bluuebunny/crossref_metadata_2025 huggingface dataset:
Speech rhythm and pitch patterns in Bengali: Implications for prosodic acquisition.
Authors: Khan Sameer ud Dowla , Yu Kristine , Roemer J’aime
Published: July 2011

Rhythmic timing, measured as durational variability of consonantal and vocalic intervals, is thought to be a cue for phonological chunking in infant acquisition [Ramus et al. (1999)], i.e., for learning stress-timing (English) versus syllable-timing (Spanish). Another source of rhythm that has been less studied in infant speech development is “macrorhythm” in pitch contours, regularity in f0 marking of prosodic structure [Jun (2005)]: Head-marking languages with variable contours on each word (English) are distinguished from edge-marking languages with a recurring pattern on each word (Korean). Bengali is a language with a moderately constrained syllable structure, and both head- and edge-marking: weak stress and a recurring rise on each content word [Khan, (in press)]. To explore how Bengali can be best categorized in terms of rhythmic timing and macrorhythm, and how speakers may adjust these properties to aid infant language acquisition, we analyzed various durational and intonational measurements in recordings of ten speakers reading a passage in both laboratory speech and simulated infant-directed speech (IDS) styles. Preliminary results suggest that while Bengali has syllable timing and strong macrorhythm, speakers do not highlight these rhythmic properties in IDS, but instead counteract them by increasing consonantal interval variability and interrupting the regular rising f0 patterns.
Perceptual study of Hawaiian in relation to rhythm classes.
Authors: Stojanovic Diana
Published: July 2011

The existence of language rhythm-classes has long been disputed. While early criticisms focused on disproving claims of isochronous units, later studies examined whether various durational measures are able to classify languages in the manner that corresponds to results of perceptual studies. Most recently, the evidence coming from perceptual studies is re-examined by testing additional languages and different tasks, in some cases failing to show support for rhythm-class hypothesis. Tasks and format of stimuli differ across perceptual studies; in particular, infant studies employ long speech samples while studies with adults use short stimuli. Longer samples allow generalization and may be required for a successful discrimination. The present study investigates perceptual classification of Hawaiian (classified with English based on durational metrics, but with Japanese based on syllable structure) with respect to three posited rhythm types. English, French, and Japanese were used as class representatives. Two variants of AAX task were used: In one, 60 s samples for both A and X language were presented at the beginning of the experiment. Classification results for Hawaiian are discussed in relation to the 60 s learning period and typicality of the stimuli.
Interaction of acoustic and neighborhood density parameters on spoken word recognition.
Authors: Cabbage Kathryn L. , Carrell Thomas D.
Published: July 2011

The neighborhood activation model (NAM) [Luce and Pisoni (1998)] predicts the speed and accuracy of word retrieval in the mental lexicon based on word frequency and neighborhood density. Data from NAM experiments have demonstrated processing advantages of dense words over sparse words. Little is known about how acoustic, versus phonetic, variability influences processing strategies used by listeners of acoustical-degraded speech such as those using cochlear implants and hearing instruments. In this experiment, neighborhood density was investigated using speech-like sentences which preserved detailed frequency information but removed most amplitude information. Results yielded no difference between dense and sparse words when presented in this time-varying sinusoidal format [Remez et al. (1980)]. Consistent with prior research, amplitude modulation increased overall intelligibility of the stimuli; however, the effect of neighborhood density was in the opposite direction from that found in natural speech. We hypothesize that this may be indicative of the neighborhood density effect, becoming secondary to a phonotactic probability effect which has previously been associated with sublexical processing tasks. These results indicate that interactions between acoustic and phonetic factors must be accounted for when designing hearing instruments and cochlear implants. Follow-up investigation with other acoustic variables is currently underway in our laboratory.
Lexical influences on response times in Japanese listeners’ recognition of spoken English words.
Authors: Yoneyama Kiyoko , Munson Benjamin
Published: July 2011

One index of phonological knowledge of an L2 is the magnitude of phonological neighborhood density (PND) effects on word recognition. Imai et al. [JASA (2005)] showed that effects of PND on word-recognition accuracy were only present in advanced Spanish-speaking L2 English learners and not in beginning learners. This poster presents a follow up to Imai et al.’s study using Japanese L1 acquiring English as an L2 (JL1-EL2) and Native English listeners (NELs). Previous reports of this work [Yoneyama and Munson, J. Phonetic. Soc. Japan (2010)] failed to replicate Imai et al.’s findings, in that we observed strong effects of frequency and neighborhood density on the performance of three groups of listeners: both beginning and advanced JL1-EL2 speakers, and native English speakers. Here we investigate the influence of lexical factors on reaction times (RTs) in spoken-word recognition in the same listeners. A preliminary analysis of a subset of data comparing the least-proficient JL1-EL2 speakers to the NELs shows larger effects of PND on RTs for the JL1-EL2 listeners than for the NELs. Together, the findings suggest that lexical and phonetic factors influence second-language word recognition in complex, language-specific ways. [Work supported by NSF Grant No. BCS 0729277.]
Perception of acoustic cues in contrastive focus.
Authors: Pincus Nadya
Published: July 2011

This research examines the ability of native English speakers to identify meanings associated with contrastive focus. Two conditions are compared: adjective and noun focus. Subjects hear either “I want the RED shirt” or “I want the red SHIRT,” where the focused element receives an L+H* pitch accent and a longer duration. They must then choose which of two continuations best follows the auditory stimulus (e.g., “I don’t like the blue shirt” versus “I don’t like the red pants”). Preliminary results, based on 23 subjects listening to 10 sentences each, show that despite substantial acoustic differences between the focused and nonfocused words for both the adjective and noun conditions, each condition’s result is vastly different. As a group, they excel at identifying the adjective-focused condition, but do poorly in the noun-focused condition. Closer examination reveals a bimodal distribution on the noun condition: One group has a high success rate (between 80%–100%) and another has a low success rate (between 20%–40%). For the low success group, conflicting acoustic cues may result in a misinterpretation as phrase-final lengthening and/or natural focus on the object, while these cues are being overridden in the high success group.
Effects of distal pitch and timing of speech and nonspeech precursors on word segmentation.
Authors: McAuley J. Devin , Dilley Laura C. , Rajarajan Prashanth , Bur Krista
Published: July 2011

Recent work shows that word segmentation is influenced by distal prosodic characteristics of the input several syllables from the segmentation point. However, the conditions under which these distal prosodic effects operate in segmentation remain obscure. In experiment 1, participants heard eight-syllable sequences with a lexically ambiguous four-syllable ending (e.g., crisis turnip versus cry sister nip). The duration and/or pitch characteristics of the initial five syllables were resynthesized in a manner predicted to favor parsing of the final syllables as either a monosyllabic or a disyllabic word; the acoustic characteristics of the final three syllables were held constant. Experiment 1 replicated earlier results showing that utterance-initial prosody influences segmentation utterance, finally, using different utterance-initial prosodic characteristics than shown previously. In experiment 2, the first four syllables of the eight-syllable sequences were replaced with complex tones, which maintained the pitch and timing of the original syllables. Initial results suggest that the presence of tones induced participants to adopt a more conservative response criterion but did not affect their sensitivity to distal pitch and timing cues relative to a control, speech-only condition. These findings have implications for understanding how listeners segment words from fluent speech. [Research funded by NSF Grant No. BCS-0847653.]
Boundary tones as gestures: Coordination relations at boundaries.
Authors: Katsika Argyro
Published: July 2011

While the temporal properties of prosodic boundaries are fairly well understood, tonal events at boundaries (phrase accents and boundary tones) have received less attention. The present study, conducted within the articulatory phonology framework [cf. Browman and Goldstein (1989)], is a first investigation of the coordination of boundary tone gestures to constriction gestures. An articulatory magnetometer study probes this coordination in a variety of constructions in Greek (causatives, yes-no questions, imperative requests, and wh-questions). The effects of stress (levels: zero, one or two syllables before the boundary) and accent (levels: accented versus deaccented utterance-final word) are examined. Results from two speakers analyzed to date indicate that the boundary tone coordinates with the onset’s release gesture or the nucleus’ opening gesture of the utterance-final syllable. The presence of accent does not affect these coordination relations, while stress does. Specifically, the latter affects the lag between the onset of the consonant gesture and the onset of the vowel gesture, and the lag between the onset of the consonant gesture and the onset of the boundary tone gesture. Such findings imply that boundary tones, albeit postlexical events, affect the intrasyllabic gestural coordination. An alternative possibility is that tonal and temporal events at prosodic boundaries interact. [Work supported by NIH.]
Reversible line-scan and circular synthetic aperture sonar processing with applications to object classification.
Authors: Marston Timothy M , Marston Philip L.
Published: July 2011

Spectral response as a function of aspect, often called “acoustic color,” is sometimes used to classify objects. The ability to utilize reversible imaging algorithms to enhance spectral features and to isolate the acoustic color plots of adjacent targets were previously demonstrated in the line-scan case [Marston et al., J. Acoust. Soc. Am. 128, 2461 (2010)]. Sometimes the resonant or ringing part of an object’s response is isolated from the spatially localized background. This was verified by experiments at Washington State University with a metal cylinder where the ringing part was modeled using quantitative ray theory. An overview of multiple inversion techniques and the importance of the assumptions behind the beam forming processes involved will be given. Current research additionally concerns the application of reversible circular synthetic aperture sonar algorithms to enhance our understanding of the relationship between CSAS images of objects and particular spectral features present in color plots. An imaging approach based on the Fourier-slice theorem, and the subsequent reversal process will be given, along with examples of the reversal process being used to associate object features with specific spectral features. [Work supported by ONR.]
Acoustic scattering from proud and buried unexploded ordnances in a cluttered environment.
Authors: Espana Aubrey L. , Williams Kevin L. , Kargl Steven G. , Zampolli Mario
Published: July 2011

Details of the surrounding environment, for examples, sediment conditions and nearby clutter, influence the ability to successfully detect and classify proud or buried targets. These issues were investigated during experiments conducted in March 2010 in a fresh water pond, during which targets were placed at varying distances from each other, in proud and buried configurations within a sand sediment. This paper will focus on a subset of these experiments involving the acoustic scattering from unexploded ordnances (UXOs) in proud and fully buried configurations in which the incident grazing angle of the sonar onto the water-sediment interface is above the critical angle. Monostatic synthetic aperture sonar (SAS) data will be presented for the case of a single, isolated UXO, as well as the situation where multiple UXOs are in close proximity to each other, hence simulating a cluttered environment. To supplement the data, finite element models have been developed for the UXO with varying levels of complexity in both target specifications (shape and material composition) and general experimental setup. These simulations reveal the level of fidelity required to achieve good data-model agreement. [Research supported by the ONR.]
Reversible bistatic and monostatic synthetic aperture sonar filtering.
Authors: Eastland Grant C. , Marston Timothy M. , Marston Philip L.
Published: July 2011

Recent investigations into understanding scattering features of proud and partially exposed cylinders have revealed many interesting properties of scattering mechanisms. In the present study, the effect of cylinder location was explored by lowering a horizontal solid aluminum cylinder through a flat free surface into a water tank while performing a vertical transducer line scan to monitor the evolution of the scattering. Interactions with the air-water interface partially simulate various aspects of interactions with flat sediment. The presence of the interface contributes many more paths for scattering to occur, even in the case of broadside illumination considered. Reversible SAS filtering of monostatic and bistatic data is able to identify different specular mechanisms involving the target and free surface, in addition to their corresponding elastic responses. Studying how these contribute to SAS images enables a clearer understanding of target classification and identification. It is also helpful to view the evolution of signal time-domain loci as a function of transducer location and target location. This investigation emphasizes broadside and near-broadside illumination. A delayed multiple scattering feature is visible involving one interfacial reflection and double reflected from the target. [Work supported by ONR.]
Comparisons of evanescent wave and propagating wave scattering based on circular synthetic aperture sonar and spectroscopy.
Authors: Plotnick Daniel S. , Marston Timothy M. , Marston Philip L.
Published: July 2011

A convenient way to generate acoustic evanescent waves is to use a tank containing immiscible liquids and an appropriately directed acoustic beam [Osterhoudt et al., IEEE J. Oceanic Eng. 33, 397–404 (2008)]. This is useful for exploring the nature of scattering induced by evanescent acoustic waves by suspending an object (or set of objects) close to the horizontal interface between the two liquids. In the present work the scattering is studied for a pair of metal spheres and other metallic objects using reversible circular synthetic aperture sonar in which the object (or set of objects) is spun around a vertical axis while the scattering is measured. In some cases it is also possible to investigate spectral features and make comparisons with measurements using ordinary propagating waves or the interference pattern produced by sets of such incident waves. These experiments are useful for gaining insight into scattering phenomena and signal processing options. [Work supported by ONR.]
Acoustics in support of policy.
Authors: Simmonds John
Published: July 2011

Policy in the United States and Europe requires development of targets and monitoring regimes for ocean exploitation. The current policies require a precautionary approach (PA)to exploitation, maximum sustainable yield (MSY) targets, and exploitation under ecosystem approach to fisheries management (EAFM). Currently underwater acoustics stock estimation is used as a major input for a number of fish stock assessments, and occasionally used directly to assess the state of a small number of stocks. This paper links policy requirements with management needs and explores how acoustics can deliver crucial information for environmental protection and safe exploitation. The current uses of acoustics in fisheries management are briefly reviewed, and the future requirements and uses required by these policies are explored.
Acoustic data from fishing vessels: What can be obtained that research vessels cannot offer?
Authors: Gerlotto Francois , Gutierrez Mariano , Josse Erwan Josse , Fablet Ronan
Published: July 2011

Use of fishing vessels and platforms of opportunity to collect acoustic-based population distribution and abundance data has been considered since the 1990s. The idea was that low data quality was compensated by large sampling throughout the year, and in areas where research vessels did not sample. We present results and hypotheses on the use of such data from the anchovy and jack mackerel fishing industry in Peru, where many fishing vessels are equipped with SIMRAD ES60 echosounders. First, fishing vessels are selected according to a criteria set. Once “optimal” fishing vessels are selected, we show that exploiting acoustic data from these ships may provide new information. The first is a correlation of fishing activity with fish biomass. Ee analyzed echograms preceding trawl sets. Data on fish biomass and spatial distribution were correlated with catch and catch structure. Additionally, environmental information in an area of 1 nm2 centered on the trawl location was added to the acoustic and fishing data. The potential to obtain information on fish length within the exploited school before fishing was explored, using direct (TS measurements around the school) or indirect (environmental signature giving probabilistic information) methods.
Comparison of DIDSON sonar based estimates of Chinook salmon escapement with other methods in the Coweeman River, Washington.
Authors: Rawding Dan , Liermann Martin
Published: July 2011

The Coweeman River supports a small spawning population of Tule fall Chinook salmon (200–2000 adults), which are listed for protection under the US Endangered Species Act but harvested in mixed stock Pacific Ocean and mainstem Columbia River fisheries. The Coweeman Chinook stock has a low proportion of hatchery spawners and has been used as an index population for other less intensively monitored Tule populations. Over the last decade the Washington Department of Fish and Wildlife (WDFW) and partners have funded numerous studies to determine effective methods to accurately estimate Chinook salmon escapement for this population. From 2007 to 2009, a DIDSON sonar was operated below the major spawning area to estimate escapement. Concurrently, independent escapement estimates were made using more traditional methods such as mark-recapture, red counts, and area-under-the curve surveys. Bayesian approaches to estimate salmon escapement for all methods will be presented, with an emphasis on the use of mixture models for sonar-based estimates and a comparison of the sonar estimates with more traditional approaches.
Observations of 5-MHz acoustic backscattering from Cochlodinium polykrikoides blooms in coastal waters.
Authors: Choi Jee Woong , Kim Eunhye , Na Jungyul , Kang Donhyug
Published: July 2011

Measurements of 5-MHz volume-backscattering strength of harmful algal blooms were made in summer 2008 at sites off the coast of Geumo island in southern Korea. The most abundant phytoplankton in the blooms was the chain-forming, toxic dinoflagellate Cochlodinium polykrikoides, which accounted for about 70% of total abundance. The measured backscattering strengths are compared with the abundance of phytoplankton estimated using an optical microscopy from water samples collected simultaneously with the acoustic measurements. The results show that the scattering strengths are highly correlated with dinoflagellate concentrations, suggesting that the acoustic method may be a useful tool for detecting the harmful algal blooms in their initial stage. Finally, the measurements are compared with the scattering strengths predicted by a fluid-sphere scattering model using a single-cell size of C. polykrikoides and the equivalent cell size of a sphere the same size as the chain-forming cells. The results imply that the chain-forming cells should be considered as single body with an equivalent cell size. [Work supported by the Korea Ocean Research and Development Institute.]
Estimating Chinook salmon passage in the Kenai River using dual frequency identification sonar.
Authors: Burwen Debby L. , Fleischman Steven J. , Miller James D.
Published: July 2011

Chinook salmon passage has been estimated in the Kenai River using fixed location, side-looking split-beam sonar since 1996. Estimation of Chinook salmon passage is complicated by the presence of smaller, more abundant species of salmon. Accurate estimates depend on the ability to distinguish large from small fish. Originally, a target strength threshold was used to classify fish species. However, in situ tethered-fish experiments revealed that target strength was a poor predictor of fish size for side-looking sonar. Measurements based on echo envelope length (duration) provided better predictive ability for tethered fish, but this could not be conclusively verified for free-swimming migratory fish. In this paper we describe our current efforts to estimate Chinook salmon passage in the Kenai River using dual frequency identification sonar (DIDSON). Length is manually measured from DIDSON still images, and from fish captured by gillnets drifted onsite. A species/age mixture model is fitted to the data to provide estimates of Chinook salmon passage. Passage estimates for large Chinook salmon can be produced using a simple threshold applied to DIDSON lengths.
Characterizing and monitoring marine nekton at Puget Sound’s renewable energy site.
Authors: Horne John K. , Parker-Stetter Sandra L. , Polagye Brian , Thomson Jim , Fresh Kurt L. , Hanson M. Brad
Published: July 2011

Marine hydrokinetic energy sites have the potential to impact distributions, dynamics, and abundances of macroinvertebrates, fish, and marine mammals (i.e., nekton). Potential impacts include changes in aggregation, avoidance, and occurrences of strikes or impingements. Understanding potential impacts requires knowledge of species-specific distributions over relevant spatial and temporal scales. Configurations and integration of technologies capable of providing images and data are not well-established, and the application of monitoring technologies is complicated by extreme flows at marine hydrokinetic energy sites. An echosounder, multibeam sonar, and acoustic camera will be used to detect, categorize, and enumerate nekton at a proposed renewable energy site in northern Admiralty Inlet, Puget Sound, WA. This is the site selected by Snohomish Public Utility District for the deployment of two OpenHydro turbines. Data from stationary instrument deployments will be compared to data from a mobile acoustic and midwater trawling survey to determine how well each technology captures spatiotemporal variation in nekon density distributions. Results of the design, deployment, retrieval, and analysis of data from these three instrument classes will be used to formulate recommendations for instrument choice, configuration, and the characterization and monitoring of pelagic nekton at any renewable energy site.
Estimating fish orientation from multiple view, limited angle, wide band reflections.
Authors: Jaffe Jules , Roberts Paul
Published: July 2011

Multiview, broadband (635–935 kHz), nearly monostatic, acoustic reflections recorded from lateral views of juvenile fish were used to infer animal orientation. Calibrated acoustic data were recorded from live fish in a laboratory while orientation was measured simultaneously via optical images. Using eight animals, 2-D data sets of target strength as a function of frequency and orientation were obtained. Fish length, lateral thickness, and dorsoventral thickness ranged from 24 to 48 mm, 3 to 7 mm, and 10 to 20 mm, respectively. Preliminary estimates of orientation were computed from the direction of the gradient of the local autocorrelation function in the target strength image. These local estimates were then median-filtered over the full system bandwidth (but still limited angle) to improve accuracy. Angular estimates were then corrected for systematic bias via a simple, one-demensional model that approximated the animals’ reflection by that of a bar target. Taken over all orientations, the average absolute error in orientation estimation is 5.6 to 17 deg, depending on the data set. Results indicate that, for most sets of views, reasonable estimates of lateral orientation can be obtained from broadband, multiview data over a set of limited angular reflections.
Some consideration about the management and economic aspect on the pinger use for mitigation the depredation phenomena in an MPA Mediterranean area.
Authors: Maccarrone Vincenzo , Filiciotto Francesco , Buffa Gaspare , Bellante Antonio , Vincenzo Di Stefano , Tranchida Giorgio , Buscaino Carmelo , Mazzola Salvatore , Buscaino Giuseppa
Published: July 2011

This study was carried out in a fishing area off the coast of southern Italy, where Tursiops truncatus is frequently observed to interact with bottom gill nets. The experiments aimed to assess the economic efficiency of the DDD pingers during the interaction between dolphins and monofilament bottom gill nets. A net was equipped with pingers and another without to measure the different biomass catch and costs. The economic effectiveness with an estimate of the costs/benefits was evaluated. We modeled a catch per unit effort (CPUE) both in economic terms as euros per 50 m of net set and in biomass terms as kilograms caught per 50 m of net set with and without pingers. The analysis of the data allowed us to evaluate the advantage of the economic aid provided by the European Fisheries Fund (EFF) for the use of pinger. The CPUE of the net equipped with pingers was statistically higher than the control net. The costs of investment for pingers could fully comeback in 74 days and with the economic assistance of EFF in 34 days of fishing. The use of pingers represents productive and economic benefits, even if their potential impact noise should be considered.
Calls of the black drum (Pogonias cromis: Sciaenidae): Geographical differences in sound production between Northern and Southern Hemisphere Populations.
Authors: Fine Michael L. , Tellechea Javier , Norbis Walter , Olsson Daniela
Published: July 2011

Because of apparent reproductive isolation between Northern and Southern hemisphere populations of the black drum Pogonius cromis, we tested the hypothesis that advertisement calls from a southern population would differ from known calls of North American populations. Additionally, we quantified disturbance and advertisement calls, their changes with fish size and sex, not previously examined in this species. Unlike most sciaenids, both sexes of P. cromis possess robust sonic muscles, and both produce disturbance calls when handled. However, only males produce an advertisement call used in courtship. The disturbance call consists of a variable train of short-duration pulses (average 23 ms). The duration, interpulse interval, and dominant frequency of pulses are similar in males and females and change developmentally: pulse duration and interpulse interval increase and dominant frequency decreases with fish size. Advertisement calls, recorded in the field and in captivity, are long-duration (average 184 ms) and tonal. Based on variation in fundamental frequency, which decreases with fish size, field choruses are composed of different-sized individuals. The duration of advertisement calls, about a third of those from Florida populations, suggests genetic differentiation between northern and southern populations.
A cabled acoustic telemetry system for detecting and tracking juvenile salmon.
Authors: Deng Zhiqun (Daniel) , Weiland Mark , Carlson Thomas , Eppard Brad , Fu Tao , Ploskey Gene
Published: July 2011

The juvenile salmon acoustic telemetry system (JSATS) is a nonproprietary technology developed by the U.S. Army Corps of Engineers, Portland District for detecting and tracking small fish. The JSATS consists of acoustic microtransmitters; autonomous, cabled, or portable receivers with hydrophones; and data management and processing applications. Each microtransmitter, surgically implanted in fish, transmits a unique 31-bit binary code encoded using BPSK at 416.7 kHz. Cabled systems are deployed at dams and used to determine passage-route and near-dam behavior for fish. Each cabled system is synchronized to a universal GPS clock and waveforms are saved to the computer before being decoded. Valid detections are separated from spurious detections using filtering processes requiring a minimum of six messages with a pulse interval matching that expected from properly functioning tags within a fixed period. Time-of-arrival information for valid detections on four hydrophones is used to solve for the 3-D position of tagged fish. For the cabled system at John Day Dam, the range for 3-D tracking is more than 100 m upstream of the dam face where hydrophones are deployed. Cabled systems have been successfully deployed on several major dams to acquire information for salmon protection and to develop more fish-friendly hydroelectric facilities. [This study was funded by the U.S. Army Corps of Engineers, Portland District.]
Ultrasonic control of nuisance species in commercial aquaculture settings.
Authors: Goodwiller Bradley T. , Chambers James P. , Lieblong Shannon N. , Beecham Rachel V.
Published: July 2011

One of the major challenges facing commercial catfish farmers is managing water quality and nuisance organisms. High stocking densities coupled with high feeding rates support many parasitic organisms. One such organism is the trematode that lives in an obligate parasitic relationship with another nuisance species, the ram’s horn snail. One method of controlling the trematodes is the use of copper sulfate to kill the snails. This is not an ideal solution as the copper will build up and eventually poison the entire pond. An alternate method, pursued here, is the use of high amplitude ultrasound to kill or mortally wound the snails. Since this method of removing the snails is mechanical, it reduces the amount of chemicals used to maintain the pond. To investigate the efficacy of this proposed technique, an ultrasound system was designed and built at NCPA to test the effects of various ultrasonic dosages on pathogens of interest. The system is designed to be tunable, allowing for the widest range of possible combinations of frequency and amplitude. The result of this setup is a well documented rubric for producing maximum efficacy for a given pathogen. Preliminary results and the experimental setup will be presented.
Comparative13C Metabolic Flux Analysis of Pyruvate Dehydrogenase Complex-Deficient,l-Valine-Producing Corynebacterium glutamicum
Authors: Bartek Tobias , Blombach Bastian , Lang Siegmund , Eikmanns Bernhard J. , Wiechert Wolfgang , Oldiges Marco , Nöh Katharina , Noack Stephan
Published: July 2011

l-Valine can be formed successfully usingC. glutamicumstrains missing an active pyruvate dehydrogenase enzyme complex (PDHC). Wild-typeC. glutamicumand four PDHC-deficient strains were compared by13C metabolic flux analysis, especially focusing on the split ratio between glycolysis and the pentose phosphate pathway (PPP). Compared to the wild type, showing a carbon flux of 69% ± 14% through the PPP, a strong increase in the PPP flux was observed in PDHC-deficient strains with a maximum of 113% ± 22%. The shift in the split ratio can be explained by an increased demand of NADPH forl-valine formation. In accordance, the introduction of theEscherichia colitranshydrogenase PntAB, catalyzing the reversible conversion of NADH to NADPH, into anl-valine-producingC. glutamicumstrain caused the PPP flux to decrease to 57% ± 6%, which is below the wild-type split ratio. Hence, transhydrogenase activity offers an alternative perspective for sufficient NADPH supply, which is relevant for most amino acid production systems. Moreover, as demonstrated forl-valine, this bypass leads to a significant increase of product yield due to a concurrent reduction in carbon dioxide formation via the PPP.
Characterization of Two Distinct Glycosyl Hydrolase Family 78 α-
l
-Rhamnosidases from Pediococcus acidilactici
Authors: Michlmayr Herbert , Brandes Walter , Eder Reinhard , Schümann Christina , del Hierro Andrés M. , Kulbe Klaus D.
Published: July 2011

α-
l
-Rhamnosidases play an important role in the hydrolysis of glycosylated aroma compounds (especially terpenes) from wine. Although several authors have demonstrated the enological importance of fungal rhamnosidases, the information on bacterial enzymes in this context is still limited. In order to fill this important gap, two putative rhamnosidase genes (
ram
and
ram2
) from
Pediococcus acidilactici
DSM 20284 were heterologously expressed, and the respective gene products were characterized. In combination with a bacterial β-glucosidase, both enzymes released the monoterpenes linalool and
cis
-linalool oxide from a muscat wine extract under ideal conditions. Additionally, Ram could release significant amounts of geraniol and citronellol/nerol. Nevertheless, the potential enological value of these enzymes is limited by the strong negative effects of acidity and ethanol on the activities of Ram and Ram2. Therefore, a direct application in winemaking seems unlikely. Although both enzymes are members of the same glycosyl hydrolase family (GH 78), our results clearly suggest the distinct functionalities of Ram and Ram2, probably representing two subclasses within GH 78: Ram could efficiently hydrolyze only the synthetic substrate
p
-nitrophenyl-α-
l
-rhamnopyranoside (
V
max
= 243 U mg
−1
). In contrast, Ram2 displayed considerable specificity toward hesperidin (
V
max
= 34 U mg
−1
) and, especially, rutinose (
V
max
= 1,200 U mg
−1
), a disaccharide composed of glucose and rhamnose. Both enzymes were unable to hydrolyze the flavanone glycoside naringin. Interestingly, both enzymes displayed indications of positive substrate cooperativity. This study presents detailed kinetic data on two novel rhamnosidases, which could be relevant for the further study of bacterial glycosidases.
D1/D2 Domain of Large-Subunit Ribosomal DNA for Differentiation of Orpinomyces spp
Authors: Dagar Sumit S. , Kumar Sanjay , Mudgil Priti , Singh Rameshwar , Puniya Anil K.
Published: July 2011

This study presents the suitability of D1/D2 domain of large-subunit (LSU) ribosomal DNA (rDNA) for differentiation of
Orpinomyces joyonii
and
Orpinomyces intercalaris
based on PCR-restriction fragment length polymorphism (RFLP). A variation of G/T in
O. intercalaris
created an additional restriction site for AluI, which was used as an RFLP marker. The results demonstrate adequate heterogeneity in the LSU rDNA for species-level differentiation.
Localization of the Clostridium difficile Cysteine Protease Cwp84 and Insights into Its Maturation Process
Authors: ChapetónMontes Diana , Candela Thomas , Collignon Anne , Janoir Claire
Published: July 2011

Clostridium difficile
is a nosocomial pathogen involved in antibiotic-associated diarrhea.
C. difficile
expresses a cysteine protease, Cwp84, which has been shown to degrade some proteins of the extracellular matrix and play a role in the maturation of the precursor of the S-layer proteins. We sought to analyze the localization and the maturation process of this protease. Two identifiable forms of the protease were found to be associated in the bacteria: a form of ∼80 kDa and a cleaved one of 47 kDa, identified as the mature protease. They were found mainly in the bacterial cell surface fractions and weakly in the extracellular fraction. The 80-kDa protein was noncovalently associated with the S-layer proteins, while the 47-kDa form was found to be tightly associated with the underlying cell wall. Our data supported that the anchoring of the Cwp84 47-kDa form is presumably due to a reassociation of the secreted protein. Moreover, we showed that the complete maturation of the recombinant protein Cwp84
30-803
is a sequential process beginning at the C-terminal end, followed by one or more cleavages at the N-terminal end. The processing sites of recombinant Cwp84 are likely to be residues Ser-92 and Lys-518. No proteolytic activity was detected with the mature recombinant protease Cwp84
92-518
(47 kDa). In contrast, a fragment including the propeptide (Cwp84
30-518
) displayed proteolytic activity on azocasein and fibronectin. These results showed that Cwp84 is processed essentially at the bacterial cell surface and that its different forms may display different proteolytic activities.
Pole Age Affects Cell Size and the Timing of Cell Division in Methylobacterium extorquens AM1
Authors: Bergmiller Tobias , Ackermann Martin
Published: July 2011

A number of recent experiments at the single-cell level have shown that genetically identical bacteria that live in homogeneous environments often show a substantial degree of phenotypic variation between cells. Often, this variation is attributed to stochastic aspects of biology—the fact that many biological processes involve small numbers of molecules and are thus inherently variable. However, not all variation between cells needs to be stochastic in nature; one deterministic process that could be important for cell variability in some bacterial species is the age of the cell poles. Working with the alphaproteobacterium
Methylobacterium extorquens
, we monitored individuals in clonally growing populations over several divisions and determined the pole age, cell size, and interdivision intervals of individual cells. We observed the high levels of variation in cell size and the timing of cell division that have been reported before. A substantial fraction of this variation could be explained by each cell's pole age and the pole age of its mother: cell size increased with increasing pole age, and the interval between cell divisions decreased. A theoretical model predicted that populations governed by such processes will quickly reach a stable distribution of different age and size classes. These results show that the pole age distribution in bacterial populations can contribute substantially to cellular individuality. In addition, they raise questions about functional differences between cells of different ages and the coupling of cell division to cell size.
Regulation of the Virulence Determinant OspC by
bbd18
on Linear Plasmid lp17 of Borrelia burgdorferi
Authors: Sarkar Amit , Hayes Beth M. , Dulebohn Daniel P. , Rosa Patricia A.
Published: July 2011

Persistent infection of a mammalian host by
Borrelia burgdorferi
, the spirochete that causes Lyme disease, requires specific downregulation of an immunogenic outer surface protein, OspC. Although OspC is an essential virulence factor needed by the spirochete to establish infection in the mammal, it represents a potent target for the host acquired immune response, and constitutive expression of OspC results in spirochete clearance. In this study, we demonstrate that a factor encoded on a linear plasmid of
B. burgdorferi
, lp17, can negatively regulate
ospC
transcription from the endogenous gene on the circular plasmid cp26 and from an
ospC
promoter-
lacZ
fusion on a shuttle vector. Furthermore, we have identified
bbd18
as the gene on lp17 that is responsible for this effect. These data identify a novel component of
ospC
regulation and provide the basis for determining the molecular mechanisms of
ospC
repression
in vivo
.
Characterization of Escherichia coli UmuC Active-Site Loops Identifies Variants That Confer UV Hypersensitivity
Authors: Hawver Lisa A. , Gillooly Caitlin A. , Beuning Penny J.
Published: July 2011

DNA is constantly exposed to chemical and environmental mutagens, causing lesions that can stall replication. In order to deal with DNA damage and other stresses,
Escherichia coli
utilizes the SOS response, which regulates the expression of at least 57 genes, including
umuDC
. The gene products of
umuDC
, UmuC and the cleaved form of UmuD, UmuD′, form the specialized
E. coli
Y-family DNA polymerase UmuD′
2
C, or polymerase V (Pol V). Y-family DNA polymerases are characterized by their specialized ability to copy damaged DNA in a process known as translesion synthesis (TLS) and by their low fidelity on undamaged DNA templates. Y-family polymerases exhibit various specificities for different types of DNA damage. Pol V carries out TLS to bypass abasic sites and thymine-thymine dimers resulting from UV radiation. Using alanine-scanning mutagenesis, we probed the roles of two active-site loops composed of residues 31 to 38 and 50 to 54 in Pol V activity by assaying the function of single-alanine variants in UV-induced mutagenesis and for their ability to confer resistance to UV radiation. We find that mutations of the N-terminal residues of loop 1, N32, N33, and D34, confer hypersensitivity to UV radiation and to 4-nitroquinoline-
N
-oxide and significantly reduce Pol V-dependent UV-induced mutagenesis. Furthermore, mutating residues 32, 33, or 34 diminishes Pol V-dependent inhibition of recombination, suggesting that these mutations may disrupt an interaction of UmuC with RecA, which could also contribute to the UV hypersensitivity of cells expressing these variants.
Transcriptional Hierarchy of Aeromonas hydrophila Polar-Flagellum Genes
Authors: Wilhelms Markus , Molero Raquel , Shaw Jonathan G. , Tomás Juan M. , Merino Susana
Published: July 2011

Aeromonas hydrophila
polar-flagellum class I gene transcription is σ
70
dependent, which is consistent with the fact that the
A. hydrophila
polar flagellum is constitutively expressed. In contrast to other bacteria with dual flagellar systems such as
Vibrio parahaemolyticus
, the
A. hydrophila
LafK protein does not compensate for the lack of the polar-flagellum regulator FlrA (
V. parahaemolyticus
FlaK homologue). This is consistent with the fact that the
A. hydrophila
FlrA mutation abolishes polar-flagellum formation in liquid and on solid surfaces but does not affect inducible lateral-flagellum formation. The results highlight that the polar- and lateral-flagellum interconnections and control networks are specific and that there are differences between the dual flagellar systems in
A. hydrophila
and
V. parahaemolyticus
. Furthermore, our results indicate that the
A. hydrophila
polar-flagellum transcriptional hierarchy (also in class II, III, and IV genes) shares some similarities with but has many important differences from the transcriptional hierarchies of
Vibrio cholerae
and
Pseudomonas aeruginosa
. The
A. hydrophila flhF
and
flhG
genes are essential for the assembly of a functional polar flagellum because in-frame mutants fail to swim in liquid medium and lack the polar flagellum. In
Vibrio
and
Pseudomonas flhG
disruption increases the number of polar flagella per cell, and
Pseudomonas flhF
disruption gives an aberrant placement of flagellum. Here, we propose the gene transcriptional hierarchy for the
A. hydrophila
polar flagellum.
Quorum Sensing Contributes to Natural Transformation of Vibrio cholerae in a Species-Specific Manner
Authors: Suckow Gaia , Seitz Patrick , Blokesch Melanie
Published: July 2011

Although it is a human pathogen,Vibrio choleraeis a regular member of aquatic habitats, such as coastal regions and estuaries. Within these environments,V. choleraeoften takes advantage of the abundance of zooplankton and their chitinous molts as a nutritious surface on which the bacteria can form biofilms. Chitin also induces the developmental program of natural competence for transformation in several species of the genusVibrio.In this study, we show thatV. choleraedoes not distinguish between species-specific and non-species-specific DNA at the level of DNA uptake. This is in contrast to what has been shown for other Gram-negative bacteria, such asNeisseria gonorrhoeaeandHaemophilus influenzae.However, species specificity with respect to natural transformation still occurs inV. cholerae. This is based on a positive correlation between quorum sensing and natural transformation. Using mutant-strain analysis, cross-feeding experiments, and synthetic cholera autoinducer-1 (CAI-1), we provide strong evidence that the species-specific signaling molecule CAI-1 plays a major role in natural competence for transformation. We suggest that CAI-1 can be considered a competence pheromone.
Characterization of anN-Acetylmuramic Acid/N-Acetylglucosamine Kinase of Clostridium acetobutylicum
Authors: Reith Jan , Berking Anne , Mayer Christoph
Published: July 2011

We report here the cloning and characterization of a cytoplasmic kinase ofClostridium acetobutylicum, named MurK (formurein sugarkinase). The enzyme has a unique specificity for both amino sugars of the bacterial cell wall,N-acetylmuramic acid (MurNAc) andN-acetylglucosamine (GlcNAc), which are phosphorylated at the 6-hydroxyl group. Kinetic analyses revealedKmvalues of 190 and 127 μM for MurNAc and GlcNAc, respectively, and akcatvalue (65.0 s−1) that was 1.5-fold higher for the latter substrate. Neither the non-N-acetylated forms of the cell wall sugars, i.e., glucosamine and/or muramic acid, nor epimeric hexoses or 1,6-anhydro-MurNAc were substrates for the enzyme. MurK displays low overall amino acid sequence identity (24%) with human GlcNAc kinase and is the first characterized bacterial representative of the BcrAD/BadFG-like ATPase family. We propose a role of MurK in the recovery of muropeptides during cell wall rescue inC. acetobutylicum. The kinase was applied for high-sensitive detection of the amino sugars in cell wall preparations by radioactive phosphorylation.
Biochemical studies on the production of neuraminidase by environmental isolates ofVibrio choleraenon-O1 from Bulgaria
Authors: Eneva Rumyana , Engibarov Stephan , Strateva Tanya , Abrashev Radoslav , Abrashev Ignat
Published: July 2011

Neuraminidase is a key factor in the infectious process of many viruses and pathogenic bacteria. The neuraminidase enzyme secreted by the etiological agent of cholera — Vibrio cholerae О1 — is well studied in contrast with the one produced by non-O1/non-O139 V. cholerae. Environmental non-O1/non-O139 V. cholerae isolates from Bulgaria were screened for production of neuraminidase. The presence of the neuraminidase gene nanH was detected in 18.5% of the strains. Тhe strain showing highest activity (30 U/mL), V. cholerae non-O1/13, was used to investigate the enzyme production in several media and at different aeration conditions. The highest production of extracellular neuraminidase was observed under microaerophilic conditions, which is possibly related to its role in the infection of intestine epithelium, where the oxygen content is low. On the other hand, this is another advantage of the microbe in such microaerophilic environments as sediments and lake mud. The highest production of intracellular neuraminidase was observed at anaerobic conditions. The ratio of extracellular to intracellular neuraminidase production in V. cholerae was investigated. The temperature optimum of the enzyme was determined to be 50 °C and the pH optimum to be 5.6–5.8.