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New Jersey Academy of Science |
2002 Senior Academy Abstracts |
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A METABOLIC INHIBITION TEST TO DETECT THE CIDAL OR STATIC EFFECTS OF ANTIMICROBIAL AGENTS. Joseph A. Adamo, Science Department, Ocean County College, Toms River, NJ 08753, and Biology Department, Georgian Court College, Lakewood, NJ 08701 and Michael A. Gealt, Department of Biological Science, Purdue University, Calumet, Hammond, IN 46233. . The usefulness of 1 mL microfuge tube cultures utilizing phenol red dextrose broth (PRDB) for a metabolic inhibition test (MIT) was demonstrated earlier. These tests (MIT) were used to determine bacteriophage titers, antibiotic sensitivity, bacterial phenotypes and the generation of transconjugants that originated in the digestive tract of microscopic nematode vectors. In the current study we attempted to establish a rapid MIT using PRDB to determine the cidal or static effect of an antibiotic. A wide range of bacterial species and the fungus Candida albicans were screened for growth in PRDB. These included gram positives and negatives, spore formers and non-spore formers. The antibiotics included Ampicillin (50mg/mL), Chloramphenicol (30mg/mL), Kanamycin (50mg/mL), Nalidixic Acid (20mg/mL), Streptomycin (25mg/mL) and Tetracycline (125mg/mL). Microfuge tubes were loaded with 1 mL PRDB with or without antibiotic and inoculated with 50-100mL of bacteria from an overnight culture. Cultures were incubated at the appropriate temperature for the particular organism. Replicating microbes generating acid caused the medium to change from a red color to a distinctive yellow. The post antibiotic treatment (PAT) consisted of centrifugation of the tubes at 14,000 rpm for 5 min in a microfuge after which the supernatant was carefully removed and fresh PRDB (without antibiotic) was added. In all cases resistant organisms showed positive growth PAT. Susceptable organisms showed positive growth PAT when the antibiotic had a static effect and negative growth when cidal.
USING BIOINFORMATICS TO CHARACTERIZE A TRUNCATED HUMAN HOMOLOGUE OF THE Drosophila Groucho GENE, hAES. Daniel Broy (student), Louis A. Scala, Department of Biology, Monmouth University, West Long Branch, NJ 07764 Screening of a human neuroblastoma cell cDNA library resulted in the cloning of a cDNA encoding a 197 amino acid polypeptide, human amino-terminal enhancer of split (hAES). The hAES gene is a variant of the Drosophila groucho and human transducin-like enhancer of split (TLE) genes. The hAES protein is similar to the groucho and TLEs in their amino-terminal domains, but lacks 500 carboxyl-termini residues including putative functional domains, such as the "WD-40" motif. We used the cDNA sequence to probe the human genome database. We had the following objectives, to localize the hAES gene within the human genome, to determine the intron/exon positions and arrangements, and to determine whether hAES is a splice variant of TLE genes. We also used genomic sequence 5’ to the start site of hAES to search databases for putative promoter sequences for insight into the regulation of this gene. Our findings indicate that hAES is located between nucleotides 28,788 and 37,892 of chromosome 19 and is processed by splicing seven exons. Chromosomal location indicates that hAES is not a splice variant of TLE genes and likely evolved from a common ancestor as a member of this gene family. The search to identify promoters of the hAES gene has revealed the presence of only the most common eukaryotic promoter. A SURVEY OF GENES EXPRESSED IN THE ICE WORM, MESENCHYTRAEUS SOLIFUGUS Angela H. Farrell (student) and Daniel H. Shain. Biology Department, Rutgers University, Camden, NJ 08102 The ice worm, Mesenchytraeus solifugus, is the only known annelid that survives exclusively in glacier ice and snow. One central question relates to the molecular evolution of gene(s) that have permitted this evolutionary step. Beginning with total RNA purified from ~50 ice worm heads, we constructed a cDNA library using SmartTM (Clonetech) methodology. A differential screen with a tail cDNA probe (i.e., cDNA from ice worm bodies lacking heads) generated multiple cDNAs that were subjected to DNA sequencing analysis. Surprisingly, many cDNAs encoded genes that are associated with energy production (e.g.., cytochrome oxidase, NAD dehydrogenase, etc.). Other genes (e.g., fibronectin) displayed significant modifications in their predicted linear amino acid sequences. Collectively, these results identify candidate genes that may have contributed to ice worm adaptation. IDENTIFICATION OF STEM CELL-SPECIFIC GENES IN LEECH. Kristi A. Hohenstein (student) and Daniel H. Shain. Biology Department, Rutgers University, Camden, NJ 08102 The glossiphoniid leech, Theromyzon rude, displays particularly large embryonic stem cells during its early development. Among these are the bilaterally-paired mesodermal (M) and neuroectodermal (N) stem cells that give rise to the bulk of segmental tissue in an adult leech. In an effort to identify differences in gene expression between these cell types, pure populations of M and N and their precursors DM and NOPQ, respectively, were dissected from developing leech embryos. Following total RNA purification, cDNA was synthesized independently from each cell type using SmartTM methodology (Clonetech), which incorporates a PCR-amplification step. Differential Display analyses were then conducted simultaneously with cDNA from all cell types. To date, ~2,000 cDNA fragments have been displayed using more than 40 primer sets. Of these, >98% are expressed in all four cell types and are likely “housekeeping” genes. Approximately 50 cDNA fragments are differentially expressed and fall into the following categories: mesoderm-specific (i.e., DM & M; 14); neuroectoderm-specific (i.e., NOPQ & N; 12); precursor-specific (i.e., DM & NOPQ; 6); stem cell-specific (i.e., M & N; 3); M-specific (2); and N-specific (11). Surprisingly, several bands (6) appear only in the DM and NOPQ precursor cells, suggesting that some mRNAs may be degraded to facilitate stem cell differentiation. These and other bands are currently being excised for DNA sequence analysis.
MOLECULAR PROPERTIES OF THE LEECH COCOON. Tarin A. Mason (student) and Daniel H. Shain. Biology Department, Rutgers University, Camden, NJ 08102 The aquatic leech, Theromyzon rude, spins cocoons underwater as it lays it eggs. Cocoons form a sleeve around the body into which eggs are deposited; cocoons are then passed over the head and sealed at both ends. We are interested in determining the amino acid composition and a partial amino acid sequence of the T.rude cocoon protein in an effort to clone its respective gene. Our initial attempts to solubilize the cocoon in strong denaturing agents including guanidinium, urea and a variety of organic solvents proved unsuccessful, suggesting that the cocoon is unusually chemically resilient. Hydrolysis with hydrochloric acid yielded an amino acid composition comprising ~12% aspartic acid, ~12% glycine, ~10% serine, ~10% proline and ~10% threonine. Partial hydrolysis with 10% acetic acid at 50蚓 generated two peptide fragments, ~35 kD and ~40 kD, that were resolved by SDS-PAGE. These were transferred onto PVDF membrane and micro-sequenced by Edmann degradation. Results from this analysis suggest that the amino termini of both fragments contain the identical seven amino sequence Ala-Ile-Asp-Glu-Ala-Ile-Lys. Surprisingly, the sequencing reaction deteriorated at the 8th position, which may relate to a folding pattern associated with its inherent chemical resiliency. Following our initial interest, a degenerate DNA oligonucleotide corresponding to the aforementioned peptide was synthesized and is currently being used to screen a genomic Southern blot. The appropriate DNA fragments will be excised and utilized for constructing a sub-genomic library in an effort to clone the T. rude cocoon gene.
AN ULTRASTRUCTURAL COMPARISON OF THE ANNELIDS, ENCHYTRAEIDAE AND HIRUDINAE Mechelle D. Regester (student) and Daniel H. Shain. Biology Department, Rutgers University, Camden, NJ 08102 The annelids Enchytraeus albidus (family: Enchytraeidae), Mesenchytraeus solifugus (family: Enchytraeidae) and Theromyzon rude (class: Hirudinae) diverged recently in evolution but occupy remarkably diverse habitats. E. albidus is a terrestrial worm that is braodly distributed; M. solifugus inhabits glacial ice along the Pacific coast of North America; and T. rude is an aquatic leech found in North and South America. By transmission electron microscopy (TEM), we have examined morphological features at the anterior end of each worm in an effort to identify similarities and differences that may contribute to their respective habitats and lifestyles. Of particular interest are sensory cells, mitochondria and the morphology of their respective head pores.
PRELIMINARY
EXAMINATION OF THE EARLY LIFE HISTORY OF THE GOOSEFISH Lophius
americanus. Peter
J. Clarke,
Kenneth W. Able (Rutgers University, Institute of Marine and Coastal Sciences, Tuckerton,
NJ), R. Christopher Chambers and David A. Witting (NOAA, National Marine Fisheries
Service, Sandy Hook, NJ) In recent years goosefish, Lophius americanus, has become an important commercial species throughout the mid Atlantic and New England region. Despite its increasing importance, little is known about its biology, and especially the important features of its early life history. Our research is directed towards the stage-specific pattern of occurrence of young goosefish. Analyses of collections and specimens (larva and juveniles to 120 mm TL) supports the following preliminary conclusions: 1) reproduction occurs in spring, larvae are distributed from the surf zone to the edge of the continental shelf and perhaps extending in to waters on the upper slope, 2) larvae are found in abundance near New Jersey beaches from May through July peaking in June, larvae have an extended pelagic period and attain sizes of approximately 12 mm TL before fin rays are complete, and 3) morphological characters continue to change in fish up to 40 to 50 mm TL. We suspect that the cessation of radical morphological change at 40 to 50 mm TL is concurrent with the settlement of individuals to the sea bottom. Post-settlement (demersal) individuals appear to be distributed widely over the entire continental shelf from the Gulf of Maine to Cape Hatteras. Goosefish appear to reach sizes of approximately 120 mm TL by the end of their first year. Future work will extend these analyses of goosefish and will include an otolith-based examination of age and growth.
LONG-TERM MONITORING OF ABIOTIC CONDITIONS IN THE MULLICA RIVER-GREAT BAY ESTUARY Kennish, M. J., Institute of Marine and Coastal Sciences, Rutgers University, New Brunswick, New Jersey 08901 and O'Donnell, S., Rutgers Marine Field Station, 132 Great Bay Boulevard, Tuckerton, New Jersey 08087 The Mullica River-Great Bay Estuary is the target of a long-term monitoring program to assess variation in physical-chemical conditions along a well-defined salinity gradient. Since 1996, measurements of an array of physical-chemical variables (i.e., temperature, salinity, dissolved oxygen, pH, turbidity, and water depth) have been made at Lower Bank and Chestnut Neck in the Mullica River and at Buoy 126 in Great Bay using unattended, automated YSI 6-series dataloggers. These instruments have recorded data semi-continuously (every 30 minutes) on the aforementioned variables, yielding an extensive database for the past five years. Analyses of these data indicate that differences in environmental conditions are generally statistically significant in oligohaline, mesohaline, and polyhaline waters of the estuarine system. This type of study is important not only because it can be used to delineate the factors responsible for biotic community changes in the estuary but also because it can provide essential data to decision-makers who address coastal resource management problems. A BATHYMETRIC AND LIMNOLOGICAL SURVEY OF ISLAND POND, ORANGE CO, NY), A DEEP NATURAL LAKE WITH A METALIMNETIC OXYGEN MINIMUM. Edward S. Kubersky, Department of Natural Sciences, Felician College, Lodi, NJ 07644 Island Pond lies within Harriman State Park, along the Appalachian Trail. Vehicle access is restricted and the watershed (4 km2) is undeveloped, consisting of forest and swamp. During a survey of the Pond in August 2001, a complete bathymetric map was produced. Although modest in surface area (22.3 ha), Island Pond is spectacularly deep (max depth 35.7m, mean depth 12.5m). It is the third deepest natural lake in southern New York State. The Pond is consequently highly voluminous (28 x 105 m3) and has a longer Water Residence Time (1.28 years) than shallow lakes with similar area and watershed.The water is pale amber due to humic substances in the watershed, and not very transparent (5% light level 2m, Secchi depth 1.5m). The metalimnion extends from 2m-6m and an unusual metalimnetic oxygen minimum exists between 3.5-4.0m. Oxygen is absent below 25m. The region of this minimum might provide a potential environment for photosynthetic purple nonsulfur bacteria. Island Pond water is "soft" (total hardness 17mg/L CaCO3), slightly acidic (pH 6.4), and has low alkalinity (10 mg/L CaCO3).Based on Total Phosphorus (TP) input, a nutrient budget was calculated using the Vollenweider-Dillon equations. Phosphorus enters from anoxic bottom sediments (57%), from the watershed (40%), and from atmospheric precipitation (3%). The model's prediction of eutrophy agreed with field observations of the Pond. SEASONAL CHANGES IN SPECIES COMPOSITION AND ABUNDANCE OF ZOOPLANKTON IN NORTHERN BARNEGAT BAY. Audrey J. McGough (student), Louise Wootton, Department of Biology, Georgian Court College, Lakewood, NJ 08701 Zooplankton are an important link between the photosynthetic phytoplankton and upper trophic levels, including many commercially important species. Little is known of the species composition of zooplankton and patterns of seasonal abundance in the Barnegat Bay. Water was collected on a weekly basis from off a bulkhead in the northern Barnegat Bay, and filtered to obtain zooplankton samples which were subsequently analyzed for species composition and abundance. Additional data was collected upon salinity, dissolved oxygen content, percent oxygen saturation and water temperature. Copepods dominated spring and early summer samples, with taxa being more evenly represented throughout the rest of the year. Dominant genera of copepods in our samples were Acartia, Calanus and Centropages. Harpacticoid copepods were also common in our samples. Copepod abundance peaked between March and April, but abundances rarely exceeded 30 individuals L-1. Other crustaceans as well as annelids and larval fish were most numerous during the months of May and June. Salinity, temperature and oxygen concentrations of the water appeared to have no impact on abundance of copepods. Sensitivity Analysis of Sustainability Indicators by Area-Based method. Loren S. Muldowney (student), Department of Environmental Sciences, Rutgers University, 14 College Farm Road, New Brunswick, NJ 08901 Sustainability is a stated goal of organizations, industries and all levels of government worldwide, but their commitment to sustainability exceeds their ability to evaluate either current status or progress toward the goal. The evaluation of sustainability is sensitive to assumptions and methods of analysis even when using identical raw data. Internal contradictions in existing evaluation models prevent widespread acceptance of the methods. Properly used area-based methods can eliminate this sensitivity and lead to an internally consistent analytical system. Evaluation of biophysical sustainability requires the comparison of a load to a load-bearing capacity. The materials and energy required for the production of goods and services can be expressed as the productive capacity of some finite amount of land. For any defined population, all goods and services used by the population are converted to equivalent land area in various use classes: energy, agriculture, forest, barren, or aquatic. Summation of all such equivalent land areas yields a measure of the society's total environmental load. A comparison of the load with the load-bearing capacity is a strong indicator of sustainability, provided each can be expressed in a common unit. The method proposed uses a common unit and remedies some of the methodological problems associated with the area-based indicator models currently in use.
WATER QUALITY IN TWO URBAN STREAMS-MOLLY ANN AND GOFFLE BROOKS, NORTHERN NEW JERSEY. Richard R. Pardi, and Amy McWatters (student), Environmental Science Program, William Paterson University, Wayne, New Jersey 07470 Monitoring of water quality variables on two urban streams within the lower Passaic River watershed – Goffle and Molly Ann Brooks – reveals substantial variations in water quantity as well as the concentration of dissolved constituents. Most dissolved ions in stream water behave quasi-conservatively with the exception of nitrate and dissolved silica whose concentrations appear to be controlled by a combination of kinetics, biological and abiolgical processes. Variations in stream discharge are likely exacerbated by ground water withdrawals for potable water supply. Rock and soil leaching laboratory experiments with local materials reveal that surface rocks and soils are not significant sources of these stream’s high alkalinities, suggesting a ground water source. High dissolved inorganic nitrate levels in ground water compare with the highest nitrate levels in streamwater and raise some concern for efforts to protect recharge zones within these sub-watersheds. INTERACTION OF PREGENOLONE SULFATE AND THYROID HORMONE WITH THE GABAA RECEPTOR. Tavia S. Cleveland (student), Stephanie R. Addison (student), and Joseph V. Martin, Biology Department, Rutgers University, Camden, NJ. In contrast to the genomic actions of thyroid hormones (THs) evident in developing brain, THs may have some nongenomic effects in adult mammalian neurons. For example, our previous results showed that THs inhibited muscimol-stimulated uptake of 36Cl- in adult rat brain cortical synaptoneurosomes and had a biphasic action on the binding of a convulsant ligand for the GABAA receptor in highly washed cerebrocortical membranes. Pregenenolone sulfate (PS), a neurosteroid, also interacts with the GABAA receptor. We tested the effects of PS and triiodothyronine (L-T3), individually and in combination, on binding of tritiated t-butyl-bicyclo-orthocarboxylate(3H-TBOB), a ligand for the convulsant site of the receptor. The cerebral cortices of mature Sprague-Dawley rats were dissected and thoroughly washed in 50 mM Tris-HCl buffer (pH 7.4). The resulting membrane preparation was incubated at 25蚓 for 60 minutes with 3H-TBOB in combination with PS (1, 10, 50 _M) and/or L-triiodothyronine (0.1-50 _M). Nonspecific binding was determined in parallel incubates including additional unlabeled picrotoxin. The membrane preparation was rapidly filtered, the filters washed and the radioactivity on the filters determined. Our results suggest that both TH and PS influence GABAA receptor binding, but by separate mechanisms of interaction with the receptor. (Supported by NSF grant IBN-981089) THE AFRICAN BUTTERFLY FISH (Pantodon buchholzi Petersi) JUMPS- IT DOES NOT GLIDE. Shannon Fornari (student), Gabriel Strain and William M. Saidel, Biology Department, Rutgers University, Camden, NJ 08102 The African butterfly fish, Pantodon buchholzi Peters, is noted for its unique ecological and phylogenetic position. The species is the lone entry in the monospecific family Pantodontidae. It lives in the waters of the Cross River system of West Africa, and as its ecological niche is just below the water surface, it has evolved an escape behavior that is a jump into the air. This ability has alternately been described as either a jump or a glide and as a result, the fish has been called a fresh water flying fish. We have been studying this behavior using video analysis at both 33 msec intervals and at 2 ms intervals. The evidence we will present demonstrates that this behavior is a ballistic motion that may be distorted by body movements while in flight. The initial acceleration is a consequence of pectoral fin abduction with a modicum of help from tail maneuvers. A fish may jump to a height exceeding its body length and may execute flips in air due to the initial inertial moment imparted at the jump. Although suggested by Pellegrin (1923), our video study conclusively shows that its pectoral fins do not flap. Its motion is ballistic, unlike the flying fish of the genera Dactylopterus or Exocoetus. GASOLINE OXYGENATES AND RELATED METABOLITES EXERT BIPHASIC INFLUENCES ON GABAA RECEPTOR FUNCTIONS. Sonal Iyer (student), M. Michael Iba, Veronica P. O'Connor (student), and Joseph V. Martin, Biology Department, Rutgers University, Camden, NJ and Department of Pharmacology and Toxicology, Environmental and Occupational Health Sciences Institute, Rutgers University, Piscataway, NJ Oxygenates such as methyl-t-butyl ether (MTBE), ethyl-t-butyl ether (ETBE), and t-amyl-methyl ether (TAME) improve combustion efficiency of gasoline . The major metabolic product of ETBE and MTBE is t-butyl alcohol (TBA). TAME metabolism produces t-amyl alcohol (TAA). Previously, our lab showed that these additives alter binding of GABAA receptor ligands. The current study examines receptor function using rat-brain synaptoneurosomes (SNS), a preparation of sacs of neuronal membrane. Various concentrations of each compound were added to SNS along with 36Cl- and muscimol (GABAA agonist). Influx of 36Cl- into the SNS was terminated after 5 seconds using picrotoxin (a GABAA channel blocker), and the radioactivity inside the SNS was measured. Each tested compound enhanced muscimol-stimulated uptake of 36Cl- at lower concentrations, while higher concentrations were less effective. The EC50 for the ethers tested were determined as 0.19 mM (ethyl ether), 0.07 mM (MTBE), 0.53 mM (ETBE), and 0.13 mM (TAME). The EC50 for the alcohols studied were determined as 4.19 mM (ethanol), 3.66 mM (TBA), and 0.64 mM (TAA). MTBE, the most commonly used additive, was most potent. The results suggest a potential role of the GABAA receptor in some of the neurotoxic effects of oxygenates. Metabolites of the additives may also contribute to the neurological symptoms. (Supported by NSF grant IBN-981089) FEEDING BEHAVIOR OF THE AFRICAN BUTTERFLY FISH (Pantodon buchholzi Petersi): A NEUROBIOLOGICAL BASIS. William M. Saidel, Biology Department, Rutgers University, Camden, NJ 08102 The African butterfly fish, Pantodon buchholzi Peters, is an obligatory surface feeder. It feeds following a visual stimulus in ‘snell’s window’. The target is presented to one eye and the information is conveyed to two regions of the brain, the optic tectum and the nucleus rostrolateralis in the diencephalon. In tracing the path of information from the retina through the brain to the motor centers, two paths have been discovered. The first is from the retina to the optic tectum and to the presumptive motor centers. This pathway is thought to convey the target presence and the spatial information as to where the target is located. The second pathway, using information from both the retina and correlated tectal information converges on nucleus rostrolateralis where it is conveyed to the midbrain nucleus interpeduncularis (IP). IP is a relatively unknown area of the brain of fishes and other vertebrates, but it is located in association with the Raphe Nuclei. Such an area is involved with motivation and activation processes. A hypothesis will be presented that suggests the pathway from retina and optic tectum to nu. rostrolateralis to IP is a switch that maintains the obligatory aspect of feeding. Its suppression accounts for the lack of feeding behavior following multiple trials in this fish and other vertebrates. ALTERATION OF 3H-MUSCIMOL BINDING AT GABAA RECEPTOR BY L-TRIIODOTHYRONINE AND ITS ANALOGS IN MEMBRANES FROM ADULT RAT BRAIN. Pradip K. Sarkar, Jenny Yuen (student), Chandria L. Mitchell (student) and Joseph V. Martin, Biology Department, Rutgers University, NJ 08102 Recent evidence suggests that thyroid hormones (THs) may have some nongenomic influences in mature mammalian brain in contrast to the classical genomic actions evident in developing brain. We have previously demonstrated that THs influence binding and activity of GABAA receptors. The present study investigates the effect of L_triiodothyronine (L-T3) and TH analogs on 3H-muscimol binding in highly washed brain membranes prepared from cerebral cortex of young adult male Sprague Dawley rats. A 1 然 dose of L-T3 significantly inhibited 3H-muscimol binding. Although L-thyroxine (L-T4) also significantly inhibited of 3H-muscimol binding, no significant changes were observed between low (100 nM) and high doses (1 然) of the TH. Surprisingly, the D-isomers of both T3 and T4 (at 100 nM or 1 然) significantly increased 3H-muscimol binding. The higher dose (1 然) of D-T3 stimulated 3H-muscimol binding less effectively as compared to lower dose (100 nM). No significant differences were noted between the stimulatory effects of 100 nM and 1 然 D-T4 on 3H-muscimol binding. Other analogs of TH tested did not significantly influence 3H-muscimol binding. The high sensitivity of 3H-muscimol binding to TH supports a functional significance of the GABAA receptor of the plasma membrane as a site for nongenomic actions of the hormones in adult brain. (Supported by NSF grant IBN-981089) Poster Presentation
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