We are grateful to anonymous reviewers for comments and suggestions that greatly improved this manuscript. We thank R. Bowes, R. Carreno, and T. Panhuis for assistance in collecting the fish. We also thank M. Lee for assistance with male preference trials. We are grateful to the Ohio Wesleyan University Department of Zoology for helpful advice and suggestions throughout this study and Arizona State University (McGraw Lab) for software advice.

Tutti gli esperimenti sono stati condotti con l&#39;approvazione del Comitato di Cura e uso istituzionale degli animali Ohio Wesleyan University. 1. Registrazione UV riflettanza di pesce utilizzando uno spettrofotometro <ol><li> Calibrare uno spettrofotometro e fonte di luce con uno standard bianco nota nella gamma di lunghezze d&#39;onda da misurare secondo le raccomandazioni dello strumento o software. </li><li> Anesthetize pesci mettendo in una soluzione allo 0,5% di etil 3-amminobenzoato metano sale di acido solfonico (MS-222) tamponato con un&#39;eguale quantità di bicarbonato di sodio fino pesci non risponde. Una volta non risponde, subito mettere il pesce su un nero, sfondo non riflettente. Nota: MS-222 è potenzialmente tossici, e guanti devono essere indossati in ogni momento. </li><li> Prendere tutte le misurazioni in una stanza buia per minimizzare luce in eccesso oltre quello della sorgente luminosa spettrometro. </li><li> riflettanza misura utilizzando una sonda a fibre ottiche tenuta contro il corpo dell&#39;organismo. Renderesicuri per formare un angolo di 45 ° coerente tra le fibre ottiche e il corpo, come è incorporato in alcuni disegni sonda. Nota: Per coerenza misura, può essere desiderabile media su più scansioni (2 - 5) per regione di body. </li><li> registrare i dati di riflettanza con adeguato software di analisi di riflettanza. Nota: il software utilizzato qui è stato impostato su: all&#39;integrazione tempo = 2.000 sec, scansioni media = 2, e pixel levigante = 5. I dati sono stati copiato e incollato ad un foglio di calcolo. </li><li> Ripetere su più zone del corpo per determinare la presenza o assenza di riflettanza UV e la gamma di variazione fenotipica di tutti gli organismi. Nota: possono essere necessarie scansioni preliminari per determinare quale, se del caso, le zone del corpo dimostrano costantemente le marcature UV. </li><li> Riportare il pesce per un acquario partecipazione ben ossigenata fino alla completa guarigione, come evidenziato da normali battiti opercolo e comportamenti di nuoto. Nota: Un protectant commerciale per ripristinare il rivestimento mucoso al corpo può essere aggiunto per aiutare nel recupero pelle. </li><li> rappresentantemangiare su più individui, tra cui entrambi i sessi, se desiderato. </li><li> Sul foglio di calcolo in cui tutti i dati sono stati copiati, i dati del diagramma utilizzando un grafico a dispersione, con riflessione sulla Y e lunghezza d&#39;onda sulla X-asse. picchi di ampiezza della gamma UV (300-400 nm) indicano la presenza di riflettanza UV sul pesce. </li></ol> 2. Prove di accoppiamento dicotomiche NOTA: effettuare osservazioni di controllo, in cui i pesci focale, filtri UV e filtri opachi sono in atto. In questi studi, quando vengono rimossi i filtri opachi, pesce focale sarà esposto a parti del serbatoio visualizzazione presenza e in assenza di luce UV, ma privi potenziali partner di accoppiamento. Per i dettagli sulla validità dei test a scelta dicotomici in questo esperimento vedere Palmer e Hankison (2015) 10, anche se nota che questi test hanno anche svantaggi 11. <ol><li> Utilizzare gli individui sessualmente ricettive come organismi focali. Utilizzare femminile P. latipinna &lt;48 ore dopo il parto 5 unnd maschi isolati per almeno 24 ore prima del test per migliorare la probabilità di motivazione sessuale. </li><li> Posizionare un singolo individuo focale in una zona neutra adiacente a due scomparti di scelta. Per P. latipinna, utilizzare un test acquario 75,7 L (30,5 cm x 30,5 cm x 75 cm; Figura 1) diviso in tre sezioni da setti plexiglas trasparenti, formando una zona centrale per l&#39;individuo focale affiancato da due compartimenti scelta di dimensioni uguali. Assicurarsi che l&#39;acqua o olfattivi segnali sono condivisi tra le singole sezioni del serbatoio. Ripetere per ogni individuo di essere osservato. </li></ol><img alt="Figura 1" src="/files/ftp_upload/54453/54453fig1.jpg" /> Figura 1. sperimentale Acquario installazione di un acquario rettangolare divisa in tre sezioni:. Un&#39;area centrale che ha tenuto i due compartimenti finali che hanno tenuto le coppie singolo oggetto focale e (la figura mostra una donna a metà con i maschi degli oggetti alle estremità). fil intercambiabileTERS che o riflettanza UV bloccato o consentito potrebbe essere collocato sopra i divisori in vetro che dividono le sezioni maschili e femminili dell&#39;acquario. Questa cifra viene modificato da Palmer e Hankison (2015) 10. <a href="http://ecsource.jove.com/files/ftp_upload/54453/54453fig1large.jpg" target="_blank">Si prega di cliccare qui per vedere una versione più grande di questa figura.</a> <ol start="3"><li> In condizioni di luce a spettro completo (illuminazione il cui spettro si estende nella gamma UV-A), isolare un potenziale partner di accoppiamento in ciascun compartimento fine (vano scelta) dell&#39;acquario di prova. Assicurarsi che potenziali partner di accoppiamento assomigliano più strettamente possibile in dimensioni, colorazione e visualizzazione caratteristiche, in modo che la differenza principale tra le due parti di accoppiamento isolati è se appaiono dietro un filtro che blocca o trasmette la luce UV. NOTA: Può essere preferibile registrare riflettanza nello spettro visibile per consentire la migliore corrispondenza dei colori possibile, alanche se qui abbinamento è stato fatto da occhio. </li><li> Ambientarsi individui sotto isolamento visivo (filtri opachi) per 15 min. Assicurarsi che UV + e - filtri sono in atto in questo momento. A seguito di acclimatazione, rimuovere i filtri opachi. Per garantire che le polarizzazioni collaterali non sono presenti (preferenza latente in un lato della vasca indipendentemente oggetto), assegnare casualmente l&#39;+ UV e UV - lati del serbatoio per ogni ogni prova. </li><li> Registrare la quantità di tempo che l&#39;individuo focale spende all&#39;interno delle zone di preferenza specifici (zone vicino i comparti di scelta, in questo caso di circa due lunghezze corpo di serie) utilizzando cronometri. Per pesce femmina, questa è la quantità di tempo che trascorre vicino ciascuno dei due maschi, e viceversa per il pesce focale maschio. NOTA: software del registratore evento può anche essere usato per la registrazione. Un quarto d&#39;ora le prove sono stati utilizzati in questo esperimento e pesce che non ha visitato ciascuna zona preferenza almeno una volta sono stati esclusi dai risultati come non risponde. </li><li> Registrare e analizzare il tempo che i pesci passano vicino a vuoto UV + e UV - scomparti per assicurare che non vi è una tendenza intrinseca verso un particolare ambiente di illuminazione che possono influenzare la scelta accoppiamento del pesce. Utilizzare appaiati t-test per determinare se gli individui preferiscono una particolare condizione UV. </li></ol>

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<li>Palmer, M. S., Hankison, S. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Use+of+ultraviolet+cues+in+female+mate+preference+in+the+sailfin+molly%2C+Poecilia+latipinna%5BTitle%5D)+AND+%22Acta+Ethol%22%5BJournal%5D)">Use of ultraviolet cues in female mate preference in the sailfin molly, Poecilia latipinna.</a> Acta Ethol. 18 (2), 153-160 (2014).</li>
<li>Wagner, W. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Measuring+female+mating+preferences%5BTitle%5D)+AND+%22Anim.+Behav%22%5BJournal%5D)">Measuring female mating preferences.</a> Anim. Behav. 55 (4), 1029-1042 (1998).</li>
<li>Rémy, A., Grégoire, A., Perret, P., Doutrelant, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Mediating+male-male+interactions%3A+the+role+of+the+UV+blue+crest+coloration+in+blue+tits%5BTitle%5D)+AND+%22Behav.+Ecol.+Sociobiol%22%5BJournal%5D)">Mediating male-male interactions: the role of the UV blue crest coloration in blue tits.</a> Behav. Ecol. Sociobiol. 64 (11), 1839-1847 (2010).</li>
<li>Guillermo-Ferreira, R., Therézio, E. M., Gehlen, M. H., Bispo, P. C., Marletta, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((The+Role+of+Wing+Pigmentation%2C+UV+and+Fluorescence+as+Signals+in+a+Neotropical+Damselfly%5BTitle%5D)+AND+%22J.+Insect+Behav%22%5BJournal%5D)">The Role of Wing Pigmentation, UV and Fluorescence as Signals in a Neotropical Damselfly.</a> J. Insect Behav. 27 (1), 67-80 (2013).</li>
<li>Ord, T. J., Stamps, J. A., Losos, J. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Convergent+evolution+in+the+territorial+communication+of+a+classic+adaptive+radiation%3A+Caribbean+Anolis+lizards%5BTitle%5D)+AND+%22Anim.+Behav%22%5BJournal%5D)">Convergent evolution in the territorial communication of a classic adaptive radiation: Caribbean Anolis lizards.</a> Anim. Behav. 85 (6), 1415-1426 (2013).</li>
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<li>Modarressie, R., Rick, I. P., Bakker, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((UV+matters+in+shoaling+decisions%5BTitle%5D)+AND+%22Proc.+Biol.+Sci%22%5BJournal%5D)">UV matters in shoaling decisions.</a> Proc. Biol. Sci. 273 (1588), 849-854 (2006).</li>
<li>Lim, M. L. M., Li, J., Li, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Effect+of+UV-reflecting+markings+on+female+mate-choice+decisions+in+Cosmophasis+umbratica%2C+a+jumping+spider+from+Singapore%5BTitle%5D)+AND+%22Behav.+Ecol%22%5BJournal%5D)">Effect of UV-reflecting markings on female mate-choice decisions in Cosmophasis umbratica, a jumping spider from Singapore.</a> Behav. Ecol. 19, 61-66 (2008).</li>
<li>Rick, I. P., Bakker, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Color+signaling+in+conspicuous+red+sticklebacks%3A+do+ultraviolet+signals+surpass+others%3F%5BTitle%5D)+AND+%22BMC+Evol.+Biol%22%5BJournal%5D)">Color signaling in conspicuous red sticklebacks: do ultraviolet signals surpass others?</a> BMC Evol. Biol. 8, 189(2008).</li>
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</ol>

The authors have nothing to disclose.

Spettrofotometria è riuscito a identificare le marcature UV su P. latipinna. Entrambi i sessi di P. latipinna possiedono marcature UV lungo i loro lati. Inoltre, alcuni maschi avevano marcature UV sulle loro pinne dorsali, tratti precedentemente trovato per essere importante nelle preferenze di accoppiamento femmina 7. Si consiglia di utilizzare spettrofotometria UV come meccanismo per rilevare la presenza di marcature UV. Ulteriori test in grado di determinare ...

Capire gli indizi e segnali utilizzati nelle interazioni sociali animali ci permette di comprendere la variazione fenotipica sia all&#39;interno che tra le specie. Questa variazione gioca un ruolo importante nei processi evolutivi come divergenza popolazione, selezione sessuale, e speciazione. Spesso, tuttavia, i ricercatori sono limitati a esplorare i segnali più evidenti di sistemi sensoriali umani, in particolare quelli nei regni visivi o uditivi. L&#39;uso della spettrofotometria, però, ci permette di espandere le nostre indagini al di là dello spettro visibile umana e in lunghezze d&#39;onda che possono essere importanti nelle interazioni sociali in altre specie. In particolare, la comunicazione a corto raggio offerta da raggi ultravioletti (UV; 300-400 nm) sensibilità ha il potenziale per essere molto vantaggiosa durante scelta del compagno 1. Molti predatori visivamente-caccia di uccelli e pesci, per esempio, sono in grado di rilevare le radiazioni UV. Nei sistemi in cui i maschi mostrano elaboratoalle femmine, questi maschi potrebbero ridurre il rischio di predazione, pur mantenendo la loro capacità di attrarre i compagni sfruttando lo spettro UV, piuttosto che sviluppare spunti rilevabile nello spettro visibile 2,3 .. Se uno non riesce a prendere in considerazione la possibilità che gli organismi comunicano con ogni altri che utilizzano questi &quot;canali di comunicazione privati&quot;, i conducenti significative di comportamento ed evoluzione possono perdere. Questo protocollo descrive un&#39;indagine l&#39;uso di segnali UV per la scelta del compagno nel sailfin Molly, Poecilia latipinna, un pesce che ha poligama non precedentemente nota capacità di rilevare UV o utilizzare le marcature UV. Questa specie di pesci ha una stretta vicinanza filogenetica ad altri pesci live-cuscinetto sensibili ai raggi UV-4 e vi sono prove che microspectroscopic P. latipinna, insieme ad altre specie molly come P. mexicana e P. formosa, in possesso di una classe di coni (fotorecettori responsabili della covisione Lor) che sono più sensibili per lunghezze d&#39;onda UV 5. In questa specie dimorfismo sessuale, la scelta femminile ha svolto un ruolo forte per l&#39;evoluzione della brillantemente pinne colorati e ingranditi dei maschi 6-9. Questa metodologia ci permette di esplorare se UV è un mezzo ulteriore per cui le femmine valutare la qualità maschile. La rilevazione e la misurazione delle marcature UV su P. latipinna utilizzando uno spettrofotometro con una sonda a fibre ottiche è descritto qui. Inoltre, se ricettive Molly femminili differenziale associano con i maschi visti attraverso un filtro ottico che trasmettono la luce a spettro completo, tra cui UV-A ([UV +], 320-700 nm) e maschi visti attraverso un filtro UV-bloccante ([UV]; 400 - 700 nm) è discusso. Questo metodo ha ampie applicazioni per scoprire i modelli di sensibilità ai raggi UV e colore in pesci e altri organismi, consentendo la ricerca in una serie di domande che coinvolgono UV e il suo ruolo nel comportamento.

<table><tbody><tr><td>Spectrophotometer, P1000</td><td>Ocean Optics</td><td></td><td>newer models are availabe</td></tr><tr><td>DT 1000 xenon UV light source</td><td>Ocean Optics</td><td></td><td>newer models are availabe</td></tr><tr><td>Ocean Optics Overture Software</td><td>Ocean Optics</td><td></td><td>newer software is available</td></tr><tr><td>R200-Angle-UV bifurcated fiber-optic probe (Guided Wave)</td><td>Ocean Optics</td><td></td><td>newer models are availabe</td></tr><tr><td>Certified reflectance standard, white</td><td>labsphere</td><td></td><td></td></tr><tr><td>75.7 L Aquarium, divided</td><td>Experimental Builder</td><td></td><td></td></tr><tr><td>Full Spectrum Bulb</td><td>Nature&#039;s Sunlight</td><td></td><td></td></tr><tr><td>UV blocking sheet</td><td>GAM UV Sheet</td><td></td><td></td></tr></tbody></table>

assays to detect uv-reflecting structures and determine their importance in mate preference using the sailfin molly poecilia latipinna

Molti organismi utilizzano spunti e segnali al di là di sensibilità umana durante le interazioni sociali. E &#39;importante prendere in considerazione come gli organismi percepiscono i loro mondi quando si cerca di capire il loro comportamento e l&#39;ecologia. La sensibilità alla spettro ultravioletto (UV; 300-400 nm) si trova in tutta generi multiplo di uccelli, pesci, rettili, anfibi, e persino mammiferi. Questo protocollo descrive una tecnica per l&#39;esame organismi per la presenza di strutture UV-riflettente ed un metodo per verificare se questi segnali sono usati come segnali sociali nel contesto della scelta del compagno. Uno spettrofotometro è utilizzato per rilevare la presenza di riflettanza UV e variazione di intensità riflettente tra individui e sessi. Un esempio di questa tecnica è presentato in cui un test a scelta dicotomica compagno espone gli individui sessualmente ricettive per gli individui di sesso opposto il cui aspetto visivo può essere manipolato dai filtri che o trasmettono a spettro completo o il blocco UV lunghezze d&#39;onda. Questosistema ha consentito per la determinazione che femminili, ma non di sesso maschile, Molly Sailfin (Poecilia latipinna) stavano usando le marcature UV come parte delle loro decisioni di accoppiamento. Questi tipi di studi servono a espandere la nostra conoscenza della gamma di organismi che utilizzano raggi UV e fornire informazioni su come UV gioca un ruolo nella loro vita.

<html> Le figure 1 e 2 mostrano l&#39;accoppiamento preferenza istituito acquario e siti di misura UV per i nostri esperimenti. Misurare riflettanza UV ha permesso per la determinazione che P. latipinna possiedono caratteristiche UV, in particolare lungo i lati dei loro corpi (Figura 3), in aggiunta alla variazione individuale in questi tratti. Una volta tratti UV sono...

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Assays to Detect UV-reflecting Structures and Determine their Importance in Mate Preference using the Sailfin Molly Poecilia latipinna

Questo protocollo descrive l&#39;uso di spettrofotometria di rilevare strutture ultravioletti che riflette sugli organismi (in questo esempio, il sailfin Molly Poecilia latipinna) e descrive test a scelta dicotomiche per i pesci che permettono inferenze da apportare sul ruolo di spunti ultravioletti durante la selezione compagno.

Metodi per la determinazione strutture UV-riflettenti e determinare la loro importanza nella Mate Preferenze utilizzando il Sailfin Molly<em&gt; Poecilia latipinna</em

Many organisms use cues and signals beyond human sensitivity during social interactions. It is important to take into account how organisms perceive their ...

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Assays to Detect UV-reflecting Structures and Determine their Importance in Mate Preference using the Sailfin Molly Poecilia latipinna

6.5K Views.  Ohio Wesleyan University.  Questo protocollo descrive l'uso di spettrofotometria di rilevare strutture ultravioletti che riflette sugli organismi (in questo esempio, il sailfin Molly Poecilia latipinna) e descrive test a scelta dicotomiche per i pesci che permettono inferenze da apportare sul ruolo di spunti ultravioletti durante la selezione compagno. 

Video: Assays to Detect UV-reflecting Structures and Determine their Importance in Mate Preference using the Sailfin Molly Poecilia latipinna

Multimodal Optical Microscopy Methods Reveal Polyp Tissue Morphology and Structure in Caribbean Reef Building Corals

An integrated suite of imaging techniques has been applied to determine the three-dimensional (3D) morphology and cellular structure of polyp tissues comprising the Caribbean reef building corals Montastraeaannularis and M. faveolata. These approaches include fluorescence microscopy (FM), serial block face imaging (SBFI), and two-photon confocal laser scanning microscopy (TPLSM). SBFI provides deep tissue imaging after physical sectioning; it details the tissue surface texture and 3D visualization to tissue depths of more than 2 mm. Complementary FM and TPLSM yield ultra-high resolution images of tissue cellular structure. Results have: (1) identified previously unreported lobate tissue morphologies on the outer wall of individual coral polyps&#160;and (2) created the first surface maps of the 3D distribution and tissue density of chromatophores and algae-like dinoflagellate zooxanthellae endosymbionts. Spectral absorption peaks of 500 nm and 675 nm, respectively, suggest that M. annularis and M. faveolata contain similar types of chlorophyll and chromatophores. However, M. annularis and M. faveolata exhibit significant differences in the tissue density and 3D distribution of these key cellular components. This study focusing on imaging methods indicates that SBFI is extremely useful for analysis of large mm-scale samples of decalcified coral tissues. Complimentary FM and TPLSM reveal subtle submillimeter&#160;scale changes in cellular distribution and density in nondecalcified coral tissue samples. The TPLSM technique affords: (1) minimally invasive sample preparation,&#160;(2) superior optical sectioning ability,&#160;and (3) minimal light absorption and scattering, while still permitting deep tissue imaging.

An integrated suite of imaging techniques has been applied to determine polyp morphology and tissue structure in the Caribbean corals Montastraeaannularis and M. faveolata. Fluorescence, serial block face,&#160;and two-photon confocal laser scanning microscopy have identified lobate structure, polyp walls, and estimated chromatophore and zooxanthellae densities and distributions.

Multimodali ottici Metodi Microscopia Reveal Polyp Tissue morfologia e struttura in Caribbean Reef costruzione Coralli

An integrated suite of imaging techniques has been applied to determine the three-dimensional (3D) morphology and cellular structure of polyp tissues ...

Ricerca

Ambiente

A Fish-feeding Laboratory Bioassay to Assess the Antipredatory Activity of Secondary Metabolites from the Tissues of Marine Organisms

Marine chemical ecology is a young discipline, having emerged from the collaboration of natural products chemists and marine ecologists in the 1980s with the goal of examining the ecological functions of secondary metabolites from the tissues of marine organisms. The result has been a progression of protocols that have increasingly refined the ecological relevance of the experimental approach. Here we present the most up-to-date version of a fish-feeding laboratory bioassay that enables investigators to assess the antipredatory activity of secondary metabolites from the tissues of marine organisms. Organic metabolites of all polarities are exhaustively extracted from the tissue of the target organism and reconstituted at natural concentrations in a nutritionally appropriate food matrix. Experimental food pellets are presented to a generalist predator in laboratory feeding assays to assess the antipredatory activity of the extract. The procedure described herein uses the bluehead, Thalassoma bifasciatum, to test the palatability of Caribbean marine invertebrates; however, the design may be readily adapted to other systems. Results obtained using this laboratory assay are an important prelude to field experiments that rely on the feeding responses of a full complement of potential predators. Additionally, this bioassay can be used to direct the isolation of feeding-deterrent metabolites through bioassay-guided fractionation. This feeding bioassay has advanced our understanding of the factors that control the distribution and abundance of marine invertebrates on Caribbean coral reefs and may inform investigations in diverse fields of inquiry, including pharmacology, biotechnology, and evolutionary ecology.

This bioassay employs a model predatory fish to assess the presence of feeding-deterrent metabolites from organic extracts of the tissues of marine organisms at natural concentrations using a nutritionally comparable food matrix.

Un pesce-alimentare Laboratorio Bioassay per valutare l&#39;attività di antipredatory Secondary metaboliti dai tessuti di organismi marini

Marine chemical ecology is a young discipline, having emerged from the collaboration of natural products chemists and marine ecologists in the 1980s with ...

Individual Culturing of Tigriopus Copepods and Quantitative Analysis of Their Mate-guarding Behavior

Copepods of the genus Tigriopus, which are common zooplankton in rocky tide pools, show precopulatory mate-guarding behavior where a male clasps a potential mate to form a pair. While this phenomenon has attracted interest of researchers, methods for its analysis have not been well described. Here we describe procedures for: 1) individual culturing and staging of Tigriopus juveniles and adults, and 2) video-based analysis of their mate-guarding behavior. The culturing method enables experimental control of paring experience of animals as well as the ability to track their development before behavioral tests. The analysis method allows quantitative evaluation of several aspects of the mate-guarding behavior, including capturing attempts by males and swimming trajectory of mate-guarding pairs. Although these methods were originally established for ethological studies on Tigriopus, with proper modifications they can also be applied to studies of other zooplankton in different research fields, such as physiology, toxicology, and ecological genetics.

Individual Culturing of Tigriopus Copepods and Quantitative Analysis of Their Mate-guarding Behavior

Mate-guarding behavior plays an important role in reproduction of intertidal copepods of the genus Tigriopus. However, methods for studying this behavior have not been well described. Here we describe methods for: 1) individual culture of virgin Tigriopus animals, and 2) quantitative analysis of their mate-guarding behavior.

Coltura individuali di copepodi Tigriopus e analisi quantitativa del loro comportamento Mate-guardia

Copepods of the genus Tigriopus, which are common zooplankton in rocky tide pools, show precopulatory mate-guarding behavior where a male clasps a ...

Probing the Limits of Egg Recognition Using Egg Rejection Experiments Along Phenotypic Gradients

Brood parasites lay their eggs in other females' nests, leaving the host parents to hatch and rear their young. Studying how brood parasites manipulate hosts into raising their young and how hosts detect parasitism provide important insights in the field of coevolutionary biology. Brood parasites, such as cuckoos and cowbirds, gain an evolutionary advantage because they do not have to pay the costs of rearing their own young. However, these costs select for host defenses against all developmental stages of parasites, including eggs, their young, and adults. Egg rejection experiments are the most common method used to study host defenses. During these experiments, a researcher places an experimental egg in a host nest and monitors how hosts respond. Color is often manipulated, and the expectation is that the likelihood of egg discrimination and the degree of dissimilarity between the host and experimental egg are positively related. This paper serves as a guide for conducting egg rejection experiments from describing methods for creating consistent egg colors to analyzing the findings of such experiments. Special attention is given to a new method involving uniquely colored eggs along color gradients that has the potential to explore color biases in host recognition. Without standardization, it is not possible to compare findings between studies in a meaningful way; a standard protocol within this field will allow for increasingly accurate and comparable results for further experiments.

This protocol provides guidelines for running egg rejection experiments: outlining techniques for painting experimental egg models to emulate the colors of natural bird eggs, conducting fieldwork, and analyzing the collected data. This protocol provides a uniform method for conducting comparable egg rejection experiments.

Sondare i limiti del riconoscimento uovo utilizzando esperimenti di rifiuto uovo lungo gradienti fenotipiche

Brood parasites lay their eggs in other females' nests, leaving the host parents to hatch and rear their young. Studying how brood parasites manipulate ...

Comportamento

Using the FishSim Animation Toolchain to Investigate Fish Behavior: A Case Study on Mate-Choice Copying In Sailfin Mollies

Over the last decade, employing computer animations for animal behavior research has increased due to its ability to non-invasively manipulate the appearance and behavior of visual stimuli, compared to manipulating live animals. Here, we present the FishSim Animation Toolchain, a software framework developed to provide researchers with an easy-to-use method for implementing 3D computer animations in behavioral experiments with fish. The toolchain offers templates to create virtual 3D stimuli of five different fish species. Stimuli are customizable in both appearance and size, based on photographs taken of live fish. Multiple stimuli can be animated by recording swimming paths in a virtual environment using a video game controller. To increase standardization of the simulated behavior, the prerecorded swimming path may be replayed with different stimuli. Multiple animations can later be organized into playlists and presented on monitors during experiments with live fish.
In a case study with sailfin mollies (Poecilia latipinna), we provide a protocol on how to conduct a mate-choice copying experiment with FishSim. We utilized this method to create and animate virtual males and virtual model females, and then presented these to live focal females in a binary choice experiment. Our results demonstrate that computer animation may be used to simulate virtual fish in a mate-choice copying experiment to investigate the role of female gravid spots as an indication of quality for a model female in mate-choice copying.
Applying this method is not limited to mate-choice copying experiments but can be used in various experimental designs. Still, its usability depends on the visual capabilities of the study species and first needs validation. Overall, computer animations offer a high degree of control and standardization in experiments and bear the potential to &#39;reduce&#39; and &#39;replace&#39; live stimulus animals as well as to &#39;refine&#39; experimental procedures.

Using the FishSim Animation Toolchain to Investigate Fish Behavior: A Case Study on Mate-Choice Copying In Sailfin Mollies

Using the novel FishSim Animation Toolchain, we present a protocol for non-invasive visual manipulation of public information in the context of mate-choice copying in sailfin mollies. FishSim Animation Toolchain provides an easy-to-use framework for the design, animation and presentation of computer-animated fish stimuli for behavioral experiments with live test fish.

Utilizzando la Toolchain di animazione FishSim per indagare il comportamento dei pesci: un caso di studio su Mate-scelta copia In Mollies Sailfin

Over the last decade, employing computer animations for animal behavior research has increased due to its ability to non-invasively manipulate the ...

Behavioral Tracking and Neuromast Imaging of Mexican Cavefish

Cave-dwelling animals have evolved a series of morphological and behavioral traits to adapt to their perpetually dark and food-sparse environments. Among these traits, foraging behavior is one of the useful windows into functional advantages of behavioral trait evolution. Presented herein are updated methods for analyzing vibration attraction behavior (VAB: an adaptive foraging behavior) and imaging of associated mechanosensors of cave-adapted tetra, Astyanax mexicanus. In addition, methods are presented for high-throughput tracking of a series of additional cavefish behaviors including hyperactivity and sleep-loss. Cavefish also show asociality, repetitive behavior and higher anxiety. Therefore, cavefish serve as an animal model for evolved behaviors. These methods use free-software and custom-made scripts that can be applied to other types of behavior. These methods provide practical and cost-effective alternatives to commercially available tracking software.

Here, we present methods for high-throughput study of a series of the Mexican cavefish behaviors and vital staining of a mechanosensory system. These methods use free-software and custom-made scripts, providing a practical and cost-effective method for the studies of behaviors.

Rilevamento comportamentale e formazione immagine di Neuromast della caverna messicano

Cave-dwelling animals have evolved a series of morphological and behavioral traits to adapt to their perpetually dark and food-sparse environments. Among ...

A Simple Behavioral Assay for Testing Visual Function in Xenopus laevis

Measurement of the visual function in the tadpoles of the frog, Xenopus laevis, allows screening for blindness in live animals. The optokinetic response is a vision-based, reflexive behavior that has been observed in all vertebrates tested. Tadpole eyes are small so the tail flip response was used as alternative measure, which requires a trained technician to record the subtle response. We developed an alternative behavior assay based on the fact that tadpoles prefer to swim on the white side of a tank when placed in a tank with both black and white sides. The assay presented here is an inexpensive, simple alternative that creates a response that is easily measured. The setup consists of a tripod, webcam and nested testing tanks, readily available in most Xenopus laboratories. This article includes a movie showing the behavior of tadpoles, before and after severing the optic nerve. In order to test the function of one eye, we also include representative results of a tadpole in which each eye underwent retinal axotomy on consecutive days. Future studies could develop an automated version of this assay for testing the vision of many tadpoles at once.

A Simple Behavioral Assay for Testing Visual Function in Xenopus laevis

Xenopus laevis tadpoles prefer swimming on the white side of a black/white tank. This behavior is guided by their vision. Based on this behavior, we present a simple assay to test the visual function of tadpoles.

Un test comportamentale semplice for Testing Funzione visiva in<em&gt; Xenopus laevis</em

Measurement of the visual function in the tadpoles of the frog, Xenopus laevis, allows screening for blindness in live animals. The optokinetic response ...

Neuroscienze

Swimming Performance Assessment in Fishes

Swimming performance tests of fish have been integral to studies of muscle energetics, swimming mechanics, gas exchange, cardiac physiology, disease, pollution, hypoxia and temperature. This paper describes a flexible protocol to assess fish swimming performance using equipment in which water velocity can be controlled. The protocol involves one to several stepped increases in flow speed that are intended to cause fish to fatigue. Step speeds and their duration can be set to capture swimming abilities of different physiological and ecological relevance. Most frequently step size is set to determine critical swimming velocity (Ucrit), which is intended to capture maximum sustained swimming ability. Traditionally this test has consisted of approximately ten steps each of 20 min duration. However, steps of shorter duration (e.g. 1 min) are increasingly being utilized to capture acceleration ability or burst swimming performance. Regardless of step size, swimming tests can be repeated over time to gauge individual variation and recovery ability. Endpoints related to swimming such as measures of metabolic rate, fin use, ventilation rate, and of behavior, such as the distance between schooling fish, are often included before, during and after swimming tests. Given the diversity of fish species, the number of unexplored research questions, and the importance of many species to global ecology and economic health, studies of fish swimming performance will remain popular and invaluable for the foreseeable future.

The lives of the majority of fish are predicated on swimming. This protocol describes techniques for capturing a range of swimming modes available to individual and schooling fish, and includes metrics associated with swimming physiology and behaviour.

La valutazione delle prestazioni nuoto in Pesci

Swimming performance tests of fish have been integral to studies of muscle energetics, swimming mechanics, gas exchange, cardiac physiology, disease, ...

Biologia

Visualization of Caenorhabditis elegans Cuticular Structures Using the Lipophilic Vital Dye DiI

The cuticle of C. elegans is a highly resistant structure that surrounds the exterior of the animal1-4. The cuticle not only protects the animal from the environment, but also determines body shape and plays a role in motility4-6. Several layers secreted by epidermal cells comprise the cuticle, including an outermost lipid layer7.
Circumferential ridges in the cuticle called annuli pattern the length of the animal and are present during all stages of development8. Alae are longitudinal ridges that are present during specific stages of development, including L1, dauer, and adult stages2,9. Mutations in genes that affect cuticular collagen organization can alter cuticular structure and animal body morphology5,6,10,11. While cuticular imaging using compound microscopy with DIC optics is possible, current methods that highlight cuticular structures include fluorescent transgene expression12, antibody staining13, and electron microscopy1. Labeled wheat germ agglutinin (WGA) has also been used to visualize cuticular glycoproteins, but is limited in resolving finer cuticular structures14. Staining of cuticular surface using fluorescent dye has been observed, but never characterized in detail15. We present a method to visualize cuticle in live C. elegans using the red fluorescent lipophilic dye DiI (1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate), which is commonly used in C. elegans to visualize environmentally exposed neurons. This optimized protocol for DiI staining is a simple, robust method for high resolution fluorescent visualization of annuli, alae, vulva, male tail, and hermaphrodite tail spike in C. elegans.

Visualization of Caenorhabditis elegans Cuticular Structures Using the Lipophilic Vital Dye DiI

We present a method to visualize cuticle in live C. elegans using the red fluorescent lipophilic dye DiI (1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate), which is commonly used in C. elegans to visualize environmentally exposed neurons. With this optimized protocol, alae and annular cuticular structures are stained by DiI and observed using compound microscopy.

Visualizzazione di Caenorhabditis Elegans Strutture cuticola Utilizzando il Dye lipofili Vital, Dii

The cuticle of C. elegans is a highly resistant structure that surrounds the exterior of the animal1-4. The cuticle not only protects the animal from the ...

Analysis of Skeletal Muscle Defects in Larval Zebrafish by Birefringence and Touch-evoke Escape Response Assays

Zebrafish (Danio rerio) have become a particularly effective tool for modeling human diseases affecting skeletal muscle, including muscular dystrophies1-3, congenital myopathies4,5, and disruptions in sarcomeric assembly6,7, due to high genomic and structural conservation with mammals8. Muscular disorganization and locomotive impairment can be quickly assessed in the zebrafish over the first few days post-fertilization. Two assays to help characterize skeletal muscle defects in zebrafish are birefringence (structural) and touch-evoked escape response (behavioral).
Birefringence is a physical property in which light is rotated as it passes through ordered matter, such as the pseudo-crystalline array of muscle sarcomeres9. It is a simple, noninvasive approach to assess muscle integrity in translucent zebrafish larvae early in development. Wild-type zebrafish with highly organized skeletal muscle appear very bright amidst a dark background when visualized between two polarized light filters, whereas muscle mutants have birefringence patterns specific to the primary muscular disorder they model. Zebrafish modeling muscular dystrophies, diseases characterized by myofiber degeneration followed by repeated rounds of regeneration, exhibit degenerative dark patches in skeletal muscle under polarized light. Nondystrophic myopathies are not associated with necrosis or regenerative changes, but result in disorganized myofibers and skeletal muscle weakness. Myopathic zebrafish typically show an overall reduction in birefringence, reflecting the disorganization of sarcomeres.
The touch-evoked escape assay involves observing an embryo&#39;s swimming behavior in response to tactile stimulation10-12. In comparison to wild-type larvae, mutant larvae frequently display a weak escape contraction, followed by slow swimming or other type of impaired motion that fails to propel the larvae more than a short distance12. The advantage of these assays is that disease progression in the same fish type can be monitored in vivo for several days, and that large numbers of fish can be analyzed in a short time relative to higher vertebrates.

The zebrafish is now an established and powerful tool for modeling muscular dystrophies, congenital myopathies, and related neuromuscular diseases. Birefringence and touch-evoked escape behavior are two common noninvasive assays used to determine the degree of muscular disorganization and locomotive impairment of zebrafish embryos during early development.

Metodi per la determinazione strutture UV-riflettenti e determinare la loro importanza nella Mate Preferenze utilizzando il Sailfin Molly

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