Robert H. Findlay

Environmental impact of salmon net-pen culture on marine benthic communities in Maine: A case study

The environmental impacts of salmon net-pen aquaculture on the benthic environment were investigated at a commercial fish farm located in coastal Maine waters. This site has a sandy mud bottom and low current velocities, is subjected to episodic sediment resuspension, and way in production for 3 yr prior to this study: We examined both the increase in carbon flux to the benthos caused by the net-pen and the effects of the elevated flux on sediment biogeochemistry and the microbenthic communities. The experimental design involved the establishment of two study sites, an ambient site ca. 100 m from the net-pen and a treatment site around the pen. Sediment traps deployed 1 m above the sediment-water interface indicated that carbon flux to the benthos was increased 1-fold to 6-fold (to a maximum of 5 g m−2d−1) at the edge of the net-pen with little or no increase in carbon flux 10 m from the pen. Unlike carbon flux rates, sediment organic matter inventories showed a complex pattern of change over time. Mineral surface area, organic carbon and nitrogen, digestible protein, and sterol content were initially (April 1991) lower beneath the pen than in ambient sediments. During 1991 ambient sediment accumulated organic matter until July after which it decreased, to a low during November. In contrast, organic matter inventories of sediment beneath the pen remained low until July and then increased to a high during November. These latter gains were associated with the development of bacterial mats at the sediment-water interface. Beneath the pen, microbial and macrofaunal communities were shifted toward those commonly associated with organic enrichment but seasonal trends and storm-related resuspension events also significantly affected these sediment communities. When abundant, most epibenthic organisms were more numerous near the pen than in adjacent ambient areas. These results suggest that net-pen aquaculture can alter the benthic ecosystem in Maine Coastal waters but indicate that the effects are spatially limited.

Digestive environments of benthic macroinvertebrate guts: Enzymes, surfactants and dissolved organic matter

Hydrolytic enzyme activity, surfactancy, and dissolved organic matter in the digestive lumens of 19 benthic echinoderm and polychaete species were examined, using consistent and quantifiable methods. Enzyme activities were compared with those of extracellular enzymes from ambient sediments. Enzyme activities ranged over five orders of magnitude, with averages decreasing in the order polychaetes > echinoderms > sediment. Highest activities in animals were usually associated with the fluid phase in midgut sections, with posteriorward decreases indicating little export to the external environment. At some phyletic levels, activity correlated inversely with animal size. Hydrolase patterns reflected food type; for example, high 1ipase:protease ratios in carnivores reflected esterified lipids in their diets. High surfactant activity was found in gut sections having high enzyme activity. Deposit feeders had the most intense surfactancy, including evidence for micelles. While enzymes reflected the biochemical nature of the digestible food substrate regardless of feeding mode (e.g., deposit vs. suspension feeder), surfactants reflected dilution of this digestible substrate with mineral grains. Dissolved organic matter levels were high, with amino acids reaching levels > 1 M and lipids commonly 1 g L-r. Among polychaete deposit-feeders, low molecular weight amino acids reflected the composition of the food substrate, but were present at much higher concentrations than could be explained by sediment present in the gut-suggesting longer residence times for fluid than for transiting sediment particles. Deposit feeder digestive fluids are better able to solubilize sedimentary food substrates than are sedimentary extracellular enzymes, owing to either more powerful solubilizing agents or to their deployment in freely diffusing, dissolved form. Gut environments may lead to chemical condensation as well as solubilization reactions.

EFFECTS OF COMMERCIAL DRAGGING ON SEDIMENTARY ORGANIC MATTER

Abstract The effect of commercial dragging on sedimentary organic matter was examined in two field experiments using different types of gear. A heavy scallop dredge caused two types of organic matter translocation—some of the surficial organic matter was exported from the drag site and the remaining material was mixed into subsurface sediments. Phospholipid analysis indicated decreases in various classes of microbial, with relative increases in the contribution of anaerobic bacteria to the microbial community. An other trawl that largely remained above the sediment-water interface caused little change in organic matter profiles, although 7 Be profiles suggest an export of the surficial horizon. Sediment mixing by some types of gear will likely result in burial of labile organic matter and hence may shift sediment metabolism toward microbial and anaerobic food chains.

The use of a classic lipid extraction method for simultaneous recovery of organic pollutants and microbial lipids from sediments

Abstract An analytical protocol is described which allows for the determination of polycyclic aromatic hydrocarbons (PAH) and phospholipid fatty acids on the same sample. This procedure allowed for the simultaneous evaluation of PAH concentration and its biological effects on the microbial community. The total extract was separated into PAH, neutral lipid, glycolipid and phospholipid fractions on a silicic acid column by sequential elution using hexane, chloroform, acetone and methanol. This method was shown to be simple, effective and reproducible, and can be applied to very small samples (20–100 mg of sediment). PAH concentration was determined directly from the hexane fraction by GC/MS analysis without further preparative steps. Microbial community structure was simultaneously determined using phospholipid ester-linked fatty acids (PLFA). This method was evaluated based on analysis of Standard Reference Material (NIST SRM 1941a), and applied to environmental samples from Eagle Harbor, Puget Sound, Washington.

The use of phospholipid fatty acids to determine microbial community structure

The most difficult task facing microbial ecologists is the quantitative description of microbial communities. This is due to the fact that, unlike higher organisms, there is little correlation between the observable structure of a microorganism and its ecological function. Hence, the tried and true method of plant and animal ecologists, careful visual observation, is often of little value when applied to the problem of describing microbial communities. Indeed, the vast majority of microbial diversity is best described in terms of their varied metabolic functions. Unfortunately, measures of metabolic processes, while yielding important ecological information, rarely yield estimates of the biomass of microorganisms effecting the measured rate. Classical methods of isolation and identification, as well as many modern molecular methods, are highly selective — that is they fail to detect most organisms in a given environment. Only two accepted methods overcome the difficulty of selectivity and provide a relatively unbiased view of the structure of complex microbial communities. These are analysis of microbial populations using ribosomal RNA (or its corresponding DNA sequences) and phospholipid fatty acid analysis. Both techniques have advantages and disadvantages and should not be viewed as competing, rather, as complementary. Community analysis using rRNA (or rDNA) sequencing can commonly detect and identify community members with a high degree of specificity (to species and even strains), as well as, detect and suggest phylogenetic affinities of as yet uncultured organisms.

Impact of a scallop drag on the sediment chemistry, microbiota, and faunal assemblages of a shallow subtidal marine benthic community

Scallops are usually obtained by means of a heavy metal dredge that is pulled over the sea bottom. Most studies of the impact of this gear type have shown that larger invertebrates, in particular, are severely disturbed. These studies, however, have been conducted on coarse sediments, ranging from sands to cobble, and have dealt only with faunal changes. In this paper the impact of a New England type scallop dredge on the fauna and sedimentary nutritional characteristics of a silty sand community is detailed. The site, in the Damariscotta River, Maine, USA, was sampled during the fall and winter prior to, then again immediately following, the dragging event, and twice more over the ensuing six months. Loss of surficial sediment, lowered food quality of the sediment (as measured by microbial populations, enzyme hydrolysable amino acids, and chlorophyll a), and changes in the faunal composition of the dragged site were observed. While some taxa returned to the drag track relatively quickly, others such as the cumaceans, phoxocephalid and photid amphipods, and nephtyid polychaetes, were not seen in abundances comparable to those of the adjacent undragged site until the food quality also recovered.

A pilot study for monitoring changes in the microbiological component of metalworking fluids as a function of time and use in the system

This article describes the results of a pilot study to examine changes in the biological component of metalworking fluids (MWF) as a function of use. Fluid samples were taken from two newly charged systems, designated BT-7415 and BT-7707, at 1-week intervals for 8 weeks and characterized with respect to the kinds and numbers of bacteria present and presence of soluble protein in cell-free supernatants. In addition, lipid extracts of pelleted cells from fluids in BT-7415 were examined by gas chromatography/mass spectroscopy for the kinds and relative amounts of phospholipid fatty acids (PLFA) present. A total of 19 different bacterial species was cultured and identified, more than half (12/19) of which were gram-negative. Total colony-forming units (CFU) reached levels of 2.2 x 10(3)/mL in BT-7415 and 2.4 x 10(5)/mL in BT-7707. The most common genus isolated was Pseudomonas. Estimations of cell numbers based on total biomass from PLFA in samples from BT-7415 indicated 1.1 x 10(7)/mL after 8 weeks of use. Both the numbers of PLFA identified and the amounts of each detected in BT-7415 increased as the fluids were used. The chromatograms were dominated by two fatty acids, the amounts of which increased with time. These fatty acids, 18:2 omega 6 and 18:1 omega 9c, are not commonly associated with pseudomonads. This suggests that there is an important component of the biological consortium in MWF is not being detected by currently used culture techniques. There was no soluble protein detected in any of the samples from either system.

Comparison of differential plating media and two chromatography techniques for the detection of histamine production in bacteria.

The bacterial enzyme histidine decarboxylase (Hdc) catalyses the conversion of histidine into histamine. This amine is essential for the biosynthesis of iron chelators (siderophores) and is an important cause of food poisoning after consumption of fish contaminated with histamine-producing bacteria. In this work we compared different methods for detecting histamine secreted by different bacterial strains. The presence of histamine in the culture supernatant of Vibrio anguillarum, which produces Hdc and secretes the histamine-containing siderophore anguibactin, was detected by thin-layer chromatography. Similar results were obtained using the culture supernatant of the Acinetobacter baumannii 19606 prototype strain that secretes the histamine-containing siderophore acinetobactin. Conversely, histamine was not detected in the culture supernatant of an isogenic V. anguillarum Hdc mutant and the A. baumannii 8399 strain that secretes a catechol siderophore different from anguibactin and acinetobactin. These results were confirmed by capillary gas chromatography/mass spectrometry. However, all these strains tested positive for histamine secretion when cultured on differential plating media containing histidine and a pH indicator, which were specifically designed for the detection of histamine-producing bacteria. The pH increase of the medium surrounding the bacterial colonies was however drastically reduced when the histidine-containing medium was supplemented with peptone, beef extract, and glucose. The histidine-containing culture supernatants of the A. baumannii and V. anguillarum strains showed an increase of about two units of pH, turned purple upon the addition of cresol red, and contained high amounts of ammonia. Escherichia coli strains, which are Hdc negative and do not use histidine as a carbon, nitrogen, and energy source, gave negative results with the differential solid medium and produced only moderate amounts of ammonia when cultured in the presence of excess histidine. This study demonstrates that, although more laborious and requiring some expensive equipment, thin-layer and gas chromatography/mass spectrometry are more accurate than differential media for detecting bacterial histamine secretion. The results obtained with these analytical methods are not affected by byproducts such as ammonia, which are generated during the degradation of histidine and produce false positive results with the differential plating media.

Lipid Analysis of the Response of a Sedimentary Microbial Community to Polycyclic Aromatic Hydrocarbons

Polycyclic aromatic hydrocarbons (PAH) are widespread environmental contaminants that can, under proper conditions, be degraded by microorganisms. The responses of a riverine sedimentary microbial community to PAH contamination were examined using an integrated biochemical assay that yielded data on PAH concentration, total microbial biomass, and microbial community structure and were interpreted using perturbation theory and the subsidy-stress gradient. Microbial mineralization of naphthalene, anthracene, fluorene, and phenanthrene was observed 24 h after their addition to all sediments sampled and ranged from 0.9 to 16.3% in ambient sediments and from 14.8 to 35.8% in contaminated sediments. Total microbial biomass, determined by phospholipid phosphate, increased in response to intermediate PAH concentration and decreased at sites with the highest PAH concentration (p < 0.05) during seven out of nine (78%) seasonal sampling periods. The two sampling periods that were not statistically different followed periods of high water and cold temperatures. Phospholipid fatty acid analysis of microbial community structure analysis indicated that increases in the relative abundance of gram-negative aerobes and heterotrophic eukaryotes were responsible, in part, for these observed increases in total microbial biomass. These findings (increased degradation rates, increased biomass at intermediate PAH concentrations, and altered community structure) indicate that a component of the microbial community responded to PAH as a usable input and are consistent with the predictions of perturbation theory and a subsidy-stress gradient.

Periphyton growth on submerged artificial substrate as a predictor of phytoplankton response to nutrient enrichment

Net-pen aquaculture provides the large quantities of fish needed to supplement an increasing consumer demand, but it is also a source of nutrient enrichment to the surrounding ecosystem. Nutrient enrichment can result in measurable increases in phytoplankton growth, but in flowing water, such measurements are difficult and may not be appropriate. An increased awareness of the link between hypernutrification and increased prevalence of nuisance algal blooms heighten this concern. We present a method based on standard lipidology techniques that uses periphyton growth as an indicator of phytoplankton response to nutrient enrichment. Chlorophyll a (Chl a) measurements of phototrophic biomass were used to determine phytoplankton and periphyton response to nutrient stimuli and periphyton growth rates at field sites near net-pen aquaculture facilities. Laboratory trials showed that phytoplankton and periphyton communities responded similarly to both nutrient-sufficient and nutrient-limiting conditions. When sufficient nutrients were present, both phytoplankton and periphyton biomass did not increase during two-day incubations. The addition of plant nutrients during periods of nutrient limitation induced increases of 122 to 175% in phytoplankton biomass and 139 to 221% in periphyton biomass. The similarity in response suggests that the periphyton response would be an accurate descriptor of phytoplankton response. Preliminary field studies showed that reproducible periphyton growth rates (coefficient of variation <10%) were obtained at growth rates greater than 20.0 ng Chl a cm−2 day−1. Periphyton growth rates ranged from 6.0 to 80.0 ng Chl a cm−2 day−1, and were locally enhanced (2.5× that of ambient station) down-current of a freshwater net-pen.