoor reaches of the oligotrophic regions. Internal production accounts for approximately 90% of the total organic carbon loaded to the system over the summer growing season. Water quality management strategies must address the stimulation of primary production by phosphorus loaded from the Fox River in any attempt to lower the standing crop of nuisance algae, improve water clarity, and reduce rates of hypolimnetic oxygen depletion in Green Bay.     

Using infrared thermography to assess seasonal trends in dorsal fin surface temperatures of free-swimming bottlenose dolphins (Tursiops truncatus) in Sarasota Bay, Florida

The temperature differential (Δ T ) between a body surface and the environment influences an organism's heat balance. In Sarasota Bay, FL, where ambient water temperature ( T _w) ranges annually from 11° to 33°C, Δ T was investigated in a resident community of bottlenose dolphins ( Tursiops truncatus ). Dorsal fin surface temperatures ( T _dfin) were measured on wild, free-swimming dolphins using infrared thermography. Field and laboratory calibration studies were also undertaken to assess the efficacy of this non-invasive technology in the marine environment. The portability of infrared thermography permitted measurements of T _dfin across the entire range of environmental temperatures experienced by animals in this region. Results indicated a positive, linear relationship between T _dfin and T _w ( r ²= 0.978, P < 0.001). On average, T _dfin was 0.9°C warmer than T _w across seasons, despite the 22°C annual range in T _w. Changes in integumentary and vascular insulation likely account for the stability of Δ T _{dfin − w} and the protection of core temperature ( T _core) across seasons. The high thermal conductivity of water may also influence this Δ T . The use of infrared thermography is an effective, non-invasive method of assessing dorsal fin skin surface temperatures (±1°C) across large numbers of wild, free-swimming dolphins throughout their thermally dynamic aquatic environment.