William F. Gilly

Oceanographic and Biological Effects of Shoaling of the Oxygen Minimum Zone

Long-term declines in oxygen concentrations are evident throughout much of the ocean interior and are particularly acute in midwater oxygen minimum zones (OMZs). These regions are defined by extremely low oxygen concentrations (<20-45 μmol kg(-1)), cover wide expanses of the ocean, and are associated with productive oceanic and coastal regions. OMZs have expanded over the past 50 years, and this expansion is predicted to continue as the climate warms worldwide. Shoaling of the upper boundaries of the OMZs accompanies OMZ expansion, and decreased oxygen at shallower depths can affect all marine organisms through multiple direct and indirect mechanisms. Effects include altered microbial processes that produce and consume key nutrients and gases, changes in predator-prey dynamics, and shifts in the abundance and accessibility of commercially fished species. Although many species will be negatively affected by these effects, others may expand their range or exploit new niches. OMZ shoaling is thus likely to have major and far-reaching consequences.

Intraspecific variation of venom injected by fish-hunting Conus snails.

SUMMARY Venom peptides from two species of fish-hunting cone snails (Conus striatus and Conus catus) were characterized using microbore liquid chromatography coupled with matrix-assisted laser desorption/ionization-time of flight-mass spectrometry and electrospray ionization-ion trap-mass spectrometry. Both crude venom isolated from the venom duct and injected venom obtained by milking were studied. Based on analysis of injected venom samples from individual snails, significant intraspecific variation (i.e. between individuals) in the peptide complement is observed. The mixture of peptides in injected venom is simpler than that in the crude duct venom from the same snail, and the composition of crude venom is more consistent from snail to snail. While there is animal-to-animal variation in the peptides present in the injected venom, the composition of any individuals injected venom remains relatively constant over time in captivity. Most of the Conus striatus individuals tested injected predominantly a combination of two neuroexcitatory peptides (s4a and s4b), while a few individuals had unique injected-venom profiles consisting of a combination of peptides, including several previously characterized from the venom duct of this species. Seven novel peptides were also putatively identified based on matches of their empirically derived masses to those predicted by published cDNA sequences. Profiling injected venom of Conus catus individuals using matrix-assisted laser desorption/ionization-time of flight-mass spectrometry demonstrates that intraspecific variation in the mixture of peptides extends to other species of piscivorous cone snails. The results of this study imply that novel regulatory mechanisms exist to select specific venom peptides for injection into prey.

Natural egg mass deposition by the Humboldt squid ( Dosidicus gigas ) in the Gulf of California and characteristics of hatchlings and paralarvae

The jumbo or Humboldt squid, Dosidicus gigas, is an important fisheries resource and a significant participant in regional ecologies as both predator and prey. It is the largest species in the oceanic squid family Ommastrephidae and has the largest known potential fecundity of any cephalopod, yet little is understood about its reproductive biology. We report the first discovery of a naturally deposited egg mass of Dosidicus gigas, as well as the first spawning of eggs in captivity. The egg mass was found in warm water (25‐278C) at a depth of 16 m and was far larger than the egg masses of any squid species previously reported. Eggs were embedded in a watery, gelatinous matrix and were individually surrounded by a unique envelope external to the chorion. This envelope was present in both wild and captive-spawned egg masses, but it was not present in artificially fertilized eggs. The wild egg mass appeared to be resistant to microbial infection, unlike the incomplete and damaged egg masses spawned in captivity, suggesting that the intact egg mass protects the eggs within. Chorion expansion was also more extensive in the wild egg mass. Hatchling behaviours included proboscis extension, chromatophore activity, and a range of swimming speeds that may allow them to exercise some control over their distribution in the wild.

Ontogeny of Copepod Predation in Juvenile Squid (Loligo opalescens)

Copepods are the major prey of juvenile squid, and small species of squid such as Loligo opalescens face a great challenge in catching these erratically moving crustaceans. We studied the ontogeny of copepod predation in laboratory-reared animals and found that mastery of copepod capture develops progressively, starting shortly after hatch with strong attacks of a simple type. Modifications of the initial basic attack lead to more specialized strategies that effectively extend the range of capture to both longer and shorter distances. This progression culminates, by approximately 40 days post-hatching, in adult-like prey capture behavior involving tentacle extension and retraction. Squid raised exclusively on easily captured Artemia nauplii and introduced to a copepod diet 40 days after hatching displayed only basic attack behavior, characteristic of very young squid. All of these attacks were unsuccessful, and very few of these animals survived the transition. Copepod capture thus appears to be a skill that must be acquired in an experience-dependent manner early in post-hatching life.

Anatomical correlates of venom production in Conus californicus

Like all members of the genus, Conus californicus has a specialized venom apparatus, including a modified radular tooth, with which it injects paralyzing venom into its prey. In this paper the venom duct and its connection to the pharynx, along with the radular sac and teeth, were examined using light and transmission electron microscopy. The general anatomy of the venom apparatus resembles that in other members of the genus, but several features are described that have not been previously reported for other species. The proximal (posterior) quarter of the venom duct is composed of a complex epithelium that may be specialized for active transport rather than secretion. The distal portion of the duct is composed of a different type of epithelium, suggestive of holocrine secretion, and the cells display prominent intracellular granules of at least two types. Similar granules fill the lumen of the duct. The passageway between the lumen of the venom duct and pharynx is a flattened branching channel that narrows to a width of 10 μm and is lined by a unique cell type of unknown function. Granular material similar to that in the venom duct was also found in the lumen of individual teeth within the radular sac. Mass spectrometry (MALDI-TOF) demonstrated the presence of putative peptides in material derived from the tooth lumen, and all of the more prominent species were also evident in the anterior venom duct. Radular teeth thus appear to be loaded with peptide toxins while they are still in the radular sac.

Diversity of conotoxin types from Conus californicus reflects a diversity of prey types and a novel evolutionary history

Most species within the genus Conus are considered to be specialists in their consumption of prey, typically feeding on molluscs, vermiform invertebrates or fish, and employ peptide toxins to immobilize prey. Conus californicus Hinds 1844 atypically utilizes a wide range of food sources from all three groups. Using DNA- and protein-based methods, we analyzed the molecular diversity of C. californicus toxins and detected a correspondingly large number of conotoxin types. We identified cDNAs corresponding to seven known cysteine-frameworks containing over 40 individual inferred peptides. Additionally, we found a new framework (22) with six predicted peptide examples, along with two forms of a new peptide type of unusual length. Analysis of leader sequences allowed assignment to known superfamilies in only half of the cases, and several of these showed a framework that was not in congruence with the identified superfamily. Mass spectrometric examination of chromatographic fractions from whole venom served to identify peptides corresponding to a number of cDNAs, in several cases differing in their degree of posttranslational modification. This suggests differential or incomplete biochemical processing of these peptides. In general, it is difficult to fit conotoxins from C. californicus into established toxin classification schemes. We hypothesize that the novel structural modifications of individual peptides and their encoding genes reflect evolutionary adaptation to prey species of an unusually wide range as well as the large phylogenetic distance between C. californicus and Indo-Pacific species.

Post-Hatching Development of Circular Mantle Muscles in the Squid Loligo opalescens

Post-hatching development of the circular muscles in the mantle of squid was studied morphometrically to identify structural changes and to quantify hyperplasia and hypertrophy of the muscle fibers. Superficial, mitochondria-rich (SMR) fibers and central, mitochondria-poor (CMP) fibers are present at hatching. Although both fiber types increase in size and, even more so, in number during post-hatching development, CMP fibers increase at a much higher rate than do SMR fibers. As a result, the relative proportion of SMR to CMP fibers shifts from about 1:1 in a hatchling to about 1:6 in an 8-week-old animal; it then apparently remains constant to adulthood. These structural changes are consistent with developmental changes in muscular activity. During slow, jet-propelled swimming, 1-week-old animals show mantle contractions that have twice the relative amplitude and frequency of those in adults. The presence of Na-channel protein in mantle muscle was detected bio-chemically by using site-directed antibodies; the protein was found to be preferentially expressed in CMP fibers. These results suggest that SMR fibers are an important source of locomotory power at hatching, but become progressively less important during the first 8 weeks of development as CMP fibers assume the dominant role in jet locomotion.

Controlled and in situ target strengths of the jumbo squid Dosidicus gigas and identification of potential acoustic scattering sources

This study presents the first target strength measurements of Dosidicus gigas, a large squid that is a key predator, a significant prey, and the target of an important fishery. Target strength of live, tethered squid was related to mantle length with values standardized to the length squared of -62.0, -67.4, -67.9, and -67.6 dB at 38, 70, 120, and 200 kHz, respectively. There were relatively small differences in target strength between dorsal and anterior aspects and none between live and freshly dead squid. Potential scattering mechanisms in squid have been long debated. Here, the reproductive organs had little effect on squid target strength. These data support the hypothesis that the pen may be an important source of squid acoustic scattering. The beak, eyes, and arms, probably via the sucker rings, also play a role in acoustic scattering though their effects were small and frequency specific. An unexpected source of scattering was the cranium of the squid which provided a target strength nearly as high as that of the entire squid though the mechanism remains unclear. Our in situ measurements of the target strength of free-swimming squid support the use of the values presented here in D. gigas assessment studies.

Locomotion and behavior of Humboldt squid, Dosidicus gigas , in relation to natural hypoxia in the Gulf of California, Mexico

SUMMARY We studied the locomotion and behavior of Dosidicus gigas using pop-up archival transmitting (PAT) tags to record environmental parameters (depth, temperature and light) and an animal-borne video package (AVP) to log these parameters plus acceleration along three axes and record forward-directed video under natural lighting. A basic cycle of locomotor behavior in D. gigas involves an active climb of a few meters followed by a passive (with respect to jetting) downward glide carried out in a fins-first direction. Temporal summation of such climb-and-glide events underlies a rich assortment of vertical movements that can reach vertical velocities of 3 m s−1. In contrast to such rapid movements, D. gigas spends more than 80% of total time gliding at a vertical velocity of essentially zero (53% at 0±0.05 m s−1) or sinking very slowly (28% at −0.05 to −0.15 m s−1). The vertical distribution of squid was compared with physical features of the local water column (temperature, oxygen and light). Oxygen concentrations of ≤20 μmol kg−1, characteristic of the midwater oxygen minimum zone (OMZ), can influence the daytime depth of squid, but this depends on location and season, and squid can ‘decouple’ from this environmental feature. Light is also an important factor in determining daytime depth, and temperature can limit nighttime depth. Vertical velocities were compared over specific depth ranges characterized by large differences in dissolved oxygen. Velocities were generally reduced under OMZ conditions, with faster jetting being most strongly affected. These data are discussed in terms of increased efficiency of climb-and-glide swimming and the potential for foraging at hypoxic depths.

Control of the spatial distribution of sodium channels in giant fiber lobe neurons of the squid.

Na+ channels are present at high density in squid giant axon but are absent from its somata in the giant fiber lobe (GFL) of the stellate ganglion. GFL cells dispersed in vitro maintain growing axons and develop a Na+ channel distribution similar to that in vivo. Tunicamycin, a glycosylation inhibitor, selectively disrupts the spatially appropriate, high level expression of Na+ channels in axonal membrane but has no effect on expression in cell bodies, which show low level, inappropriate expression in vitro. This effect does not appear to involve alteration in Na+ channel turnover or axon viability. K+ channel distribution is unaffected. Thus, glycosylation appears to be involved in controlling Na+ channel localization in squid neurons.