Estimating density of calling male Eleutherodactylus coqui in Hawaii from audio recordings of the nighttime frog chorus

Analysis of sound pressure data obtained from audio recordings of night-time Eleutherodactylus coqui chorusing was used to estimate population density of calling male coqui frogs in the major Hawaiian island populations. Study plots 400 m² in size (20 by 20 m; n = 32) were established on the islands of Hawaii and Maui to provide venues for field counting calling male frogs to investigate the ground truth relationship with the recorded sound pressure data. Male coqui frogs emit a two-note ‘co-qui’ advertisement call at regular intervals. Exploiting the temperature-related and relatively consistent male coqui call repetition rate and the average emitted ‘co’ call sound pressure level (70.4 dB at 1 m), cue counts obtained from a correlation process available in the program Raven were also used to estimate the density of calling male coqui. The study methodology included comparing linear regression results between the field-counted frogs and the number of frogs obtained from the cue count method. Both methods revealed a linear relationship between calling male population density and associated average sound pressure of the resulting chorus.     

ADAPTATION TO SEVERE CONDITIONS OF PROPAGATION: LONG-DISTANCE DISTRESS CALLS; AND COURTSHIP CALLS OF A COLONIAL SEABIRD

ABSTRACT

The boatwhistle sound produced by male toadfish during the reproductive season attracts females to the nest site. Boatwhistles consist of a series of rapidly produced, amplitude modulated ‘pulses’ of sound that are generated by specific muscles of the gas bladder. Previous studies have shown that boatwhistle characteristics vary with temperature and geographic location. This study investigated the normal range of variation in boatwhistles produced by males in a northern population of O. tau. Multiple boatwhistles were recorded from individuals at different sites around Waquoit Bay, Massachusetts. Multiple boatwhistles were recorded at different sites to assess the range of variation in boatwhistle production from this population. Pulse repetition rates varied from a low of 125 pulses/sec at 16 °C to a high of 219 pulses/sec at 21 °C. Careful examination of recordings from different sites indicated that individuals vary in the durations of their boatwhistles as well as in the pulse repetition rates during the very consistently produced second segment of the call. In particular, pulse repetition rate (PRR) varied significantly (p < 0.001) among most individuals recorded at the same temperatures. Psychophysical testing as well as behavioral choice experiments are needed to assess the relative importance of PRR and spectral cues in species recognition and/or mate choice.     

Changes of movement behavior and HSP70 gene expression in the hemocytes of the mud crab (Scylla paramamosain) in response to acoustic stimulation

Offshore anthropogenic activities often produce high levels of noise below 1000 Hz, which can be serious threats to aquatic crustaceans based on the knowledge about their acoustic sensitive bandwidth. This study simulated noise with main frequency band similar to common underwater engineering noises, and examined its effects on movement behavior and physiological response, indicated by heat shock protein 70 (HSP70) gene expression, of the mud crab Scylla paramamosain. Experiments were conducted in the tanks, equipped with hydrophone, transducer, and video recording system, using juvenile S. paramamosain. Three acoustic stimulations with ascending levels were separately imposed on the animals. Results showed that the linear sweep with the sound power spectral density greater than 110 dB re 1μPa²/Hz in total bandwidth (100–1000 Hz), and 155 dB re 1μPa²/Hz within 600–800 Hz, could increase locomotor activities and HSP70 gene expression significantly.     

Singing of <Emphasis Type="Italic">Neoconocephalus robustus</Emphasis> as an example of deterministic chaos in insects

We use nonlinear time series analysis methods to analyse the dynamics of the sound-producing apparatus of the katydid Neoconocephalus robustus. We capture the dynamics by analysing a recording of the singing activity. First, we reconstruct the phase space from the sound recording and test it against determinism and stationarity. After confirming determinism and stationarity, we show that the maximal Lyapunov exponent of the series is positive, which is a strong indicator for the chaotic behaviour of the system. We discuss that methods of nonlinear time series analysis can yield instructive insights and foster the understanding of acoustic communication among insects.     

Effects of ionizing radiation and preservation on biomechanical properties of human costal cartilage

Tissue banks around the world store human cartilage obtained from cadaveric donors for use in diverse reconstructive surgical procedures. To ensure this tissue is sterile at the time of distribution, tissues may be sterilized by ionizing radiation. In this work, we evaluate the physical changes in deep frozen costal cartilage (?70 °C) or costal cartilage preserved in high concentrations of glycerol (>98 %) followed by a terminal sterilization process using ionizing radiation, at 3 different doses (15, 25 and 50 kGy). Tension and compression tests were carried out to determine the mechanical changes related both to the different preservation methods and irradiation doses. For both methods of preservation, tension strength was increased by about 24 %, when cartilage tissue was irradiated with 15 kGy. Deep frozen samples, when irradiated with 25 or 50 kGy, had a decrease in their mechanical performance, albeit to a lesser extent than when tissues were preserved in high concentration of glycerol and equally irradiated. In conclusion, processing in high concentration of glycerol did not increase tissue protection against radiation damage; while cartilage preserved in high concentrations of glycerol withstands radiation up to 25 kGy, deep frozen human costal cartilage may be sterilized with a doses up to 50 kGy without significant mechanical impact.     

Florivory by a floriphilic katydid,Phaneroptera brevis,induces changes in a leaf trait in Lantana camara

1. Plants can induce a response when they are attacked by herbivores. Although the induction of responses by herbivory in both flowers and leaves is relatively well studied, whether florivory (feeding of flowers) can also induce responses in flowers and leaves is less well explored and there are still unanswered questions. These include whether plants exhibit different levels of induced responses depending on the length of exposure to the florivores. 2. To address this knowledge gap, this study used a tropical floriphilic katydid, Phaneroptera brevis, and its non‐native food plant, Lantana camara. Nursery experiments were performed in which dry matter content and anthocyanin concentration of the flower (corolla and stamens), leaf dry matter content, and leaf blade punch resistance were measured at three time phases before and after exposing the plant to the katydid individuals for 0–7 days. 3. It was demonstrated that increasing the length (days) of exposure to the katydid individual leads to higher levels of induced plant response (leaf blade punch resistance), but only in the leaves. It was also shown that higher levels of induced plant response owing to the increase in the length of exposure to the katydid individual was not observed beyond the first set of leaves developed after the exposure. 4. These results address the knowledge gap and show that plants can exhibit different levels of induced responses depending on the length of exposure to florivores. This study thus highlights the far‐reaching importance of florivory on plants.     

Host range of Lepidelphax pistiae (Hemiptera: Delphacidae) and its potential impact on Pistia stratiotes L. (Araceae)

Pistia stratiotes L. (Araceae) is a floating aquatic plant that has become invasive in Florida. It is primarily controlled with herbicides, but two biocontrol agents have previously been released to assist in management of this species. A new potential biocontrol agent from Argentina, Lepidelphax pistiae Remes Lenicov (Hemiptera: Delphacidae), has been evaluated comprehensively for specificity after initial host range studies done in its native range indicated that it is likely specific to P. stratiotes. Host range studies indicated that this insect is specific to P. stratiotes, with no survival or reproduction occurring on any of the 42 other plant species tested. Impact studies indicated that this insect can significantly damage P. stratiotes at medium and high population densities, which were comparable to those seen in its native range. Lepidelphax pistiae is sufficiently specific enough to warrant release and has a high probability of aiding management of P. stratiotes populations in Florida.     

Diagnostics of nocturnal calls of Sciaena umbra (L., fam. Sciaenidae) in a nearshore Mediterranean marine reserve

Calls emitted by the brown meagre Sciaena umbra (L., fam. Sciaenidae) were recorded at the Natural Marine Reserve of Miramare (Trieste, Italy) in seven nocturnal surveys (12-h continuous sampling) during the summer of 2009. Calls consist of pulses, with the main energy content below 2 kHz and mean peak frequency of c. 270 Hz. Pulses were short, with an average duration of 20 ms and a pulse period of 100 ms. Sounds lasted approximately 500 ms. Three types of sound patterns were recognized: irregular (I), regular (R) and the chorus (C). Their acoustic parameters are described showing that I, R and C differ in pulse duration, pulse peak frequency and pulse period. Occurrence of the three call types changes throughout the night: the R pattern occurred mainly at dawn and dusk, C predominated after nightfall, while I calls were produced sporadically during the whole nocturnal period. Our results indicate that S. umbra has a pronounced nocturnal rhythm in vocalizing behaviour and highlight how the diagnostic time–frequency pattern of S. umbra calls can be used to identify the species in the field. Considering that the abundance of S. umbra is currently declining, the information presented here will be relevant in developing non-invasive and low-cost monitoring acoustic systems for managing S. umbra conservation and fishery along the Mediterranean Sea.     

Long‐term monitoring of captive red drum Sciaenops ocellatus reveals that calling incidence and structure correlate with egg deposition

In the present study, quantitative data were collected to clarify the relationship between calling, call structure and eggs produced in a captive population of red drum Sciaenops ocellatus. Sciaenops ocellatus were held in four tanks equipped with long‐term acoustic loggers to record underwater sound throughout a simulated reproductive season. Maximal sound production of captive S. ocellatus occurred when the photoperiod shifted from 13·0 to 12·5 h of light, and the water temperature decreased to c. 25° C. These captive settings are similar to the amount of daylight and water temperatures observed during the autumn, which is the primary spawning period for S. ocellatus. Sciaenops ocellatus exhibited daily patterns of calling with peak sound production occurring in the evenings between 0·50 h before dark and 1·08 h after dark. Spawning occurred only on evenings in which S. ocellatus were calling, and spawning was more productive when S. ocellatus produced more calls with longer durations and more pulses. This study provides ample evidence that sound production equates to spawning in captive S. ocellatus when calls are longer than 0·8 s and contain more than seven pulses. The fact that more calling, longer calls and higher sound pressure levels are associated with spawns that are more productive indicates that acoustic metrics can provide quantitative information on spawning in the wild.     

Production and Purification of Recombinant Enteropeptidase Expressed in an Insect–Baculovirus Cell System

Enteropeptidase (EC 3.4.21.9) is the glycoprotein enzyme in the small intestine that triggers the activation of the zymogens in pancreatic juice by converting trypsinogen into trypsin. Because of its physiological significance, there have been many studies on the expression, purification, and characterization of enteropeptidase from different species. The baculovirus expression system has been commonly used in research communities and scientific industries for the production of high levels of recombinant proteins, which require posttranslational modifications for functional activity. In the present study, we isolated bovine enteropeptidase catalytic subunit gene from Bos taurus indicus (GenBank accession no. KC756844), and cloned it in pFast Bac HT “A” baculovirus expression donor vector, under the polyhedrin promoter. Recombinant bovine enteropeptidase was expressed in SF-9 insect cells with high expression levels. Recombinant enteropeptidase was purified using Ni-NTA affinity chromatography. A 6-mg quantity of pure active protein was obtained from 100 mL culture using this approach. Its activity and kinetic parameters were determined by cleavage of its fluorogenic substrate Gly-(Asp) 4-Lys-β-naphthylamide. The recombinant bovine enteropeptidase showed a K _m value of 0.75 ± 0.02 mM with K _cat 25 ± 1 s.     

Effects of Metarhizium anisopliae on Meccus pallidipennis (Hemiptera: Reduviidae) over different types of wall surfaces

The entomopathogenic fungus Metarhizium anisopliae is virulent for the insect triatomine Meccus pallidipennis. To evaluate the functionality of a fungal formulation (vegetable oil and emulsifiers) of this fungus, virulence was assayed by insect mortality on the pronotum of third instar nymphs (N3) M. pallidipennis in laboratory conditions and ST₅₀ was calculated. Mortality was evaluated directly: 100%, 97.33% and 98.66% mortalities were caused by formulation, emulsified formulation and fungal conidia, respectively, at day 8 of insect infection. Another bioassay was carried out in simulated external conditions (peridomicility) using red and gray brick walls, a stone fence and mountain soil (experimental units). These simulated conditions were infected with 10?ml of a 1?×?10⁹?conidia/ml emulsified formulation by means of a manual sprinkler prior to the placement of the nymphs. Ten N3 M. pallidipennis were deposited in each experimental unit and insect mortality was monitored every 12?h for 22 days. Each treatment was replicated four times. With the red brick wall, a mortality of 90% at day 22 and a ST₅₀ of 15 days were obtained on N3 M. pallidipennis; with the gray brick wall, 100% mortality and a ST₅₀ of 13 days; and with the stone fence, 88.33% mortality and a ST₅₀ of 21 days. The results obtained in this research work indicate that the formulation with conidia of the M. anisopliae strain EH-473/4 may be auxiliary in the development of strategies for the control of Chagas disease insect transmitters such as M. pallidipennis.     

Acoustic signalling in female fish: factors influencing sound characteristics in croaking gouramis

The characteristics of sounds produced by fishes are influenced by several factors such as size. The current study analyses factors affecting structural properties of acoustic signals produced by female croaking gouramis Trichopsis vittata during agonistic interactions. Female sounds (although seldom analysed separately from male sounds) can equally be used to investigate factors affecting the sound characteristics in fish. Sound structure, dominant frequency and sound pressure levels (SPL) were determined and correlated to body size and the order in which sounds were emitted. Croaking sounds consisted of series of single-pulsed or double-pulsed bursts, each burst produced by one pectoral fin. Main energies were concentrated between 1.3 and 1.5 kHz. The dominant frequency decreased with size, as did the percentage of single-pulsed bursts within croaking sounds. The SPL and the number of bursts within a sound were independent of size but decreased significantly with the order of their production. Thus, acoustic signals produced at the beginning of agonistic interactions were louder and consisted of more bursts than subsequent ones. Our data indicate that body size affects the dominant frequency and structure of sounds. The increase in the percentage of double-pulsed bursts with size may be due to stronger pectoral muscles in larger fish. In contrast, ongoing fights apparently result in muscle fatigue and subsequently in a decline in the number of bursts and SPL. The factor ‘order of sound production’ points to an intra-individual variability of sounds and should be considered in future studies.     

Localization of virtual stimuli moving in the median plane by listeners of different age

Capability for identification of direction of movement of sound images (“upward” or “downward”) was studied in listeners of two age groups: 19–27 year old (11 subjects) and 55–73 year old (9 subjects). Various sound models of movement in the median plane were used as stimuli. Initially, a model of movement was developed based on filtration of broadband noise pulses by sets of “non-individualized,” i. e., measured in other listeners, head-related transfer functions. These functions corresponded to consecutive positions of the sound sources with the 5.6° step between the space points with coordinates of elevation from ?45° to 45°. The signals generated on the basis of transfer function sets of 23 subjects were distinguished by regularity and the value of the spectral minimum shift as well as by dynamic changes of the spectral maximum. These dynamic changes were taken into account at creation of “synthetic” signals. In these signals, the vertical movement of sound images in the median plane was simulated either by a consecutive shift of the spectral minimum in broadband noise pulses or by a combination of the spectral minimum shift with a simultaneous change of the spectral maximum width and power. The obtained data have shown that young listeners with the high capability for vertical localization could identify direction of the sound image movement based on displacement of the spectral minimum in the broadband noise. For identification of direction of the sound image movement, the younger listeners with poor capabilities for vertical localization and the older listeners used dynamic changes of the signal power, which were connected mainly with the spectral maximum range.     

Effects of Carbofuran on Lantana camara and its biocontrol agent,Teleonemia scrupulosa

This laboratory-based study sought to determine the efficacy of using carbofuran in an exclusion experiment aimed at assessing the impacts of biocontrol agents on Lantana camara L. (sensu lato) (Verbenaceae). Two separate experiments were conducted, the first one on insect-free plants, to determine the effects of carbofuran solely on plant growth; and the second one, on Teleonemia scrupulosa Stål (Hemiptera: Tingidae) infested plants, with the objective of determining the impact of carbofuran on this biocontrol agent, as well as its impacts on plant growth. Carbofuran granules (10% a.i.) were applied at 7?g/m² a.i. to the potting medium. It was found that carbofuran did not have a significant effect on plant growth. Total removal of T. scrupulosa from exclusion plants (carbofuran-treated plants) was not achieved; however the low level of leaf feeding lesions on those plants indicated that carbofuran had considerably reduced the insect’s population density. Results from a bioassay showed 100% and 40% T. scrupulosa mortality on leaves collected from carbofuran-treated and control plants, respectively, within three weeks of exposure. Analysis of chemical residue levels in the leaf material revealed that carbofuran potency only persisted for about three weeks, and was detectable at trace levels (<0.1?mg/kg). It was therefore concluded that carbofuran was effective at reducing the population of T. scrupulosa on its host plant, but only briefly. Carbofuran should be applied at least once every three weeks or at a higher dosage in order to maintain a low insect population for the duration of an experiment or to achieve total exclusion. For better insect removal, one should consider combining carbofuran and foliar insecticides.     

Context Specific Signaling with Different Frequencies - Directed to Different Receivers? A Case Study in <Emphasis Type="Italic">Gonatoxia</Emphasis> Katydids (Orthoptera<Emphasis Type="Italic">,</Emphasis> Phaneropteridae)

In katydids (Orthoptera: Tettigonioidea) of the subfamily Phaneropterinae females ready to mate initiate a duet, announcing her position to the male singer, but also potentially to eavesdropping rivals. In many species the male seems to defend the communication by adding self-produced imitations of a female response. If these signals occur within the male sensory time-window after the female song, they can disturb the orientation of rivals. In two species of the genus Gonatoxia, males and females use short, relatively narrow-banded sounds (width 2–7 kHz 10 dB below peak). Male song and female response, however, differ considerably in peak frequency. In G. maculata, the peak frequency of the last part of the male song (13 kHz) is between that of the first part (15 kHz) and the female response (9 kHz), in G. helleri the last part (9 kHz; assumed imitation) and the female song are identical in peak frequency and by a factor two lower than the first part (19 kHz). The male stridulatory file of this species is correspondingly modified and differs from all other members of the genus. The imitation of spectral properties of the female response is not known from any other katydid.     

Intra-population variation and geographic correlation in Canthon humectus hidalgoensis using FTIR-ATR spectroscopy

In some cases external morphology is not sufficient to discern between populations of a species, as occurs in the dung beetle Canthon humectus hidalgoensis Bates; and much less to determine phenotypic distances between them. FTIR-ATR spectroscopy show several advantages over other identification techniques (e.g. morphological, genetic, and cuticular hydrocarbons analysis) due to the non-invasive manner of the sample preparation, the relative speed of sample analysis and the low-cost of this technology. The infrared spectrum obtained is recognized to give a unique ‘fingerprint’ because vibrational spectra are specific and unique to the molecular nature of the sample. In our study, results showed that proteins, amino acids and aromatic ethers of insect exocuticle have promising discriminative power to discern between different populations of C. h. hidalgoensis. Furthermore, the correlation between geographic distances between populations and the chemical distances obtained by proteins + amino acids + aromatic ethers was statistically significant, showing that the spectral and spatial information available of the taxa together with appropriated chemometric methods may help to a better understanding of the identity, structure, dynamics and diversity of insect populations.     

Otoacoustic emissions from insect ears having just one auditory neuron

Sensitive hearing organs often employ nonlinear mechanical sound processing which produces distortion-product otoacoustic emissions. Such emissions are also recorded from insect tympanal organs. Here we report high frequency distortion-product emissions, evoked by stimulus frequencies up to 95 kHz, from the tympanal organ of a notodontid moth, Ptilodon cucullina, which contains only a single auditory receptor neuron. The 2f1–f2 distortion-product emission reaches sound levels above 40 dB SPL. Most emission growth functions show a prominent notch of 20 dB depth (n = 20 trials), accompanied by an average phase shift of 119°, at stimulus levels between 60 and 70 dB SPL, which separates a low- and a high-level component. The emissions are vulnerable to topical application of ethyl ether which shifts growth functions by about 20 dB towards higher stimulus levels. For the mammalian cochlea, Lukashkin and colleagues have proposed that distinct level-dependent components of nonlinear amplification do not necessarily require interaction of several cellular sources but could be due to a single nonlinear source. In notodontids, such a physiologically vulnerable source could be the single receptor cell. Potential contributions from accessory cells to the nonlinear properties of the scolopidial hearing organ are still unclear.     

Cryoprotection of lipid membranes for high-resolution solid-state NMR studies of membrane peptides and proteins at low temperature

Solid-state NMR spectra of membrane proteins often show significant line broadening at cryogenic temperatures. Here we investigate the effects of several cryoprotectants to preserve the spectral resolution of lipid membranes and membrane peptides at temperatures down to ~200 K. Trehalose, glycerol, dimethylsulfoxide (DMSO), dimethylformamide (DMF), and polyethylene glycol (PEG), were chosen. These compounds are commonly used in protein crystallography and cryobiology. ¹³C and ¹H magic-angle-spinning spectra of several types of lipid membranes show that DMSO provides the best resolution enhancement over unprotected membranes and also best retards ice formation at low temperature. DMF and PEG-400 show slightly weaker cryoprotection, while glycerol and trehalose neither prevent membrane line broadening nor prevent ice formation under the conditions of our study. Neutral saturated-chain phospholipids are the most amenable to cryoprotection, whereas negatively charged and unsaturated lipids attenuate cryoprotection. ¹³C–¹H dipolar couplings and ³¹P chemical shift anisotropies indicate that high spectral resolution at low temperature is correlated with stronger immobilization of the lipids at high temperature, indicating that line narrowing results from reduction of the conformational space sampled by the lipid molecules at high temperature. DMSO selectively narrowed the linewidths of the most disordered residues in the influenza M2 transmembrane peptide, while residues that exhibit narrow linewidths in the unprotected membrane are less impacted. A relatively rigid β-hairpin antimicrobial peptide, PG-1, showed a linewidth increase of ~0.5 ppm over a ~70 K temperature drop both with and without cryoprotection. Finally, a short-chain saturated lipid, DLPE, exhibits excellent linewidths, suggesting that it may be a good medium for membrane protein structure determination. The three best cryoprotectants found in this work—DMSO, PEG, and DMF—should be useful for low-temperature membrane-protein structural studies by SSNMR without compromising spectral resolution.