Counter-gradient variation in gene expression between fish populations facilitates colonization of low-dissolved oxygen environments.

The role of phenotypic plasticity during colonization remains unclear due to the shifting importance of plasticity across timescales. In the early stages of colonization, plasticity can facilitate persistence in a novel environment; but over evolutionary time, processes such as genetic assimilation may reduce variation in plastic traits such that species with a longer evolutionary history in an environment can show lower levels of plasticity than recent invaders. Therefore, comparing species in the early stages of colonization to long-established species provides a powerful approach for uncovering the role of phenotypic plasticity during different stages of colonization. We compared gene expression between low-dissolved oxygen (DO) and high-DO populations of two cyprinid fish: Enteromius apleurogramma, a species that has undergone a recent range expansion, and E. neumayeri, a long-established native species in the same region. We sampled tissue either immediately after capture from the field or after a 2-week acclimation under high-DO conditions, allowing us to test for both evolved and plastic differences in low-DO vs high-DO populations of each species. We found that most genes showing candidate-evolved differences in gene expression did not overlap with those showing plastic differences in gene expression. However, in the genes that did overlap, there was counter-gradient variation such that plastic and evolved gene expression responses were in opposite directions in both species. Additionally, E. apleurogramma had higher levels of plasticity and evolved divergence in gene expression between field populations. We suggest that the higher level of plasticity and counter-gradient variation may have allowed rapid genetic adaptation in E. apleurogramma and facilitated colonization. This study shows how counter-gradient variation may impact the colonization of divergent oxygen environments.

Turbidity drives plasticity in the eyes and brains of an African cichlid.

Natural variation in environmental turbidity correlates with variation in the visual sensory system of many fishes, suggesting that turbidity may act as a strong selective agent on visual systems. Since many aquatic systems experience increased turbidity due to anthropogenic perturbations, it is important to understand the degree to which fish can respond to rapid shifts in their visual environment, and whether such responses can occur within the lifetime of an individual. We examined whether developmental exposure to turbidity (clear, <5 NTU; turbid, ∼9 NTU) influenced the size of morphological structures associated with vision in the African blue-lip cichlid Pseudocrenilabrus multicolor. Parental fish were collected from two sites (clear swamp, turbid river) in western Uganda. F1 broods from each population were split and reared under clear and turbid rearing treatments until maturity. We measured morphological traits associated with the visual sensory system (eye diameter, pupil diameter, axial length, brain mass, optic tectum volume) over the course of development. Age was significant in explaining variation in visual traits even when standardized for body size, suggesting an ontogenetic shift in the relative size of eyes and brains. When age groups were analyzed separately, young fish reared in turbid water grew larger eyes than fish reared in clear conditions. Population was important in the older age category, with swamp-origin fish having relatively larger eyes and optic lobes relative to river-origin fish. Plastic responses during development may be important for coping with a more variable visual environment associated with anthropogenically induced turbidity.

Inshore and offshore marine migration pathways of Atlantic salmon post-smolts from multiple rivers in Scotland, England, Northern Ireland, and Ireland.

The migratory behavior of Atlantic salmon (Salmo salar) post-smolts in coastal waters is poorly understood. In this collaborative study, 1914 smolts, from 25 rivers, in four countries were tagged with acoustic transmitters during a single seasonal migration. In total, 1105 post-smolts entered the marine study areas and 438 (39.6%) were detected on a network of 414 marine acoustic receivers and an autonomous underwater vehicle. Migration pathways (defined as the shortest distance between two detections) of up to 575 km and over 100 days at sea were described for all 25 populations. Post-smolts from different rivers, as well as individuals from the same river, used different pathways in coastal waters. Although difficult to generalize to all rivers, at least during the year of this study, no tagged post-smolts from rivers draining into the Irish Sea were detected entering the areas of sea between the Hebrides and mainland Scotland, which is associated with a high density of finfish aquaculture. An important outcome of this study is that a high proportion of post-smolts crossed through multiple legislative jurisdictions and boundaries during their migration. This study provides the basis for spatially explicit assessment of the impact risk of coastal pressures on salmon during their first migration to sea.

The weakly electric fish, Apteronotus albifrons, actively avoids experimentally induced hypoxia.

Anthropogenic environmental degradation has led to an increase in the frequency and prevalence of aquatic hypoxia (low dissolved oxygen concentration, DO), which may affect habitat quality for water-breathing fishes. The weakly electric black ghost knifefish, Apteronotus albifrons, is typically found in well-oxygenated freshwater habitats in South America. Using a shuttle-box design, we exposed juvenile A. albifrons to a stepwise decline in DO from normoxia (> 95% air saturation) to extreme hypoxia (10% air saturation) in one compartment and chronic normoxia in the other. On average, A. albifrons actively avoided the hypoxic compartment below 22% air saturation. Hypoxia avoidance was correlated with upregulated swimming activity. Following avoidance, fish regularly ventured back briefly into deep hypoxia. Hypoxia did not affect the frequency of their electric organ discharges. Our results show that A. albifrons is able to sense hypoxia at non-lethal levels and uses active avoidance to mitigate its adverse effects.

The effect of normoxia exposure on hypoxia tolerance and sensory sampling in a swamp-dwelling mormyrid fish.

Effects of energetic limitations on the performance of sensory systems are generally difficult to quantify. Weakly electric fishes provide an ideal model system to quantify the effects of metabolic stressors on sensory information acquisition, because they use an active-sensing strategy that permits easy measurement of the sensing effort. These fishes discharge an electric signal and sense perturbations of the resulting electric field. We used the mormyrid Petrocephalus degeni to quantify the relationship between routine metabolic rate and the rate of sensory sampling (rate of electric organ discharge, EOD) while under progressive hypoxia by quantifying the critical oxygen tension (PC-MR) and the critical electric organ discharge threshold (PC-EOD). PC-MR was significantly higher in fish acclimated to normoxia for over 40 days compared to animals tested within 1-5 days of capture from a hypoxic swamp, which suggests high costs of maintaining hypoxia tolerance; however, there was no acclimation effect on PC-EOD. All P. degeni reached their PC-EOD prior to their PC-MR. However, below the respective critical tension value, EOD rate decreased more gradually than the metabolic rate suggesting that the fish were increasing the proportion of their energy budget allocated to acquiring sensory information as dissolved-oxygen levels dropped. Trade-offs between sensory sampling and other physiological functions are also suggested by the increase in routine EOD rate with long-term normoxia acclimation, in contrast to metabolic rate, which showed no significant changes. These results highlight the relationship between sensory sampling and metabolic rate in response to progressive hypoxia and the plasticity of hypoxia tolerance.

Divergence in aerobic scope and thermal tolerance is related to local thermal regime in two populations of introduced Nile perch (Lates niloticus).

We tested whether thermal tolerance and aerobic performance differed between two populations of Nile perch (Lates niloticus) originating from the same source population six decades after their introduction into two lakes in the Lake Victoria basin in East Africa. We used short-term acclimation of juvenile fish to a range of temperatures from ambient to +6°C, and performed critical thermal maximum (CTmax ) and respirometry tests to measure upper thermal tolerance, resting and maximum metabolic rates, and aerobic scope (AS). Across acclimation temperatures, Nile perch from the cooler lake (Lake Nabugabo, Uganda) tended to have lower thermal tolerance (i.e., CTmax ) and lower aerobic performance (i.e., AS) than Nile perch from the warmer waters of Lake Victoria (Bugonga region, Uganda). Effects of temperature acclimation were more pronounced in the Lake Victoria population, with the Lake Nabugabo fish showing less thermal plasticity in most metabolic traits. Our results suggest phenotypic divergence in thermal tolerance between these two introduced populations in a direction consistent with an adaptive response to local thermal regimes.

Effects of hypoxia on aerobic metabolism and active electrosensory acquisition in the African weakly electric fish Marcusenius victoriae.

Environmental hypoxia has effected numerous and well-documented anatomical, physiological and behavioural adaptations in fishes. Comparatively little is known about hypoxia's impacts on sensing because it is difficult to quantify sensory acquisition in vivo. Weakly electric fishes, however, rely heavily on an easily-measurable sensory modality-active electric sensing-whereby individuals emit and detect electric organ discharges (EODs). In this study, hypoxia tolerance of a mormyrid weakly electric fish, Marcusenius victoriae, was assessed by examining both its metabolic and EOD rates using a critical threshold (pcrit ) paradigm. The routine metabolic rate was 1.42 mg O2 h-1 , and the associated critical oxygen tension was 14.34 mmHg. Routine EOD rate was 5.68 Hz with an associated critical tension of 15.14 mmHg. These metabolic indicators of hypoxia tolerance measured in this study were consistent with those in previous studies on M. victoriae and other weakly electric fishes. Furthermore, our results suggest that some aerobic processes may be reduced in favour of maintaining the EOD rate under extreme hypoxia. These findings underscore the importance of the active electrosensory modality to these hypoxia-tolerant fish.

Understanding Maladaptation by Uniting Ecological and Evolutionary Perspectives.

Evolutionary biologists have long trained their sights on adaptation, focusing on the power of natural selection to produce relative fitness advantages while often ignoring changes in absolute fitness. Ecologists generally have taken a different tack, focusing on changes in abundance and ranges that reflect absolute fitness while often ignoring relative fitness. Uniting these perspectives, we articulate various causes of relative and absolute maladaptation and review numerous examples of their occurrence. This review indicates that maladaptation is reasonably common from both perspectives, yet often in contrasting ways. That is, maladaptation can appear strong from a relative fitness perspective, yet populations can be growing in abundance. Conversely, resident individuals can appear locally adapted (relative to nonresident individuals) yet be declining in abundance. Understanding and interpreting these disconnects between relative and absolute maladaptation, as well as the cases of agreement, is increasingly critical in the face of accelerating human-mediated environmental change. We therefore present a framework for studying maladaptation, focusing in particular on the relationship between absolute and relative fitness, thereby drawing together evolutionary and ecological perspectives. The unification of these ecological and evolutionary perspectives has the potential to bring together previously disjunct research areas while addressing key conceptual issues and specific practical problems.

Cardiac plasticity influences aerobic performance and thermal tolerance in a tropical, freshwater fish at elevated temperatures.

Fishes faced with novel thermal conditions often modify physiological functioning to compensate for elevated temperatures. This physiological plasticity (thermal acclimation) has been shown to improve metabolic performance and extend thermal limits in many species. Adjustments in cardiorespiratory function are often invoked as mechanisms underlying thermal plasticity because limitations in oxygen supply have been predicted to define thermal optima in fishes; however, few studies have explicitly linked cardiorespiratory plasticity to metabolic compensation. Here, we quantified thermal acclimation capacity in the commercially harvested Nile perch (Lates niloticus) of East Africa, and investigated mechanisms underlying observed changes. We reared juvenile Nile perch for 3 months under two temperature regimes, and then measured a series of metabolic traits (e.g. aerobic scope) and critical thermal maximum (CTmax) upon acute exposure to a range of experimental temperatures. We also measured morphological traits of heart ventricles, gills and brains to identify potential mechanisms for compensation. We found that long-term (3 month) exposure to elevated temperature induced compensation in upper thermal tolerance (CTmax) and metabolic performance (standard and maximum metabolic rate, and aerobic scope), and induced cardiac remodeling in Nile perch. Furthermore, variation in heart morphology influenced variations in metabolic function and thermal tolerance. These results indicate that plastic changes enacted over longer exposures lead to differences in metabolic flexibility when organisms are acutely exposed to temperature variation. Furthermore, we established functional links between cardiac plasticity, metabolic performance and thermal tolerance, providing evidence that plasticity in cardiac capacity may be one mechanism for coping with climate change.

Effects of hypoxia on swimming and sensing in a weakly electric fish.

Low dissolved oxygen (hypoxia) can severely limit fish performance, especially aerobically expensive behaviours including swimming and acquisition of sensory information. Fishes can reduce oxygen requirements by altering these behaviours under hypoxia, but the underlying mechanisms can be difficult to quantify. We used a weakly electric fish as a model system to explore potential effects of hypoxia on swim performance and sensory information acquisition, which enabled us to non-invasively record electric signalling activity used for active acquisition of sensory information during swimming. To quantify potential effects of hypoxia, we measured critical swim speed (Ucrit) and concurrent electric signalling activity under high- and low-dissolved oxygen concentrations in a hypoxia-tolerant African mormyrid fish, Marcusenius victoriae Fish were maintained under normoxia for 6 months prior to experimental treatments, and then acclimated for 8 weeks to normoxia or hypoxia and tested under both conditions (acute: 4 h exposure). Acute hypoxia exposure resulted in a significant reduction in both Ucrit and electric signalling activity in fish not acclimated to hypoxia. However, individuals acclimated to chronic hypoxia were characterized by a higher Ucrit under both hypoxia and normoxia than fish acclimated to normoxia. Following a 6 month re-introduction to normoxia, hypoxia-acclimated individuals still showed increased performance under acute hypoxic test conditions, but not under normoxia. Our results highlight the detrimental effects of hypoxia on aerobic swim performance and sensory information acquisition, and the ability of fish to heighten aerobic performance through acclimation processes that can still influence performance even months after initial exposure.

Evolution of a Strategy for the Enantioselective Total Synthesis of (+)-Psiguadial B.

(+)-Psiguadial B is a diformyl phloroglucinol meroterpenoid that exhibits antiproliferative activity against the HepG2 human hepatoma cancer cell line. This full account details the evolution of a strategy that culminated in the first enantioselective total synthesis of (+)-psiguadial B. A key feature of the synthesis is the construction of the trans-cyclobutane motif by a Wolff rearrangement with in situ catalytic, asymmetric trapping of the ketene. An investigation of the substrate scope of this method to prepare enantioenriched 8-aminoquinolinamides is disclosed. Three routes toward (+)-psiguadial B were evaluated that featured the following key steps: (1) an ortho-quinone methide hetero-Diels-Alder cycloaddition to prepare the chroman framework, (2) a Prins cyclization to form the bridging bicyclo[4.3.1]decane system, and (3) a modified Norrish-Yang cyclization to generate the chroman. Ultimately, the successful strategy employed a ring-closing metathesis to form the seven-membered ring and an intramolecular O-arylation reaction to complete the polycyclic framework of the natural product.

Elevated temperature and acclimation time affect metabolic performance in the heavily exploited Nile perch of Lake Victoria.

Increasing water temperatures owing to anthropogenic climate change are predicted to negatively impact the aerobic metabolic performance of aquatic ectotherms. Specifically, it has been hypothesized that thermal increases result in reductions in aerobic scope (AS), which lead to decreases in energy available for essential fitness and performance functions. Consequences of warming are anticipated to be especially severe for warm-adapted tropical species as they are thought to have narrow thermal windows and limited plasticity for coping with elevated temperatures. In this study we test how predicted warming may affect the aerobic performance of Nile perch (Lates niloticus), a commercially harvested fish species in the Lake Victoria basin of East Africa. We measured critical thermal maxima (CTmax) and key metabolic variables such as AS and excess post-exercise oxygen consumption (EPOC) across a range of temperatures, and compared responses between acute (3-day) exposures and 3-week acclimations. CTmax increased with acclimation temperature; however, 3-week-acclimated fish had higher overall CTmax than acutely exposed individuals. Nile perch also showed the capacity to increase or maintain high AS even at temperatures well beyond their current range; however, acclimated Nile perch had lower AS compared with acutely exposed fish. These changes were accompanied by lower EPOC, suggesting that drops in AS may reflect improved energy utilization after acclimation, a finding that is supported by improvements in growth at high temperatures over the acclimation period. Overall, the results challenge predictions that tropical species have limited thermal plasticity, and that high temperatures will be detrimental because of limitations in AS.

Hypoxia acclimation increases novelty response strength during fast-starts in the African mormyrid, Marcusenius victoriae.

Many fishes perform quick and sudden swimming maneuvers known as fast-starts to escape when threatened. In pulse-type weakly electric fishes these responses are accompanied by transient increases in the rate of electric signal production known as novelty responses. While novelty responses may increase an individual's information about their surroundings, they are aerobically powered and may come at a high energetic cost when compared to fast-starts, which rely primarily on anaerobic muscle. The juxtaposition between two key aspects of fast-starts in these fishes - the aerobic novelty response and the anaerobic swimming performance - makes them an interesting model for studying effects of hypoxia on escape performance and sensory information acquisition. We acclimated the hypoxia-tolerant African mormyrid Marcusenius victoriae to either high or low dissolved oxygen (DO) levels for 8weeks, after which fast-starts and novelty responses were quantified under both high (normoxic) and low-DO (hypoxic) test conditions. Hypoxia-acclimated fish exhibited higher maximum curvature than normoxia-acclimated fish. Displacement of normoxia-acclimated fish was not reduced under acute hypoxic test conditions. Novelty responses were given upon each startle, whether or not the fish performed a fast-start; however, novelty responses associated with fast-starts were significantly stronger than those without, suggesting a functional link between fast-start initiation and the motor control of the novelty response. Overall, hypoxia-acclimated individuals produced significantly stronger novelty responses during fast-starts. We suggest that increased novelty response strength in hypoxia-acclimated fish corresponds to an increased rate of sensory sampling, which may compensate for potential negative effects of hypoxia on higher-level processing.

Enantioselective Total Synthesis of (+)-Psiguadial B.

The first enantioselective total synthesis of the cytotoxic natural product (+)-psiguadial B is reported. Key features of the synthesis include (1) the enantioselective preparation of a key cyclobutane intermediate by a tandem Wolff rearrangement/asymmetric ketene addition, (2) a directed C(sp(3))-H alkenylation reaction to strategically forge the C1-C2 bond, and (3) a ring-closing metathesis to build the bridging bicyclo[4.3.1]decane terpene framework.

Effects of thermal increase on aerobic capacity and swim performance in a tropical inland fish.

Rising water temperature associated with climate change is increasingly recognized as a potential stressor for aquatic organisms, particularly for tropical ectotherms that are predicted to have narrow thermal windows relative to temperate ectotherms. We used intermittent flow resting and swimming respirometry to test for effects of temperature increase on aerobic capacity and swim performance in the widespread African cichlid Pseudocrenilabrus multicolor victoriae, acclimated for a week to a range of temperatures (2°C increments) between 24 and 34°C. Standard metabolic rate (SMR) increased between 24 and 32°C, but fell sharply at 34°C, suggesting either an acclimatory reorganization of metabolism or metabolic rate depression. Maximum metabolic rate (MMR) was elevated at 28 and 30°C relative to 24°C. Aerobic scope (AS) increased between 24 and 28°C, then declined to a level comparable to 24°C, but increased dramatically 34°C, the latter driven by the drop in SMR in the warmest treatment. Critical swim speed (Ucrit) was highest at intermediate temperature treatments, and was positively related to AS between 24 and 32°C; however, at 34°C, the increase in AS did not correspond to an increase in Ucrit, suggesting a performance cost at the highest temperature.

Saliva management options for difficult-to-wean people with tracheostomy following severe acquired brain injury (ABI): A review of the literature.

PRIMARY OBJECTIVE: To evaluate the evidence on saliva management options in those people who have a tracheostomy in situ following an acquired brain injury and to ascertain whether any of these treatments may facilitate tracheostomy decannulation. METHODS: The search was conducted on Medline, Embase, Cinahl and Central databases since 1990 and the evidence has been critiqued and summarized. Saliva management options were identified and analysed to see whether they had evidence or clinical support for the population. MAIN OUTCOMES AND RESULTS: There is a paucity of evidence in this area and clinical decision-making requires evidence from other populations. Saliva management issues in this population are most likely to be related to dysphagia. Treatment options include behavioural/compensatory therapies which should be tried in all cases, with adjunct pharmaceutical therapies or surgical options to reduce saliva volume as clinically appropriate. CONCLUSION AND FUTURE DIRECTIONS: This group of people is complex and requires a multi-disciplinary team to guide decision-making. High quality control studies looking at the effectiveness of dysphagia therapy and guidelines regarding botulinum toxin injections are recommended.

Effects of acute hypoxia exposure and acclimation on the thermal tolerance of an imperiled Canadian minnow.

Elevated water temperatures and low dissolved oxygen (hypoxia) are pervasive stressors in aquatic systems that can be exacerbated by climate change and anthropogenic activities, and there is growing interest in their interactive effects. To explore this interaction, we quantified the effects of acute and long-term hypoxia exposure on the critical thermal maximum (CTmax) of Redside Dace (Clinostomus elongatus), a small-bodied freshwater minnow with sparse populations in the Great Lakes Basin of Canada and designated as Endangered under Canada's Species at Risk Act. Fish were held at 18°C and acclimated to four levels of dissolved oxygen (>90%, 60%, 40%, and 20% air saturation). CTmax was measured after 2 and 10 weeks of acclimation and after 3.5 weeks of reoxygenation, and agitation behavior was quantified during CTmax trials. Aquatic surface respiration behavior was also quantified at 14 weeks of acclimation to oxygen treatments. Acute hypoxia exposure decreased CTmax in fish acclimated to normoxia (>90% air saturation), but acclimation to hypoxia reduced this effect. There was no effect of acclimation oxygen level on CTmax when measured in normoxia, and there was no effect of exposure time to hypoxia on CTmax. Residual effects of hypoxia acclimation on CTmax were not seen after reoxygenation. Agitation behavior varied greatly among individuals and was not affected by oxygen conditions. Fish performed aquatic surface respiration with low frequency, but performed it earlier when acclimated to higher levels of oxygen. Overall, this work sheds light on the vulnerability of fish experiencing acute hypoxia and heat waves concurrently.

Persisting in papyrus: size, oxidative stress, and fitness in freshwater organisms adapted to sustained hypoxia.

Aquatic hypoxia is generally viewed as stressful for aerobic organisms. However, hypoxia may also benefit organisms by decreasing cellular stress, particularly that related to free radicals. Thus, an ideal habitat may have the minimum O2 necessary to both sustain aerobic metabolism and reduce the need to scavenge free radicals and repair free radical damage. The ability of aquatic organisms to sustain aerobic metabolism relates in part to the ability to maximize gas diffusion, which can be facilitated by small body size when O2 uptake occurs across the body surface, by a large gill surface area, or by the ability to use atmospheric air. We use water-breathing organisms in chronically hypoxic papyrus (Cyperus papyrus) swamps of East Africa to test the hypothesis that cellular-level benefits of hypoxia may translate into increased fitness, especially for small organisms. A review of recent studies of fingernail clams (Sphaerium sp.) shows that clams living in sustained hypoxia have minimized oxidative stress and that these cellular-level benefits may lead to increased fitness. We suggest that organisms in the extreme conditions in the papyrus swamps provide a unique opportunity to challenge the conventional classification of hypoxic habitats as 'stressful' and normoxic habitats as 'optimal.'

Cycles of fusion and fission enabled rapid parallel adaptive radiations in African cichlids.

Although some lineages of animals and plants have made impressive adaptive radiations when provided with ecological opportunity, the propensities to radiate vary profoundly among lineages for unknown reasons. In Africa's Lake Victoria region, one cichlid lineage radiated in every lake, with the largest radiation taking place in a lake less than 16,000 years old. We show that all of its ecological guilds evolved in situ. Cycles of lineage fusion through admixture and lineage fission through speciation characterize the history of the radiation. It was jump-started when several swamp-dwelling refugial populations, each of which were of older hybrid descent, met in the newly forming lake, where they fused into a single population, resuspending old admixture variation. Each population contributed a different set of ancient alleles from which a new adaptive radiation assembled in record time, involving additional fusion-fission cycles. We argue that repeated fusion-fission cycles in the history of a lineage make adaptive radiation fast and predictable.

Holding water steroid hormones in the African cichlid fish Pseudocrenilabrus multicolor victoriae.

Measuring hormone levels multiple times on the same individual across different life stages or treatments can facilitate our understanding of hormonal regulation of physiological and behavioral events. The conventional method of hormone measurement requires blood sampling, which is potentially lethal to small individuals. In fishes, there is an alternative non-invasive method of hormone measurement using the release of hormones across gill membranes from blood into holding water. Validation of this method is required to evaluate its application value to different species. In the present study we used the maternal mouth-brooding African cichlid fish, Pseudocrenilabrus multicolor victoriae to (i) investigate whether handling involved in using the holding water technique is a stressor by measuring excreted cortisol in male and female P. multicolor handled one or multiple times, (ii) validate use of this technique by quantifying the relationship between plasma and holding water measures of sex hormones in male P. multicolor, and (iii) demonstrate the biological relevance of this technique using excreted levels of sex hormones in female P. multicolor across different reproductive stages. Excreted cortisol and estradiol levels did not differ between fish handled one or more times, suggesting that the repeated sampling approach over the breeding cycle that we propose to use does not affect the excreted level of the hormone of interest. Measurements from plasma and holding water samples were positively related for both testosterone and estradiol, indicating that the holding water technique is a reliable index of plasma hormone levels. Excreted sex hormone levels varied with reproductive state, suggesting that the technique is a useful, non-invasive measure of sex hormone levels in P. multicolor.