Development and assays estradiol equivalent concentration from prawn (p-EEQ) in river prawn, Macrobrachium nipponense, in Taiwan.

The present study established a fast and convenient bioassay method for aqueous ecosystems using the prawn estradiol equivalent concentration (p-EEQ) of male Macrobrachium nipponense, which produce vitellogenin (VTG) after exposure to xeno-estrogens. This method was then used to determine the concentrations of xeno-estrogen pollutants in the rivers of Taiwan. To establish the calibration curve for the concentrations based on the p-EEQ, the induced VTG content was determined using the alkali-labile phosphate method after male M. nipponense were exposed to 0, 10, 100, 1,000 and 10,000ng/L of 17ß-estradiol for 1, 3, 5, 7, 10 and 14 days, respectively. The results of the experiments showed that the induced VTG content in all of the experimental groups stabilized after 10 days, except for the 10,000ng/L experimental group, in which the induced VTG content decreased after 10 days. A 17ß-estradiol-VTG10day response curve was then established based on the induced VTG content in the 0, 10, 100 and 1000ng/L experimental groups at day 10. After establishing the curve, male M. nipponense were captured from the upper, middle and lower reaches of the Chuo-shui River, the Beigang River, the Jishui River, the Agongdian River and the Sichong River in Taiwan, and the VTG content in these prawns was determined. In addition, the p-EEQ in the waters was determined based on the VTG content, and the estradiol equivalent concentration (EEQ) in the waters was also measured immediately after sampling using the solid-phase extraction-enzyme-linked immunosorbent assay (SPE-ELISA) method. The results showed that the p-EEQ in the middle and lower reaches of the rivers in certain parts of Taiwan ranged from 38 to 400ng/L, and the detection rate was 100%. Moreover, the EEQ ranged from 7.9 to 92.9ng/L, and the detection rate was 42.9%, indicating that most of the middle and lower reaches of the rivers in Taiwan were polluted by xeno-estrogens. The 17ß-estradiol concentrations determined based on the p-EEQ were all higher than those based on the EEQ (SPE-ELISA method). The results of the present study showed that the use of M. nipponense to determine the p-EEQ in environmental waters provided advantages that included a high detection rate, high sensitivity and convenience. However, the p-EEQ cannot be used in waters that do not contain M. nipponense.

Rutting vocalizations of Formosan sika deer Cervus nippon taiouanus-acoustic structure, seasonal and diurnal variations, and individuality.

In sika deer Cervus nippon, rutting vocalizations play an important role in breeding behavior. This study investigated two types of rutting vocalizations, the moan and the howl, of the Formosan sika deer C. n. taiouanus, including the acoustic characteristics of the vocalizations, the diurnal and seasonal variations of vocal activity, and individual acoustic variation and identification. The results showed that the sound levels were approximately 81-88 dB(A) for the moan and 92-96 dB(A) for the howl, at a distance of 7 m from the sources. From October 2006 to January 2007, eight days of continuous observations were conducted to record the type and amount of vocalizations. Both moan and howl began to occur in the middle of October and reached peaks in the middle and end of November. Thereafter, few vocalizations were recorded until mid-January 2007. Moreover, we found that 74.5% of the first portion of moan, 65.3% of the second portion of moan, and 64.2% of howl could be identified on an individual basis by using discriminant analysis with 200 iterations of cross-validation test. These results revealed that the sounds differed among individuals, and also that they could be correctly identified. Our findings add to the scientific knowledge of sika deer behavior and provide the basis for a novel method of monitoring sika deer populations.

Female preference for song frequency in the cicada Mogannia formosana Matsumura (Hemiptera: Cicadidae).

In chorusing species, sound frequency has been suggested as a decisive cue for male body size in female mate choice. However, few studies on the female choice of male song frequency have been conducted in cicadas, in which males of most species sing in chorusing groups to attract females for mating. In this study, we investigated female mate choice for song frequency and body size of males of a chorusing cicada, Mogannia formosana, by phonotaxis experiments using two-choice tests and field observation, respectively. The choice proportion of effective responses and the proximity to the stimulus were used to assess the phonotaxis preference. In the field, two types of males (copulating vs. random) were caught, and their body sizes were compared. In phonotaxis experiments, the results revealed that females showed a preference for low-frequency song by approaching the low-frequency stimuli to a significantly greater proximity. By comparing the body sizes of copulating vs. random males, no significant differences were found. However, the copulating males had significantly different body shapes, as expressed by pronotum width regression by body length, from those of random males. We concluded that the sound frequency of male songs and male body shape in M. formosana can be used as cues of mate quality during female mate choice within a chorus. Additionally, we suggest that females of this species might use multiple cues for mate choice in different ranges of communication and that body size might not be the sole criterion of mate quality.

Determinants of establishment success: Comparing alien and native freshwater fishes in Taiwan.

Many parts of Asia, including Taiwan, have suffered severely from freshwater fish invasions. However, few studies using an assemblage approach have been conducted in the region so far that would help to prioritize suitable preventive actions. In this study, we focused on the invasion process from the import stage to the establishment stage, and defined establishment success as the success during this predefined process. We used datasets of freshwater fish assemblages in Taiwan to (1) compare established versus non-established alien species to distinguish the determinants of establishment success, and (2) to use these determinants to test a life history hypothesis which predicts that the magnitudes of the determinants should be significantly different between established alien species and native species. We collated a dataset for freshwater fish species which were imported into Taiwan (n = 118) of which some successfully established (n = 26), and another dataset for freshwater fish species native to Taiwan (n = 77). For each imported species, we collected data for 17 variables, including two phylogenetic, two human-use, two invasion history, and 11 life history variables. We then used decision tree methods, which have advantages in analyzing datasets with many variables of mixed types without the need to make assumptions about data distributions and input data for missing values. Our results showed that aquaculture use and maximum body length were the most important determinants for predicting establishment success of alien freshwater fish in Taiwan. Comparing five important determinants between established alien versus native species showed that the established alien species were significantly more often used in aquaculture, were associated with a higher number of established countries, and had a larger body length and greater highest water temperature tolerance than the native species. We thus conclude that our results provided evidence to support our stated hypothesis. We suggest that aquaculture use, measures of body size, and the number of previously invaded countries may alert researchers and conservation managers to species with a high establishment potential, especially for countries with similar conditions as those in Taiwan.

Acoustic preadaptation to transmit vocal individuality of savanna nightjars in noisy urban environments.

As urbanization has expanded dramatically, the impacts of urban noise on wildlife have drawn increasing attention. However, previous studies have focused primarily on diurnal songbirds and much less on nocturnal nonpasserines such as nightjars. The savanna nightjar has recently successfully colonized urban areas in Taiwan. Using 1925 calls recorded from 67 individuals, we first investigated the individual differences of the acoustic structures; and, for those acoustic variables with significant individual differences, we examined the correlation between the acoustic structures and the ambient noise levels. We then compared the transmission efficacy of vocal individuality among three sets of acoustic variables: all acoustic variables, noise-related variables, and noise-unrelated variables. Using seven artificial frequency-shifted calls to represent seven different individuals in playback-recording experiments, we also investigated the transmission efficacy of vocal individuality and variable accuracy in three different urban noise levels (high, medium, low). We found that all 30 acoustic variables derived from the acoustic structures demonstrated significant individual differences, and 14 frequency-based variables were negatively correlated with ambient noise levels. Although transmission efficacy was significantly affected by urban noise, individuality information was still transmitted with high accuracy. Furthermore, the noise-unrelated structures (which included the maximum frequency, the maximum amplitude frequency, and the mean frequency of the call) had a significantly higher transmission efficacy of vocal individuality than the noise-related variables (which included the minimum frequency, the frequency at the start and the end of the call) in both field observation and playback-recording experiments. We conclude that these noise-unrelated acoustic features may be one of the key preadaptations for this nocturnal nonpasserine to thrive so successfully in its newly adopted urban environment.

Male dimorphism in three sympatric species of Onthophagus (Coleoptera: Scarabaeidae).

Onthophagus trituber, O. taurinus, and O. proletarius, three sympatric dung beetle species native to southern Taiwan, exhibit normally distributed body size (pronotum width) but non-normally distributed horn length due to the presence of horned and hornless morphs. The scaling relationships between horn length and body size were established by using sigmoidal and segmented linear estimation, and the horned/hornless ratio in each of the species was estimated. The ratios estimated by sigmoidal curves showed that all three species were biased toward the hornless morph, whereas the ratios from segmented lines showed that only one species (O. taurinus) was biased toward the hornless morph. Nevertheless, the results of the two methods of estimation were concordant in ranking the horned morph proportions among the three species; O. proletarius had the largest proportion of horned morphs, whereas O. taurinus had the smallest, suggesting that the fewer horned morph in O. taurinus likely resulted from both intra- and inter-specific competition.

Contrasting determinants for the introduction and establishment success of exotic birds in Taiwan using decision trees models.

BACKGROUND: Biological invasions have become a major threat to biodiversity, and identifying determinants underlying success at different stages of the invasion process is essential for both prevention management and testing ecological theories. To investigate variables associated with different stages of the invasion process in a local region such as Taiwan, potential problems using traditional parametric analyses include too many variables of different data types (nominal, ordinal, and interval) and a relatively small data set with too many missing values. METHODS: We therefore used five decision tree models instead and compared their performance. Our dataset contains 283 exotic bird species which were transported to Taiwan; of these 283 species, 95 species escaped to the field successfully (introduction success); of these 95 introduced species, 36 species reproduced in the field of Taiwan successfully (establishment success). For each species, we collected 22 variables associated with human selectivity and species traits which may determine success during the introduction stage and establishment stage. For each decision tree model, we performed three variable treatments: (I) including all 22 variables, (II) excluding nominal variables, and (III) excluding nominal variables and replacing ordinal values with binary ones. Five performance measures were used to compare models, namely, area under the receiver operating characteristic curve (AUROC), specificity, precision, recall, and accuracy. RESULTS: The gradient boosting models performed best overall among the five decision tree models for both introduction and establishment success and across variable treatments. The most important variables for predicting introduction success were the bird family, the number of invaded countries, and variables associated with environmental adaptation, whereas the most important variables for predicting establishment success were the number of invaded countries and variables associated with reproduction. DISCUSSION: Our final optimal models achieved relatively high performance values, and we discuss differences in performance with regard to sample size and variable treatments. Our results showed that, for both the establishment model and introduction model, the number of invaded countries was the most important or second most important determinant, respectively. Therefore, we suggest that future success for introduction and establishment of exotic birds may be gauged by simply looking at previous success in invading other countries. Finally, we found that species traits related to reproduction were more important in establishment models than in introduction models; importantly, these determinants were not averaged but either minimum or maximum values of species traits. Therefore, we suggest that in addition to averaged values, reproductive potential represented by minimum and maximum values of species traits should be considered in invasion studies.

Interspecific comparison of traffic noise effects on dove coo transmission in urban environments.

Most previous studies concerning avian adaptation to anthropogenic noise have focused on songbirds, but few have focused on non-songbirds commonly found in urban environments such as doves. We conducted field playback-recording experiments on the perch-coos of five dove species, including four native Taiwan species (the spotted dove, Spilopelia chinensis, the oriental turtle-dove, Streptopelia orientalis, the red collared-dove, Streptopelia tranquebarica, and the emerald dove, Chalcophaps indica) and one species not native to Taiwan (the zebra dove, Geopelia striata) to evaluate the detection and recognition of dove coos in habitats with differing levels of traffic noise. Our results suggest that traffic noise has selected dominant urban species such as the spotted dove to temporally and spatially adjust cooing to reduce the masking effects of traffic noise and rare urban species such as the emerald dove to avoid areas of high traffic noise. Additionally, although the zebra dove had the highest coo frequency among the study species, its coos showed the highest detection value but not the highest recognition value. We conclude that traffic noise is an important factor in shaping the distribution of rare and dominant dove species in urban environments through its significant effects on coo transmission.

Growth, Diet Composition and Reproductive Biology of the Invasive Freshwater Fish Chevron snakehead Channa striata on a Subtropical Island.

Kuan-chung Li, Bao-sen Shieh, Yuh-wen Chiu, Da-ji Huang, and Shih-hsiung Liang (2016) The Chevron snakehead (Channa striata) has been invading Taiwan for over 30 years, and it is currently widely distributed across diverse aquatic habitats within the island. Due to its strong environmental adaptability and carnivorous diet, C. striata has caused great negative impacts to the biodiversity of native fishes and aquatic organisms in Taiwan. To effectively restrain its spatial distribution and population, the objective of this study was to investigate the growing conditions, diet composition, and reproductive biology of C. striata in the field. In total, 294 individuals were collected from wetlands, irrigation canals, streams, and reservoirs in southern Taiwan from September 2008 to December 2010. Among 272 sex-identified individuals, more females (164) were collected than males (108). The morphological differences between the sexes could not be distinguished by the 10 body measurements recorded. Diverse food items, including snails, odonates, fishes, amphibians, and reptiles, were identified in the stomachs of 35 individuals. The minimum body length of sexually mature C. striata females exhibited at a standard length of 24.5 cm (total length 28 cm). The appearance of mature oocytes were mainly observed from July to November in 2009 and from April to October in 2010. Greater absolute fecundity (oocyte/individual) was estimated in Taiwan for C. striata than in its original distribution range possibly due to less water level fluctuation in the sampling habitats of Taiwan. The relative fecundity (oocyte/g) for C. striata was considered lower but within the documented range in Taiwan when compared with its original habitat in Malaysia. To effectively manage C. striata in Taiwan, regionally eradiating young and adult individuals, especially during the reproductive season and educating people to stop releasing it in the wild are possible ways to restrain and control the further spread of this exotic fish in Taiwan.

Acoustic and temporal partitioning of cicada assemblages in city and mountain environments.

Comparing adaptations to noisy city environments with those to natural mountain environments on the community level can provide significant insights that allow an understanding of the impact of anthropogenic noise on invertebrates that employ loud calling songs for mate attraction, especially when each species has its distinct song, as in the case of cicadas. In this study, we investigated the partitioning strategy of cicada assemblages in city and mountain environments by comparing the acoustic features and calling activity patterns of each species, recorded using automated digital recording systems. Our comparison of activity patterns of seasonal and diel calling revealed that there was no significant temporal partitioning of cicada assemblages in either environment. In addition, there was no correlation between the acoustic distance based on spectral features and temporal segregation. Heterospecific spectral overlap was low in both city and mountain environments, although city and mountain cicada assemblages were subject to significantly different levels of anthropogenic or interspecific noise. Furthermore, for the common species found in both environments, the calling activity patterns at both seasonal and diel time scales were significantly consistent across sites and across environments. We suggest that the temporal calling activity is constrained by endogenous factors for each species and is less flexible in response to external factors, such as anthropogenic noise. As a result, cicada assemblages in city environments with low species diversity do not demonstrate a more significant temporal partitioning than those in mountain environments with high species diversity.

Burkholderia pseudomallei infection induces the expression of apoptosis-related genes and proteins in mouse macrophages.

BACKGROUND/PURPOSE: In this study, we addressed whether the production of apoptosis-related genes and proteins is induced in mouse macrophages infected with Burkholderia pseudomallei cells. METHODS: Mouse macrophages were infected with B. pseudomallei cells at 0.5 hours, 1 hour, 2 hours, 4 hours, and 6 hours, respectively, followed by real-time polymerase chain reaction (PCR) array analysis. The amount of apoptosis-related proteins (caspase-3, caspase -8, caspase -9, Bax, and Bcl-2) was confirmed by Western blot. RESULTS: After infection, an increase of these proteins was observed. The expression levels of other apoptosis-related genes were also determined by PCR array. Experimental results revealed that the messenger RNA levels of tumor necrosis factor ligand (e.g., tnfsf10 and tnfrs10b) and fas were increased, whereas the expression levels of some antiapoptosis genes such as Birc5, Hells, and Bnip3 were decreased. CONCLUSION: Our study results demonstrate that the apoptosis-related genes and proteins in mouse macrophages were modulated by B. pseudomallei.

Biodegradation of methyl tert-butyl ether (MTBE) by Enterobacter sp. NKNU02.

We previously isolated and identified Enterobacter sp. NKNU02 as a methyl tert-butyl ether (MTBE)-degrading bacterial strain from gasoline-contaminated water. In this study, tert-butyl alcohol, acetic acid, 2-propanol, and propenoic acid were detected using gas chromatography/mass spectrometry when MTBE was degraded by rest cells of Enterobacter sp. NKNU02 cells. We also found that biodegradation of MTBE was decreased, but not totally inhibited in mixtures of benzene, toluene, ethylbenzene and xylene. The effects of MTBE on the biology of Enterobacter sp. NKNU02 were elucidated using 2D proteomic analysis. The cytoplasmic proteins isolated from these MTBE-treated and -untreated cells were carried out for proteomic analysis. Results showed that there were 6 differential protein spots and 8 differential protein spots, respectively, as compared to their corresponding control (without MTBE addition), at the indicated incubation times when 40% and 60% of 100 mg/L of MTBE had been removed, Among these proteins, nine were successfully identified with matrix-assisted laser desorption ionization-time of flight-mass spectrometry. Proteins identified included extracellular solute-binding protein, periplasmic-binding protein ytfQ, cationic amino acid ABC transporter, isocitrate dehydrogenase, cysteine synthase A, alkyl hydroperoxide reductase (AhpC), transaldolase, and alcohol dehydrogenase. Based on these differential proteins, we discuss the bacterial responses to MTBE at the molecular level.

Cloning, expression, and characterization of cadmium-induced metallothionein-2 from the earthworms Metaphire posthuma and Polypheretima elongata.

In this study we report the sequences of MT-2 cDNA from two species of Megascoleidae earthworms, Metaphire posthuma and Polypheretima elongata, by mRNA differential display after exposure of the organisms to cadmium. Complementary (c)DNA was verified as the MT-2 gene by the characteristics of its predicted translation product, namely a high cysteine content, conserved CXC motifs, and a molecular weight of around 8 kDa. Amino acid sequence alignment revealed a conserved TKCCG in the cloned MT-2 of both megascolecid earthworms instead of the corresponding conserved TQCCG found in lumbricid earthworms. The cDNAs corresponding to the two megascolecid MT-2 genes were expressed, and the MT-2 proteins were purified for biochemical characterization. The binding of Cu2+ exhibited monophasic kinetics and those of Zn2+ and Cd2+ biphasic kinetics. The proteins bound more tightly to Cd2+ than to Zn2+ and more tightly still to Cu2+. Zn-MT and apo-MT were the most effective at scavenging free radicals, followed by Cd-MT. In conclusion, MT-2s from M. posthuma and P. elongata showed unique sequence features compared to those of lumbricid earthworms. These earthworms could be used to evaluate heavy-metal pollution in soil due to the inducible MT-2 by cadmium exposure.