While renewable biofuels can reduce negative effects of fossil fuel energy consumption, the magni... more While renewable biofuels can reduce negative effects of fossil fuel energy consumption, the magnitude of their benefits depends on the magnitude of N 2 O emissions. High variability of N 2 O emissions overpowers efforts to curb uncertainties in estimating N 2 O fluxes from biofuel systems. In this study, we explored (a) N 2 O production via bacterial denitrification and (b) N 2 O emissions from soils under several contrasting bioenergy cropping systems, with specific focus on explaining N 2 O variations by accounting for soil pore characteristics. Intact soil samples were collected after 9 years of implementing five biofuel systems: continuous corn with and without winter cover crop, monoculture switchgrass, poplars, and early-successional vegetation. After incubation, N 2 O emissions were measured and bacterial denitrification was determined based on the site-preference method. Soil pore characteristics were quantified using X-ray computed microtomography. Three bioenergy systems with low plant diversity, that is, corn and switchgrass systems, had low porosities, low organic carbon contents, and large volumes of poorly aerated soil. In these systems, greater volumes of poorly aerated soil were associated with greater bacterial denitrification, which in turn was associated with greater N 2 O emissions (R 2 = 0.52, p < 0.05). However, the two systems with high plant diversity, that is, poplars and early-successional vegetation, over the 9 years of implementation had developed higher porosities and organic carbon contents. In these systems, volumes of poorly aerated soil were positively associated with N 2 O emissions without a concomitant increase in bacterial denitrification. Our results suggest that changes in soil pore architecture generated by long-term implementation of contrasting bioenergy systems may affect the pathways of N 2 O production, thus, change associations between N 2 O emissions and other soil properties. Plant diversity appears as one of the factors determining which microscale soil characteristics will influence the amounts of N 2 O emitted into the atmosphere and, thus, which can be used as effective empirical predictors.
In this paper, 1 Tables 2 and 3 should have appeared as below: TABLE 2 Interlaboratory comparison... more In this paper, 1 Tables 2 and 3 should have appeared as below: TABLE 2 Interlaboratory comparison of USGS51 and USGS52 N 2 O isotopic measurements. Individual laboratory uncertainties are standard deviations of minimum of three repeated measurements against the laboratory reference gas and do not include any calibration uncertainties of the laboratory reference gas with respect to Air-N 2 and VSMOW Laboratory code Name δ 15 N Air-N2 (mUr) δ 18 O VSMOW (mUr) δ 15 N α Air-N2 (mUr) δ 15 N β Air-N2 (mUr) S P (mUr) Methodology
Within pure cultures of microbes our prior research has demonstrated that the isotopomer composit... more Within pure cultures of microbes our prior research has demonstrated that the isotopomer composition of nitrous oxide can provide a basis to apportion the origin of this important greenhouse gas to nitrification and denitrification. This approach is based on the observation that nitrous oxide derived from denitrification and nitrification have distinct values (approx. 0 and 30 0/00, respectively) in Site
We used stable isotope analysis to investigate the foraging ecology of coastal bottlenose dolphin... more We used stable isotope analysis to investigate the foraging ecology of coastal bottlenose dolphins (Tursiops truncatus) in relation to a series of anthropogenic disturbances. We first demonstrated that stable isotopes are a faithful indicator of habitat use by comparing muscle isotope values to behavioral foraging data from the same individuals. d 13 C values increased, while d 34 S and d 15 N values decreased with the percentage of feeding observations in seagrass habitat. We then utilized stable isotope values of muscle to assess temporal variation in foraging habitat from 1991 to 2010 and collagen from tooth crown tips to assess the time period 1944 to 2007. From 1991 to 2010, d 13 C values of muscle decreased while d 34 S values increased indicating reduced utilization of seagrass habitat. From 1944 to 1989 d 13 C values of the crown tip declined significantly, likely due to a reduction in the coverage of seagrass habitat and d 15 N values significantly increased, a trend we attribute to nutrient loading from a rapidly increasing human population. Our results demonstrate the utility of using marine mammal foraging habits to retrospectively assess the extent to which anthropogenic disturbance impacts coastal food webs.
Hydrogenases catalyze the reversible formation of H2 from electrons and protons with high efficie... more Hydrogenases catalyze the reversible formation of H2 from electrons and protons with high efficiency. Understanding the relationships between H2 production, H2 uptake, and H2-H2O exchange can provide insight into the metabolism of microbial communities in which H2 is an essential component in energy cycling. We used stable H isotopes (1H and 2H) to probe the isotope effects associated with three [FeFe]-hydrogenases and three [NiFe]-hydrogenases. All six hydrogenases displayed fractionation factors for H2 formation that were significantly less than 1, producing H2 that was severely depleted in 2H relative to the substrate, water. Consistent with differences in their active site structure, the fractionation factors for each class appear to cluster, with the three [NiFe]-hydrogenases (α = 0.27–0.40) generally having smaller values than the three [FeFe]-hydrogenases (α = 0.41–0.55). We also obtained isotopic fractionation factors associated with H2 uptake and H2-H2O exchange under conditions similar to those utilized for H2 production, providing a more complete picture of the reactions catalyzed by hydrogenases. The fractionation factors determined in our studies can be used as signatures for different hydrogenases to probe their activity under different growth conditions and to ascertain which hydrogenases are most responsible for H2 production and/or uptake in complex microbial communities.
Two commercial Bifidobacterium bifidum (Bf-1 and Bf-6) were cultured in 12% (wt/vol) reconstitute... more Two commercial Bifidobacterium bifidum (Bf-1 and Bf-6) were cultured in 12% (wt/vol) reconstituted nonfat dry milk (NDM) containing 5% (wt/wt) honey, sucrose, fructose, or glucose. Inoculated samples were incubated anaerobically at 37 degrees C for 48 h. Samples were collected at 12-h intervals and examined for (i) specific growth rate, (ii) pH, and (iii) levels of fermentation end products (lactic and acetic acids) as measured by high-performance liquid chromatography (HPLC). Viability of the organisms during 28 days of refrigerated storage at 4 degrees C was also assessed at 7-day intervals. Growth promotion and acid production were greatest when Bf-1 and Bf-6 were grown in the presence of honey. For both Bf-1 and Bf-6, retention of viability was greatest up to 14 days of refrigerated storageat 4 degrees C when they were grown and stored in the presence of honey compared to other sweeteners.
Five human intestinal Bifidobacterium spp., B. longum, B. adolescentis, B. breve, B. bifidum, and... more Five human intestinal Bifidobacterium spp., B. longum, B. adolescentis, B. breve, B. bifidum, and B. infantis, were cultured in reinforced clostridial medium (control) and in reinforced clostridial medium supplemented with 5% (wt/vol) honey, fructooligosaccharide (FOS), galactooligosaccharide (GOS), and inulin. Inoculated samples were incubated anaerobically at 37degrees C for 48 h. Samples were collected at 12-h intervals and examined for specific growth rate. Levels of fermentation end products (lactic and acetic acids) were measured by high-pressure liquid chromatography. Honey enhanced the growth of the five cultures much like FOS, GOS, and inulin did. Honey, FOS, GOS, and inulin were especially effective (P < 0.05) in sustaining the growth of these cultures after 24 h of incubation as compared with the control treatment. Overall, the effects of honey on lactic and acetic acid production by intestinal Bifidobacterium spp. were similar to those of FOS, GOS, and inulin.
This study examines resource use (diet, habitat use, and trophic level) within and among demograp... more This study examines resource use (diet, habitat use, and trophic level) within and among demographic groups (males, females, and juveniles) of bottlenose dolphins (Tursiops truncatus). We analyzed the d 13 C and d 15 N values of 15 prey species constituting 84% of the species found in stomach contents. We used these data to establish a trophic enrichment factor (TEF) to inform dietary analysis using a Bayesian isotope mixing model. We document a TEF of 0& and 2.0& for d 13 C and d 15 N, respectively. The dietary results showed that all demographic groups relied heavily on low trophic level seagrass-associated prey. Bayesian standard ellipse areas (SEA b) were calculated to assess diversity in resource use. The SEA b of females was nearly four times larger than that of males indicating varied resource use, likely a consequence of small home ranges and habitat specialization. Juveniles possessed an intermediate SEA b , generally feeding at a lower trophic level compared to females, potentially an effect of natal philopatry and immature foraging skills. The small SEA b of males reflects a high degree of specialization on seagrass associated prey. Patterns in resource use by the demographic groups are likely linked to differences in the relative importance of social and ecological factors.
ABSTRACT This study examines resource use (diet, habitat use, and trophic level) within and among... more ABSTRACT This study examines resource use (diet, habitat use, and trophic level) within and among demographic groups (males, females, and juveniles) of bottlenose dolphins (Tursiops truncatus). We analyzed the δ13C and δ15N values of 15 prey species constituting 84% of the species found in stomach contents. We used these data to establish a trophic enrichment factor (TEF) to inform dietary analysis using a Bayesian isotope mixing model. We document a TEF of 0‰ and 2.0‰ for δ13C and δ15N, respectively. The dietary results showed that all demographic groups relied heavily on low trophic level seagrass-associated prey. Bayesian standard ellipse areas (SEAb) were calculated to assess diversity in resource use. The SEAb of females was nearly four times larger than that of males indicating varied resource use, likely a consequence of small home ranges and habitat specialization. Juveniles possessed an intermediate SEAb, generally feeding at a lower trophic level compared to females, potentially an effect of natal philopatry and immature foraging skills. The small SEAb of males reflects a high degree of specialization on seagrass associated prey. Patterns in resource use by the demographic groups are likely linked to differences in the relative importance of social and ecological factors.
The relative importance of individual microbial pathways in nitrous oxide (N(2)O) production is n... more The relative importance of individual microbial pathways in nitrous oxide (N(2)O) production is not well known. The intramolecular distribution of (15)N in N(2)O provides a basis for distinguishing biological pathways. Concentrated cell suspensions of Methylococcus capsulatus Bath and Nitrosomonas europaea were used to investigate the site preference of N(2)O by microbial processes during nitrification. The average site preference of N(2)O formed during hydroxylamine oxidation by M. capsulatus Bath (5.5 +/- 3.5 per thousand) and N. europaea (-2.3 +/- 1.9 per thousand) and nitrite reduction by N. europaea (-8.3 +/- 3.6 per thousand) differed significantly (ANOVA, f((2,35)) = 247.9, p = 0). These results demonstrate that the mechanisms for hydroxylamine oxidation are distinct in M. capsulatus Bath and N. europaea. The average delta(18)O-N(2)O values of N(2)O formed during hydroxylamine oxidation for M. capsulatus Bath (53.1 +/- 2.9 per thousand) and N. europaea (-23.4 +/- 7.2 per thousand) and nitrite reduction by N. europaea (4.6 +/- 1.4 per thousand) were significantly different (ANOVA, f((2,35)) = 279.98, p = 0). Although the nitrogen isotope value of the substrate, hydroxylamine, was similar in both cultures, the observed fractionation (delta(15)N) associated with N(2)O production via hydroxylamine oxidation by M. capsulatus Bath and N. europaea (-2.3 and 26.0 per thousand, respectively) provided evidence that differences in isotopic fractionation were associated with these two organisms. The site preferences in this study are the first measured values for isolated microbial processes. The differences in site preference are significant and indicate that isotopomers provide a basis for apportioning biological processes producing N(2)O.
A technique for measurement of the stable isotope composition of dissolved organic carbon (DOC) i... more A technique for measurement of the stable isotope composition of dissolved organic carbon (DOC) in stream water, using an elemental analyzer (EA) coupled to an isotope ratio mass spectrometer (IRMS), is described. Stream water samples were concentrated by rotary evaporation, acidified to remove dissolved inorganic carbon (DIC), and dried in silver cups prior to analysis. Precision was evaluated with standards (alanine and humic acid), and with stream water samples with varying 13 C enrichment. Standards and samples were also prepared in sealed quartz tubes for high-temperature combustion (HTC) and analyzed by dual inlet for comparison. The d 13 C values of natural abundance standards and samples measured by the two techniques differed by 0.3%. Isotopically enriched stream water samples analyzed using the EA and HTC methods had comparable d 13 C and atom % values (differed by 6%). Expensive quartz vessels and the extensive time required for sample lyophilization, combustion and cryogenic separation of combustion products before isotopic analysis for the HTC method are eliminated with the EA method. Overall, the EA method is simpler, faster, and more economical than the HTC method, and it can be used for 13 C-enriched as well as natural abundance samples.
Hydrogenases catalyze the reversible formation of H(2), and they are key enzymes in the biologica... more Hydrogenases catalyze the reversible formation of H(2), and they are key enzymes in the biological cycling of H(2). H isotopes have the potential to be a very useful tool in quantifying hydrogen ion trafficking in biological H(2) production processes, but there are several obstacles that have thus far limited the application of this tool. Here, we describe a new method that overcomes some of these barriers and is specifically designed to measure isotopic fractionation during enzyme-catalyzed H(2) evolution. A key feature of this technique is that purified hydrogenases are employed, allowing precise control over the reaction conditions and therefore a high level of precision. In addition, a custom-designed high-throughput gas chromatograph/isotope ratio mass spectrometer is employed to measure the isotope ratio of the H(2). Using our new approach, we determined that the fractionation factor for H(2) production by the [NiFe]-hydrogenase from Desulfovibrio fructosovorans is 0.273 ± 0.006. This result indicates that, as expected, protons are highly favored over deuterium ions during H(2) evolution. Potential applications of this newly developed method are discussed.
ABSTRACT We used stable isotope analysis to investigate the foraging ecology of coastal bottlenos... more ABSTRACT We used stable isotope analysis to investigate the foraging ecology of coastal bottlenose dolphins (Tursiops truncatus) in relation to a series of anthropogenic disturbances. We first demonstrated that stable isotopes are a faithful indicator of habitat use by comparing muscle isotope values to behavioral foraging data from the same individuals. δ13C values increased, while δ34S and δ15N values decreased with the percentage of feeding observations in seagrass habitat. We then utilized stable isotope values of muscle to assess temporal variation in foraging habitat from 1991 to 2010 and collagen from tooth crown tips to assess the time period 1944 to 2007. From 1991 to 2010, δ13C values of muscle decreased while δ34S values increased indicating reduced utilization of seagrass habitat. From 1944 to 1989 δ13C values of the crown tip declined significantly, likely due to a reduction in the coverage of seagrass habitat and δ15N values significantly increased, a trend we attribute to nutrient loading from a rapidly increasing human population. Our results demonstrate the utility of using marine mammal foraging habits to retrospectively assess the extent to which anthropogenic disturbance impacts coastal food webs.
Two commercial strains of Bifidobacterium spp (Bf-1 and Bf-6) were cultured in 12% (w/w) reconsti... more Two commercial strains of Bifidobacterium spp (Bf-1 and Bf-6) were cultured in 12% (w/w) reconstituted nonfat dry milk (NDM) containing 0, 0.5, 1.0, 3.0, or 5.0% (w/v) fructooligosaccharide (FOS), galactooligosaccharide (GOS), and inulin. Inoculated samples were incubated anaerobically at 37°C for 48 h. Growth and activity of the cultures in the presence of FOS, GOS, and inulin were determined. Viability of each strain was assessed after 4 weeks of refrigerated storage at 4°C. Growth promotion, enhancement of activity and retention of viability were greatest when Bifidobacterium Bf-1 and Bf-6 were grown in the presence of FOS followed in a descending order by GOS and inulin. The effects of oligosaccharides and inulin increased with increasing carbohydrate concentration and was maximal at 5% (w/v).
Polyclonal antibodies (PAb) prepared against bovine milk alkaline phosphatase (ALP) were used to ... more Polyclonal antibodies (PAb) prepared against bovine milk alkaline phosphatase (ALP) were used to develop a competitive indirect (CI) ELISA. Anti-ALP PAb were specific for milk ALP and did not react with ALP from E. coli or bovine and calf intestinal mucosa. Anti-ALP PAb were 20% cross-reactive with bovine placenta ALP. The anti-ALP antibodies also did not recognize bovine serum albumin, acid glycoprotein, ovalbumin, ferritin, and casein, although some cross-reactivity was observed with whey protein isolate. Anti-ALP PAbs reacted with deglycosylated native ALP, but did not recognize ALP denatured at 100 degrees C in 2% SDS or deglycosylated denatured ALP. When buffered solutions of milk ALP were heated at 70 degrees C, ALP activity decreased at a faster rate than ALP content determined by CI-ELISA. The ELISA differentiated between native and heat denatured ALP. Further studies are warranted to determine if an ELISA can be used to verify pasteurization of fluid milk.
While renewable biofuels can reduce negative effects of fossil fuel energy consumption, the magni... more While renewable biofuels can reduce negative effects of fossil fuel energy consumption, the magnitude of their benefits depends on the magnitude of N 2 O emissions. High variability of N 2 O emissions overpowers efforts to curb uncertainties in estimating N 2 O fluxes from biofuel systems. In this study, we explored (a) N 2 O production via bacterial denitrification and (b) N 2 O emissions from soils under several contrasting bioenergy cropping systems, with specific focus on explaining N 2 O variations by accounting for soil pore characteristics. Intact soil samples were collected after 9 years of implementing five biofuel systems: continuous corn with and without winter cover crop, monoculture switchgrass, poplars, and early-successional vegetation. After incubation, N 2 O emissions were measured and bacterial denitrification was determined based on the site-preference method. Soil pore characteristics were quantified using X-ray computed microtomography. Three bioenergy systems with low plant diversity, that is, corn and switchgrass systems, had low porosities, low organic carbon contents, and large volumes of poorly aerated soil. In these systems, greater volumes of poorly aerated soil were associated with greater bacterial denitrification, which in turn was associated with greater N 2 O emissions (R 2 = 0.52, p < 0.05). However, the two systems with high plant diversity, that is, poplars and early-successional vegetation, over the 9 years of implementation had developed higher porosities and organic carbon contents. In these systems, volumes of poorly aerated soil were positively associated with N 2 O emissions without a concomitant increase in bacterial denitrification. Our results suggest that changes in soil pore architecture generated by long-term implementation of contrasting bioenergy systems may affect the pathways of N 2 O production, thus, change associations between N 2 O emissions and other soil properties. Plant diversity appears as one of the factors determining which microscale soil characteristics will influence the amounts of N 2 O emitted into the atmosphere and, thus, which can be used as effective empirical predictors.
In this paper, 1 Tables 2 and 3 should have appeared as below: TABLE 2 Interlaboratory comparison... more In this paper, 1 Tables 2 and 3 should have appeared as below: TABLE 2 Interlaboratory comparison of USGS51 and USGS52 N 2 O isotopic measurements. Individual laboratory uncertainties are standard deviations of minimum of three repeated measurements against the laboratory reference gas and do not include any calibration uncertainties of the laboratory reference gas with respect to Air-N 2 and VSMOW Laboratory code Name δ 15 N Air-N2 (mUr) δ 18 O VSMOW (mUr) δ 15 N α Air-N2 (mUr) δ 15 N β Air-N2 (mUr) S P (mUr) Methodology
Within pure cultures of microbes our prior research has demonstrated that the isotopomer composit... more Within pure cultures of microbes our prior research has demonstrated that the isotopomer composition of nitrous oxide can provide a basis to apportion the origin of this important greenhouse gas to nitrification and denitrification. This approach is based on the observation that nitrous oxide derived from denitrification and nitrification have distinct values (approx. 0 and 30 0/00, respectively) in Site
We used stable isotope analysis to investigate the foraging ecology of coastal bottlenose dolphin... more We used stable isotope analysis to investigate the foraging ecology of coastal bottlenose dolphins (Tursiops truncatus) in relation to a series of anthropogenic disturbances. We first demonstrated that stable isotopes are a faithful indicator of habitat use by comparing muscle isotope values to behavioral foraging data from the same individuals. d 13 C values increased, while d 34 S and d 15 N values decreased with the percentage of feeding observations in seagrass habitat. We then utilized stable isotope values of muscle to assess temporal variation in foraging habitat from 1991 to 2010 and collagen from tooth crown tips to assess the time period 1944 to 2007. From 1991 to 2010, d 13 C values of muscle decreased while d 34 S values increased indicating reduced utilization of seagrass habitat. From 1944 to 1989 d 13 C values of the crown tip declined significantly, likely due to a reduction in the coverage of seagrass habitat and d 15 N values significantly increased, a trend we attribute to nutrient loading from a rapidly increasing human population. Our results demonstrate the utility of using marine mammal foraging habits to retrospectively assess the extent to which anthropogenic disturbance impacts coastal food webs.
Hydrogenases catalyze the reversible formation of H2 from electrons and protons with high efficie... more Hydrogenases catalyze the reversible formation of H2 from electrons and protons with high efficiency. Understanding the relationships between H2 production, H2 uptake, and H2-H2O exchange can provide insight into the metabolism of microbial communities in which H2 is an essential component in energy cycling. We used stable H isotopes (1H and 2H) to probe the isotope effects associated with three [FeFe]-hydrogenases and three [NiFe]-hydrogenases. All six hydrogenases displayed fractionation factors for H2 formation that were significantly less than 1, producing H2 that was severely depleted in 2H relative to the substrate, water. Consistent with differences in their active site structure, the fractionation factors for each class appear to cluster, with the three [NiFe]-hydrogenases (α = 0.27–0.40) generally having smaller values than the three [FeFe]-hydrogenases (α = 0.41–0.55). We also obtained isotopic fractionation factors associated with H2 uptake and H2-H2O exchange under conditions similar to those utilized for H2 production, providing a more complete picture of the reactions catalyzed by hydrogenases. The fractionation factors determined in our studies can be used as signatures for different hydrogenases to probe their activity under different growth conditions and to ascertain which hydrogenases are most responsible for H2 production and/or uptake in complex microbial communities.
Two commercial Bifidobacterium bifidum (Bf-1 and Bf-6) were cultured in 12% (wt/vol) reconstitute... more Two commercial Bifidobacterium bifidum (Bf-1 and Bf-6) were cultured in 12% (wt/vol) reconstituted nonfat dry milk (NDM) containing 5% (wt/wt) honey, sucrose, fructose, or glucose. Inoculated samples were incubated anaerobically at 37 degrees C for 48 h. Samples were collected at 12-h intervals and examined for (i) specific growth rate, (ii) pH, and (iii) levels of fermentation end products (lactic and acetic acids) as measured by high-performance liquid chromatography (HPLC). Viability of the organisms during 28 days of refrigerated storage at 4 degrees C was also assessed at 7-day intervals. Growth promotion and acid production were greatest when Bf-1 and Bf-6 were grown in the presence of honey. For both Bf-1 and Bf-6, retention of viability was greatest up to 14 days of refrigerated storageat 4 degrees C when they were grown and stored in the presence of honey compared to other sweeteners.
Five human intestinal Bifidobacterium spp., B. longum, B. adolescentis, B. breve, B. bifidum, and... more Five human intestinal Bifidobacterium spp., B. longum, B. adolescentis, B. breve, B. bifidum, and B. infantis, were cultured in reinforced clostridial medium (control) and in reinforced clostridial medium supplemented with 5% (wt/vol) honey, fructooligosaccharide (FOS), galactooligosaccharide (GOS), and inulin. Inoculated samples were incubated anaerobically at 37degrees C for 48 h. Samples were collected at 12-h intervals and examined for specific growth rate. Levels of fermentation end products (lactic and acetic acids) were measured by high-pressure liquid chromatography. Honey enhanced the growth of the five cultures much like FOS, GOS, and inulin did. Honey, FOS, GOS, and inulin were especially effective (P < 0.05) in sustaining the growth of these cultures after 24 h of incubation as compared with the control treatment. Overall, the effects of honey on lactic and acetic acid production by intestinal Bifidobacterium spp. were similar to those of FOS, GOS, and inulin.
This study examines resource use (diet, habitat use, and trophic level) within and among demograp... more This study examines resource use (diet, habitat use, and trophic level) within and among demographic groups (males, females, and juveniles) of bottlenose dolphins (Tursiops truncatus). We analyzed the d 13 C and d 15 N values of 15 prey species constituting 84% of the species found in stomach contents. We used these data to establish a trophic enrichment factor (TEF) to inform dietary analysis using a Bayesian isotope mixing model. We document a TEF of 0& and 2.0& for d 13 C and d 15 N, respectively. The dietary results showed that all demographic groups relied heavily on low trophic level seagrass-associated prey. Bayesian standard ellipse areas (SEA b) were calculated to assess diversity in resource use. The SEA b of females was nearly four times larger than that of males indicating varied resource use, likely a consequence of small home ranges and habitat specialization. Juveniles possessed an intermediate SEA b , generally feeding at a lower trophic level compared to females, potentially an effect of natal philopatry and immature foraging skills. The small SEA b of males reflects a high degree of specialization on seagrass associated prey. Patterns in resource use by the demographic groups are likely linked to differences in the relative importance of social and ecological factors.
ABSTRACT This study examines resource use (diet, habitat use, and trophic level) within and among... more ABSTRACT This study examines resource use (diet, habitat use, and trophic level) within and among demographic groups (males, females, and juveniles) of bottlenose dolphins (Tursiops truncatus). We analyzed the δ13C and δ15N values of 15 prey species constituting 84% of the species found in stomach contents. We used these data to establish a trophic enrichment factor (TEF) to inform dietary analysis using a Bayesian isotope mixing model. We document a TEF of 0‰ and 2.0‰ for δ13C and δ15N, respectively. The dietary results showed that all demographic groups relied heavily on low trophic level seagrass-associated prey. Bayesian standard ellipse areas (SEAb) were calculated to assess diversity in resource use. The SEAb of females was nearly four times larger than that of males indicating varied resource use, likely a consequence of small home ranges and habitat specialization. Juveniles possessed an intermediate SEAb, generally feeding at a lower trophic level compared to females, potentially an effect of natal philopatry and immature foraging skills. The small SEAb of males reflects a high degree of specialization on seagrass associated prey. Patterns in resource use by the demographic groups are likely linked to differences in the relative importance of social and ecological factors.
The relative importance of individual microbial pathways in nitrous oxide (N(2)O) production is n... more The relative importance of individual microbial pathways in nitrous oxide (N(2)O) production is not well known. The intramolecular distribution of (15)N in N(2)O provides a basis for distinguishing biological pathways. Concentrated cell suspensions of Methylococcus capsulatus Bath and Nitrosomonas europaea were used to investigate the site preference of N(2)O by microbial processes during nitrification. The average site preference of N(2)O formed during hydroxylamine oxidation by M. capsulatus Bath (5.5 +/- 3.5 per thousand) and N. europaea (-2.3 +/- 1.9 per thousand) and nitrite reduction by N. europaea (-8.3 +/- 3.6 per thousand) differed significantly (ANOVA, f((2,35)) = 247.9, p = 0). These results demonstrate that the mechanisms for hydroxylamine oxidation are distinct in M. capsulatus Bath and N. europaea. The average delta(18)O-N(2)O values of N(2)O formed during hydroxylamine oxidation for M. capsulatus Bath (53.1 +/- 2.9 per thousand) and N. europaea (-23.4 +/- 7.2 per thousand) and nitrite reduction by N. europaea (4.6 +/- 1.4 per thousand) were significantly different (ANOVA, f((2,35)) = 279.98, p = 0). Although the nitrogen isotope value of the substrate, hydroxylamine, was similar in both cultures, the observed fractionation (delta(15)N) associated with N(2)O production via hydroxylamine oxidation by M. capsulatus Bath and N. europaea (-2.3 and 26.0 per thousand, respectively) provided evidence that differences in isotopic fractionation were associated with these two organisms. The site preferences in this study are the first measured values for isolated microbial processes. The differences in site preference are significant and indicate that isotopomers provide a basis for apportioning biological processes producing N(2)O.
A technique for measurement of the stable isotope composition of dissolved organic carbon (DOC) i... more A technique for measurement of the stable isotope composition of dissolved organic carbon (DOC) in stream water, using an elemental analyzer (EA) coupled to an isotope ratio mass spectrometer (IRMS), is described. Stream water samples were concentrated by rotary evaporation, acidified to remove dissolved inorganic carbon (DIC), and dried in silver cups prior to analysis. Precision was evaluated with standards (alanine and humic acid), and with stream water samples with varying 13 C enrichment. Standards and samples were also prepared in sealed quartz tubes for high-temperature combustion (HTC) and analyzed by dual inlet for comparison. The d 13 C values of natural abundance standards and samples measured by the two techniques differed by 0.3%. Isotopically enriched stream water samples analyzed using the EA and HTC methods had comparable d 13 C and atom % values (differed by 6%). Expensive quartz vessels and the extensive time required for sample lyophilization, combustion and cryogenic separation of combustion products before isotopic analysis for the HTC method are eliminated with the EA method. Overall, the EA method is simpler, faster, and more economical than the HTC method, and it can be used for 13 C-enriched as well as natural abundance samples.
Hydrogenases catalyze the reversible formation of H(2), and they are key enzymes in the biologica... more Hydrogenases catalyze the reversible formation of H(2), and they are key enzymes in the biological cycling of H(2). H isotopes have the potential to be a very useful tool in quantifying hydrogen ion trafficking in biological H(2) production processes, but there are several obstacles that have thus far limited the application of this tool. Here, we describe a new method that overcomes some of these barriers and is specifically designed to measure isotopic fractionation during enzyme-catalyzed H(2) evolution. A key feature of this technique is that purified hydrogenases are employed, allowing precise control over the reaction conditions and therefore a high level of precision. In addition, a custom-designed high-throughput gas chromatograph/isotope ratio mass spectrometer is employed to measure the isotope ratio of the H(2). Using our new approach, we determined that the fractionation factor for H(2) production by the [NiFe]-hydrogenase from Desulfovibrio fructosovorans is 0.273 ± 0.006. This result indicates that, as expected, protons are highly favored over deuterium ions during H(2) evolution. Potential applications of this newly developed method are discussed.
ABSTRACT We used stable isotope analysis to investigate the foraging ecology of coastal bottlenos... more ABSTRACT We used stable isotope analysis to investigate the foraging ecology of coastal bottlenose dolphins (Tursiops truncatus) in relation to a series of anthropogenic disturbances. We first demonstrated that stable isotopes are a faithful indicator of habitat use by comparing muscle isotope values to behavioral foraging data from the same individuals. δ13C values increased, while δ34S and δ15N values decreased with the percentage of feeding observations in seagrass habitat. We then utilized stable isotope values of muscle to assess temporal variation in foraging habitat from 1991 to 2010 and collagen from tooth crown tips to assess the time period 1944 to 2007. From 1991 to 2010, δ13C values of muscle decreased while δ34S values increased indicating reduced utilization of seagrass habitat. From 1944 to 1989 δ13C values of the crown tip declined significantly, likely due to a reduction in the coverage of seagrass habitat and δ15N values significantly increased, a trend we attribute to nutrient loading from a rapidly increasing human population. Our results demonstrate the utility of using marine mammal foraging habits to retrospectively assess the extent to which anthropogenic disturbance impacts coastal food webs.
Two commercial strains of Bifidobacterium spp (Bf-1 and Bf-6) were cultured in 12% (w/w) reconsti... more Two commercial strains of Bifidobacterium spp (Bf-1 and Bf-6) were cultured in 12% (w/w) reconstituted nonfat dry milk (NDM) containing 0, 0.5, 1.0, 3.0, or 5.0% (w/v) fructooligosaccharide (FOS), galactooligosaccharide (GOS), and inulin. Inoculated samples were incubated anaerobically at 37°C for 48 h. Growth and activity of the cultures in the presence of FOS, GOS, and inulin were determined. Viability of each strain was assessed after 4 weeks of refrigerated storage at 4°C. Growth promotion, enhancement of activity and retention of viability were greatest when Bifidobacterium Bf-1 and Bf-6 were grown in the presence of FOS followed in a descending order by GOS and inulin. The effects of oligosaccharides and inulin increased with increasing carbohydrate concentration and was maximal at 5% (w/v).
Polyclonal antibodies (PAb) prepared against bovine milk alkaline phosphatase (ALP) were used to ... more Polyclonal antibodies (PAb) prepared against bovine milk alkaline phosphatase (ALP) were used to develop a competitive indirect (CI) ELISA. Anti-ALP PAb were specific for milk ALP and did not react with ALP from E. coli or bovine and calf intestinal mucosa. Anti-ALP PAb were 20% cross-reactive with bovine placenta ALP. The anti-ALP antibodies also did not recognize bovine serum albumin, acid glycoprotein, ovalbumin, ferritin, and casein, although some cross-reactivity was observed with whey protein isolate. Anti-ALP PAbs reacted with deglycosylated native ALP, but did not recognize ALP denatured at 100 degrees C in 2% SDS or deglycosylated denatured ALP. When buffered solutions of milk ALP were heated at 70 degrees C, ALP activity decreased at a faster rate than ALP content determined by CI-ELISA. The ELISA differentiated between native and heat denatured ALP. Further studies are warranted to determine if an ELISA can be used to verify pasteurization of fluid milk.
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Papers by Hasand Gandhi