Darting and marking techniques for an arboreal forest monkey,Cercopithecus ascanius

Techniques are described for capturing and marking redtail monkeys (Cercopithecus ascanius). An air-powered darting rifle and syringe darts loaded with a Ketamine-Rompun mixture were used for capture. Ketamine was used for maintaining anesthesia. Monkeys were darted 48 times and captured 27 times. In 24 of the 27 captures, the monkeys were released unharmed. Adult males were marked with radiotransmitters attached to collars of nylon webbing. Females received nylon webbing collars with colorcoded plastic washers for identification.     

Monoclonal antibody to a protein of the nucleus and mitotic spindle of mammalian cells

We report here the isolation of a monoclonal antibody, J17, that reacts with a conserved vertebrate protein antigen that is present in the spindle apparatus during mitosis but found within the nucleus during interphase. Immunofluorescence microscopy demonstrates that the J17 antigen is found in numerous punctate regions that are distinct from nucleoli. Furthermore, this antigen is not directly associated with kinetochores, the nuclear envelope, or with metaphase chromosomes. — Antibody J17 immunoprecipitates a single polypeptide of very high molecular weight (over 250000) from K562 human erythroleukemia cells pulse-labeled with ¹⁴C-leucine. This polypeptide is converted quantitatively to a stable 220-kilodalton product within one cellular generation. We discuss the possible relevance of this processing event for transport into the nucleus. The J17 antigen is synthesized throughout the cell cycle in Chinese hamster ovary cells.     

Viability and Isolation of Marine Bacteria by Dilution Culture: Theory, Procedures, and Initial Results

Dilution culture, a method for growing the typical small bacteria from natural aquatic assemblages, has been developed. Each of 11 experimental trials of the technique was successful. Populations are measured, diluted to a small and known number of cells, inoculated into unamended sterilized seawater, and examined three times for the presence of 10⁴ or more cells per ml over a 9-week interval. Mean viability for assemblage members is obtained from the frequency of growth, and many of the cultures produced are pure. Statistical formulations for determining viability and the frequency of pure culture production are derived. Formulations for associated errors are derived as well. Computer simulations of experiments agreed with computed values within the expected error, which verified the formulations. These led to strategies for optimizing viability determinations and pure culture production. Viabilities were usually between 2 and 60% and decreased with >5 mg of amino acids per liter as carbon. In view of difficulties in growing marine oligobacteria, these high values are noteworthy. Significant differences in population characteristics during growth, observed by high-resolution flow cytometry, suggested substantial population diversity. Growth of total populations as well as of cytometry-resolved subpopulations sometimes were truncated at levels of near 10⁴ cells per ml, showing that viable cells could escape detection. Viability is therefore defined as the ability to grow to that population; true viabilities could be even higher. Doubling times, based on whole populations as well as individual subpopulations, were in the 1-day to 1-week range. Data were examined for changes in viability with dilution suggesting cell-cell interactions, but none could be confirmed. The frequency of pure culture production can be adjusted by inoculum size if the viability is known. These apparently pure cultures produced retained the size and apparent DNA-content characteristic of the bulk of the organisms in the parent seawater. Three cultures are now available, two of which have been carried for 3 years. The method is thus seen as a useful step for improving our understanding of typical aquatic organisms.     

Identification of a locus involved in meningococcal lipopolysaccharide biosynthesis by deletion mutagenesis

A novel method for insertion/deletion mutagenesis in meningococci was devised. This consisted of ligating a digest of total chromosomal DNA to a 1.1 kb restriction fragment containing an erythromycin-resistance marker ( ermC ), and subsequent transformation of the ligation mixture into the homologous meningococcal strain H44/76. Southern blotting of a number of the resulting erythromycin-resistant transformants demonstrated that all carried the ermC gene inserted at different positions in the chromosome. Mutants with a specific phenotype were identified by screening with the anti-lipopolysaccharide (LPS) monoclonal antibody MN4A8B2, which is specific for immunotype L3. In this way, two independent L3-negative mutant strains were isolated. In transformation experiments with chromosomal DNA from these mutants, erythromycin-resistance and lack of MN4A8B2 reactivity were always linked, showing that the insertion/deletion was in a locus involved in LPS biosynthesis. On SDS–PAGE, the mutant LPS displayed an electrophoretic mobility intermediate between that produced by the previously isolated galE and rfaF mutant strains. Chemical analysis of the mutant LPS revealed that the structure was probably lipid A–(KDO)₂–(Hep)₂. Chromosomal DNA flanking the ermC insertion in these two mutant strains was cloned, and used as probe for the isolation of the corresponding region of the wild-type strain. From hybridization and polymerase chain reaction (PCR) analysis, it could be concluded that both mutations map to the same locus. The affected gene probably encodes the glycosyltransferase necessary for adding N -acetylglucosamine to heptose.     

Dissolved Hydrocarbons and Related Microflora in a Fjordal Seaport: Sources, Sinks, Concentrations, and Kinetics

The continuous addition of toluene as a solute of treated ballast water from oil tankers into a well-defined estuary facilitated the study of the dynamics of dissolved hydrocarbon metabolism in seawater. Most rates of toluene oxidation were in the range of 1 to 30 pg/liter per h at 0.5 μg of toluene per liter. Near the ballast water injection point, a layer of warm ballast water, rich in bacteria, that was trapped below the less-dense fresh surface water was located. Toluene residence times were approximately 2 weeks in this layer, 2 years elsewhere in Port Valdez, and 2 decades in the surface water of a more oceanic receiving estuary adjacent. Mixing was adequate for a steady-state treatment which showed that 98% of the toluene was flushed from Port Valdez before metabolism and gave a steady-state concentration of 0.18 μg/liter. Total bacterial biomass from direct counts and organism size data was usually near 0.1 mg/liter, but ranged up to 0.8 mg/liter in the bacteria-rich layer. The origin of bacteria in this layer was traced to growth in oil tanker ballast during shipments. The biomass of toluene oxidizers in water samples was estimated from the average affinity of pure-culture isolates for toluene (28 liters per g of cells per h) and observed toluene oxidation kinetics. Values ranged from nearly all of the total bacterial biomass within the bacteria-rich layer down to 0.2% at points far removed. Because the population of toluene oxidizers was large with respect to the amount of toluene consumed and because water from a nearby nonpolluted estuary was equally active in facilitating toluene metabolism, we searched for an additional hydrocarbon source. It was found that terpenes could be washed from spruce trees by simulated rainfall, which suggested that riparian conifers provide an additional and significant hydrocarbon source to seawater.     

The dimer-monomer equilibrium constant for [125I]β nerve growth factor

The equilibrium constant for $\text{[}{}^{125}\text{I]β}$ nerve growth factor was determined using polyacrylamide gel electrophoresis to separate the monomer and dimer. Various concentrations of the radiolabelled nerve growth factor were incubated for 24 and 48 hours. The equilibrium constants obtained for both incubation periods were the same, 3.2 ± 1.4 × 10^?11$\text{M}$ and 2.6 ± 1.6 × 10^?11$\text{M}$, respectively. Thus, at physiological concentrations the β nerve growth factor is in the dimeric form almost exculsively.     

Reducing Biotic and Abiotic Land‐Use Legacies to Restore Invaded,Abandoned Citrus Groves

Land‐use legacies associated with agriculture, such as increased soil fertility and elevated soil pH, promote invasions by non‐native plant species on former agricultural lands. Restoring natural soil conditions (i.e. low fertility and low pH) may be an effective, long‐term method to control and reduce the abundance of non‐native and ruderal species that invade abandoned agricultural lands. In this study, we examined how soil manipulation treatments of lowering soil fertility with carbon additions and lowering soil pH by applying sulfur affect non‐native and ruderal native plant species abundance in two former citrus groves in central Florida. Non‐native plant biomass was removed by one of two methods (tilling or topsoil removal), and was combined with a soil amendment of sulfur, carbon, sulfur + carbon, or none. The biomass removal treatments significantly decreased non‐native abundance, with topsoil removal as the most effective. Carbon additions did not affect soil fertility or vegetation. Sulfur and sulfur + carbon additions significantly decreased soil pH in both groves for at least 1 year post‐treatment; however, we did not see a significant vegetation response. Overall, our results suggest that removing vegetation by tilling and topsoil removal is an effective method for reducing non‐target species cover. Although we did not see a response of vegetation to our treatments, we were able to restore the initial soil characteristics, which can be a first step toward complete restoration.     

Simultaneous knockout of Slo3 and CatSper1 abolishes all alkalization- and voltage-activated current in mouse spermatozoa

During passage through the female reproductive tract, mammalian sperm undergo a maturation process termed capacitation that renders sperm competent to produce fertilization. Capacitation involves a sequence of changes in biochemical and electrical properties, the onset of a hyperactivated swimming behavior, and development of the ability to undergo successful fusion and penetration with an egg. In mouse sperm, the development of hyperactivated motility is dependent on cytosolic alkalization that then results in an increase in cytosolic Ca²⁺. The elevation of Ca²⁺ is thought to be primarily driven by the concerted interplay of two alkalization-activated currents, a K⁺ current (KSPER) composed of pore-forming subunits encoded by the Kcnu1 gene (also termed Slo3) and a Ca²⁺ current arising from a family of CATSPER subunits. After deletion of any of four CATSPER subunit genes (CATSPER1–4), the major remaining current in mouse sperm is alkalization-activated KSPER current. After genetic deletion of the Slo3 gene, KSPER current is abolished, but there remains a small voltage-activated K⁺ current hypothesized to reflect monovalent flux through CATSPER. Here, we address two questions. First, does the residual outward K⁺ current present in the Slo3 ^−/− sperm arise from CATSPER? Second, can any additional membrane K⁺ currents be detected in mouse sperm by patch-clamp methods other than CATSPER and KSPER? Here, using mice bred to lack both SLO3 and CATSPER1 subunits, we show conclusively that the voltage-activated outward current present in Slo3 ^−/− sperm is abolished when CATSPER is also deleted. Any leak currents that may play a role in setting the resting membrane potential in noncapacitated sperm are likely smaller than the pipette leak current and thus cannot be resolved within the limitation of the patch-clamp technique. Together, KSPER and CATSPER appear to be the sole ion channels present in mouse sperm that regulate membrane potential and Ca²⁺ influx in response to alkalization.