Role of the Digestive Gland in Ink Production in Four Species of Sea Hares: An Ultrastructural Comparison

The ultrastructure of the digestive gland of several sea hare species that produce different colored ink (Aplysia californica produces purple ink, A. juliana white ink, A. parvula both white and purple ink, while Dolabrifera dolabrifera produces no ink at all) was compared to determine the digestive gland’s role in the diet-derived ink production process. Rhodoplast digestive cells and their digestive vacuoles, the site of digestion of red algal chloroplast (i.e., rhodoplast) in A. californica, were present and had a similar ultrastructure in all four species. Rhodoplast digestive cell vacuoles either contained a whole rhodoplast or fragments of one or were empty. These results suggest that the inability to produce colored ink in some sea hare species is not due to either an absence of appropriate digestive machinery, that is, rhodoplast digestive cells, or an apparent failure of rhodoplast digestive cells to function. These results also propose that the digestive gland structure described herein occurred early in sea hare evolution, at least in the common ancestor to the genera Aplysia and Dolabrifera. Our data, however, do not support the hypothesis that the loss of purple inking is a synapomorphy of the white-ink-producing subgenus Aplysia

Ultrastructural Comparison of Processing of Protein and Pigment in the Ink Gland of Four Species of Sea Hares

The ink glands of four sea hare species (Aplysia californica, A. parvula, A. juliana, and Dolabrifera dolabrifera) were compared to determine where ink protein is synthesized, how it is incorporated into protein storage vesicles, and the degree of variation in the structure of the ink gland. Ink protein was synthesized in RER cells and stored in amber and white vesicles. Lack of competent RER cells in the ink gland of D. dolabrifera was correlated with the absence of ink protein. Ink protein had similar characteristics in all three Aplysia species but, again, it was absent in D. dolabrifera. Its uptake involved pinocytosis by protein vesicle cell membranes. Granulate cells showed little variation in structure among the four species, the opposite was the case for RER cells. The conversion of the red algal pigment, phycoerythrin, to phycoerythrobilin (PEB) occurs in the digestive gland but the change of PEB to aplysioviolin (APV), the form of pigment released by the ink gland, occurs in the ink gland itself by both granulate cells and pigment vesicles. The literature describes five types of vesicles based upon color and contents in the ink gland of these four species. We report only three types of vesicle: colored (purple), protein (white and amber), and transparent (includes clear vesicles)

Hearing Loss in Stranded Odontocete Dolphins and Whales

The causes of dolphin and whale stranding can often be difficult to determine. Because toothed whales rely on echolocation for orientation and feeding, hearing deficits could lead to stranding. We report on the results of auditory evoked potential measurements from eight species of odontocete cetaceans that were found stranded or severely entangled in fishing gear during the period 2004 through 2009. Approximately 57% of the bottlenose dolphins and 36% of the rough-toothed dolphins had significant hearing deficits with a reduction in sensitivity equivalent to severe (70–90 dB) or profound (>90 dB) hearing loss in humans. The only stranded short-finned pilot whale examined had profound hearing loss. No impairments were detected in seven Risso's dolphins from three different stranding events, two pygmy killer whales, one Atlantic spotted dolphin, one spinner dolphin, or a juvenile Gervais' beaked whale. Hearing impairment could play a significant role in some cetacean stranding events, and the hearing of all cetaceans in rehabilitation should be tested

Robust estimation of bacterial cell count from optical density

Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data

Multi-component chemical defense in seahares (Gastropoda:Opisthobranchia): antipredator compounds act as both honest and deceptive signals to multiple predator species

Thesis (Ph. D.)--University of Washington, 2002Many organisms produce chemical defenses to deter predation, yet the behavioral, neurophysiological, and cellular mechanisms of chemical defenses are largely unexplored. Animals do not face a single hypothetical predator, but rather a diversity of predators whose behaviors and sensory systems can be highly divergent. Do prey chemical defenses function differently versus different predator species? Can chemical defenses be honest signals to some species and deceitful to others? These issues were examined using seahares (Aplysia spp. and Stylocheilus spp.) and their cnidarian, crustacean, and vertebrate predators. Seahares release defensive secretions called ink and opaline from independent glands. In behavioral assays, seahares with full glands versus those with depleted glands had a significant survival advantage against sea anemones, crabs, and lobsters, but not against fishes. Isolated secretions and their components were used to explore mechanisms of defense against sea anemones and lobsters. A 60 kDa glycoprotein from ink was isolated, cloned, sequenced, and expressed. This protein ("escapin") was responsible for the aversive reaction of sea anemones to ink via lysis of anemone cells. It also had antibacterial effects against Gram positive and negative bacteria. Escapin is the first reported antipredator protein of any organism.Ink and opaline protected seahares against crustaceans in a different and unusual way---by stimulating them to feed. Opaline was as stimulatory as homogenates of squid and shrimp in behavioral studies and electrophysiological assays of chemosensory neurons of lobsters. The attractiveness of opaline and ink is likely due to extremely high levels of free amino acids. In particular the highly stimulatory amino acid, taurine, is three orders of magnitude more concentrated in opaline than seahare haemolymph. These secretions are a supernormal feeding stimulus that can act as a sensory trap, exploiting the chemosensory systems of crustaceans. Via this novel chemical defense "phagomimicry", crustaceans are deceived into attending to a false food stimulus from the secretions while dropping the seahare and thereby incurring a nutritional cost. Thus chemical defenses can function differently versus different predator species, acting as honest signals (as escapin does to sea anemones) to some and deceitful signals (as phagomimicry does to crustaceans) to others

Dependence Flow Graphs: An Algebraic Approach to Program Dependencies

The topic of intermediate languages for optimizing and parallelizing compilers has received much attention lately. In this paper, we argue that any good representation of a program must have two crucial properties: first, it must be a data structure that can be rapidly traversed to determine dependence information, and second this representation must be a program in its own right, with a parallel, local model of execution. In this paper, we illustrate the importance of these points by examining algorithms for a standard optimization --- global constant propagation. We discuss the problems in working with current representations. Then, we propose a novel representation called the dependence flow graph which has each of the properties mentioned above. We show that this representation leads to a simple algorithm, based on abstract interpretation, for solving the constant propagation problem. Our algorithm is simpler than, and as efficient as, the best known algorithms for this problem. An..

Network Analysis in Disorders of Consciousness: Four Problems and One Proposed Solution (Exponential Random Graph Models).

In recent years, the study of the neural basis of consciousness, particularly in the context of patients recovering from severe brain injury, has greatly benefited from the application of sophisticated network analysis techniques to functional brain data. Yet, current graph theoretic approaches, as employed in the neuroimaging literature, suffer from four important shortcomings. First, they require arbitrary fixing of the number of connections (i.e., density) across networks which are likely to have different "natural" (i.e., stable) density (e.g., patients vs. controls, vegetative state vs. minimally conscious state patients). Second, when describing networks, they do not control for the fact that many characteristics are interrelated, particularly some of the most popular metrics employed (e.g., nodal degree, clustering coefficient)-which can lead to spurious results. Third, in the clinical domain of disorders of consciousness, there currently are no methods for incorporating structural connectivity in the characterization of functional networks which clouds the interpretation of functional differences across groups with different underlying pathology as well as in longitudinal approaches where structural reorganization processes might be operating. Finally, current methods do not allow assessing the dynamics of network change over time. We present a different framework for network analysis, based on Exponential Random Graph Models, which overcomes the above limitations and is thus particularly well suited for clinical populations with disorders of consciousness. We demonstrate this approach in the context of the longitudinal study of recovery from coma. First, our data show that throughout recovery from coma, brain graphs vary in their natural level of connectivity (from 10.4 to 14.5%), which conflicts with the standard approach of imposing arbitrary and equal density thresholds across networks (e.g., time-points, subjects, groups). Second, we show that failure to consider the interrelation between network measures does lead to spurious characterization of both inter- and intra-regional brain connectivity. Finally, we show that Separable Temporal ERGM can be employed to describe network dynamics over time revealing the specific pattern of formation and dissolution of connectivity that accompany recovery from coma

Dependence Flow Graphs: An Algebraic Approach to Program Dependencies

The topic of intermediate languages for optimizing and parallelizing compilers has received much attention lately. In this paper, we argue that any good representation of a program must have two crucial properties: first, it must be a data structure that can be rapidly traversed to determine dependence information, and second this representation must be a program in its own right, with a parallel, local model of execution. In this paper, we illustrate the importance of these points by examining algorithms for a standard optimization --- global constant propagation. We discuss the problems in working with current representations. Then, we propose a novel representation called the dependence flow graph which has each of the properties mentioned above. We show that this representation leads to a simple algorithm, based on abstract interpretation, for solving the constant propagation problem. Our algorithm is simpler than, and as e#cient as, the best known algorithms for this problem. An interesting feature of our representation is that it naturally incorporates the best aspects of many other representations, including continuation-passing style, data and program dependence graphs, static single assignment form and dataflow program graphs

Users Get Routed: Traffic Correlation on Tor by Realistic Adversaries

We present the first analysis of the popular Tor anonymity network that indicates the security of typical users against reasonably realistic adversaries in the Tor network or in the underlying Internet. Our results show that Tor users are far more susceptible to compromise than indicated by prior work. Specific contributions of the paper include (1) a model of various typical kinds of users, (2) an adversary model that includes Tor network relays, autonomous systems (ASes), Internet exchange points (IXPs), and groups of IXPs drawn from empirical study, (3) metrics that indicate how secure users are over a period of time, (4) the most accurate topological model to date of ASes and IXPs as they relate to Tor usage and network configuration, (5) a novel realistic Tor path simulator (TorPS), and (6) analyses of security making use of all the above. To show that our approach is useful to explore alternatives and not just Tor as currently deployed, we also analyze a published alternative path selection algorithm, Congestion-Aware Tor. We create an empirical model of Tor congestion, identify novel attack vectors, and show that it too is more vulnerable than previously indicated