M.J.W. Heetkamp

An exploration on greenhouse gas and ammonia production by insect species suitable for animal or human consumption.

Background Greenhouse gas (GHG) production, as a cause of climate change, is considered as one of the biggest problems society is currently facing. The livestock sector is one of the large contributors of anthropogenic GHG emissions. Also, large amounts of ammonia (NH3), leading to soil nitrification and acidification, are produced by livestock. Therefore other sources of animal protein, like edible insects, are currently being considered. Methodology/Principal Findings An experiment was conducted to quantify production of carbon dioxide (CO2) and average daily gain (ADG) as a measure of feed conversion efficiency, and to quantify the production of the greenhouse gases methane (CH4) and nitrous oxide (N2O) as well as NH3 by five insect species of which the first three are considered edible: Tenebrio molitor, Acheta domesticus, Locusta migratoria, Pachnoda marginata, and Blaptica dubia. Large differences were found among the species regarding their production of CO2 and GHGs. The insects in this study had a higher relative growth rate and emitted comparable or lower amounts of GHG than described in literature for pigs and much lower amounts of GHG than cattle. The same was true for CO2 production per kg of metabolic weight and per kg of mass gain. Furthermore, also the production of NH3 by insects was lower than for conventional livestock. Conclusions/Significance This study therefore indicates that insects could serve as a more environmentally friendly alternative for the production of animal protein with respect to GHG and NH3 emissions. The results of this study can be used as basic information to compare the production of insects with conventional livestock by means of a life cycle analysis.

Effect of eggshell temperature and oxygen concentration on survival rate and nutrient utilization in chicken embryos

Environmental conditions during incubation such as temperature and O(2) concentration affect embryo development that may be associated with modifications in nutrient partitioning. Additionally, prenatal conditions can affect postnatal nutrient utilization. Using broiler chicken embryos, we studied the effects of eggshell temperature (EST; 37.8 or 38.9 degrees C) and O(2) (17, 21, or 25%) applied from d 7 until 19 of incubation in a 2 x 3 factorial design. Effects of these factors on embryonic survival, development, and nutrient utilization were assessed in the pre- and posthatch period. High EST reduced yolk-free body mass compared with normal EST (36.1 vs. 37.7 g), possibly through reduced incubation duration (479 vs. 487 h) and lower efficiency of protein utilization for growth (83.6 vs. 86.8%). Increasing O(2) increased yolk-free body mass (from 35.7 to 38.3 g) at 12 h after emergence from the eggshell, but differences were larger between the low and normal O(2) than between the normal and high O(2). This might be due to the lower efficiency of nutrient utilization for growth at low O(2). However, the effects of O(2) that were found at 12 h were less pronounced at 48 h posthatch. When O(2) was shifted to 21% for all treatments at d 19 of incubation, embryos incubated at low O(2) used nutrients more efficiently than those incubated at normal or high O(2). An additional negative effect on survival and chick development occurred when embryos were exposed to a combination of high EST and low O(2). Possible explanations include reduced nutrient availability for hatching, decreased body development to fulfill the energy-demanding hatching process, and higher incidence of malpositions. In conclusion, EST and O(2) during incubation affect nutrient utilization for growth, which may explain differences in survival and development. Embryos raised under suboptimal environmental conditions in the prenatal period may develop adaptive mechanisms that still continue in the posthatch period.

High Environmental Temperature Increases Glucose Requirement in the Developing Chicken Embryo

Environmental conditions during the perinatal period influence metabolic and developmental processes in mammals and avian species, which could impact pre- and postnatal survival and development. The current study investigated the effect of eggshell temperature (EST) on glucose metabolism in broiler chicken embryos. Broiler eggs were incubated at a high (38.9°C) or normal (37.8°C) EST from day 10.5 of incubation onward and were injected with a bolus of [U-13C]glucose in the chorio-allantoic fluid at day 17.5 of incubation. After [U-13C]glucose administration, 13C enrichment was determined in intermediate pools and end-products of glucose metabolism. Oxidation of labeled glucose occurred for approximately 3 days after injection. Glucose oxidation was higher in the high than in the normal EST treatment from day 17.6 until 17.8 of incubation. The overall recovery of 13CO2 tended to be 4.7% higher in the high than in the normal EST treatment. An increase in EST (38.9°C vs 37.8°C) increased 13C enrichment in plasma lactate at day 17.8 of incubation and 13C in hepatic glycogen at day 18.8 of incubation. Furthermore, high compared to normal EST resulted in a lower yolk-free body mass at day 20.9 (−2.74 g) and 21.7 (−3.81 g) of incubation, a lower hepatic glycogen concentration at day 18.2 (−4.37 mg/g) and 18.8 (−4.59 mg/g) of incubation, and a higher plasma uric acid concentration (+2.8 mg/mL/+43%) at day 21.6 of incubation. These results indicate that the glucose oxidation pattern is relatively slow, but the intensity increased consistently with an increase in developmental stage of the embryo. High environmental temperatures in the perinatal period of chicken embryos increased glucose oxidation and decreased hepatic glycogen prior to the hatching process. This may limit glucose availability for successful hatching and could impact body development, probably by increased gluconeogenesis from glucogenic amino acids to allow anaerobic glycolysis.

Effects of fermentable starch and straw-enriched housing on energy partitioning of growing pigs

Both dietary fermentable carbohydrates and the availability of straw bedding potentially affect activity patterns and energy utilisation in pigs. The present study aimed to investigate the combined effects of straw bedding and fermentable carbohydrates (native potato starch) on energy partitioning in growing pigs. In a 2 × 2 factorial arrangement, 16 groups of 12 pigs (approximately 25 kg) were assigned to either barren housing or housing on straw bedding, and to native or pregelatinised potato starch included in the diet. Pigs were fed at approximately 2.5 times maintenance. Nitrogen and energy balances were measured per group during a 7-day experimental period, which was preceded by a 30-day adaptation period. Heat production and physical activity were measured during 9-min intervals. The availability of straw bedding increased both metabolisable energy (ME) intake and total heat production (P < 0.001). Housing conditions did not affect total energy retention, but pigs on straw bedding retained more energy as protein (P < 0.01) and less as fat (P < 0.05) than barren-housed pigs. Average daily gain (P < 0.001), ME intake (P < 0.001) and energy retention (P < 0.01) were lower in pigs on the native potato starch diet compared to those on the pregelatinised potato starch diet. Pigs on the pregelatinised potato starch diet showed larger fluctuations in heat production and respiration quotient over the 24-h cycle than pigs on the native potato starch diet, and a higher activity-related energy expenditure. The effect of dietary starch type on activity-related heat production depended, however, on housing type (P < 0.05). In barren housing, activity-related heat production was less affected by starch type (16.1% and 13.7% of total heat production on the pregelatinised and native potato starch diet, respectively) than in straw-enriched housing (21.1% and 15.0% of the total heat production on the pregelatinised and native potato starch diet, respectively). In conclusion, the present study shows that the availability both of straw bedding and of dietary starch type, fermentable or digestible, affects energy utilisation and physical activity of pigs. The effects of housing condition on protein and fat deposition suggest that environmental enrichment with long straw may result in leaner pigs. The lower energy expenditure on the physical activity of pigs on the native potato starch diet, which was the most obvious in straw-housed pigs, likely reflects a decrease in foraging behaviour related to a more gradual supply of energy from fermentation processes.

Design of climate respiration chambers, adjustable to the metabolic mass of subjects

Open-circuit respiration chambers can be used to measure gas exchange and to calculate heat production (Q) of humans and animals. When studying short-term changes in Q, the size of the respiration chamber in relation to the subject of study is a point of concern. The washout time of a chamber, defined as the proportion of the chamber size to the rate of ventilation, needs to be minimised for accurate measurement of short term changes in Q. To date, most respiration chambers have a fixed size, limiting their use for different species, sizes and number of subjects, thus hampering studying the short term dynamics of Q. This chapter presents various approaches to the design, construction and testing of respiration chambers, adjustable to the metabolic mass inside. As investment costs for constructing respiration chambers are high, flexibility in the use of chambers can contribute substantially to an efficient use of resources. Furthermore, an outline is given to sensor criteria and calibration and finally, the validation of a whole indirect-calorimetric system is described. Air leak tolerance is defined and attention is paid to caretaking of animals, excreta collection and animal and personnel welfare and safety. Respiration facilities, recently constructed at Wageningen University are presented as an example. Briefly, four 45 m2 climate chambers can be used, e.g. for heat or cold stress experiments, to incubate eggs or as a hygiene barrier. Within each chamber, one or two smaller airtight, size adaptable respiration rooms, can be built in where ambient temperature, humidity and ventilation rate can be controlled independently. In each respiration room a wide range of ventilation flow rates can be accomplished and both hypobaric and hyperbaric air pressure control can be established, allowing energy metabolism experiments with specific pathogen free animals (hyperbaric) or trials with infectious agents (hypobaric).

Asynchronous Supply of Indispensable Amino Acids Reduces Protein Deposition in Milk-Fed Calves

A balanced supply of indispensable amino acids (AA) is required for efficient protein synthesis. Different absorption kinetics (e.g., free vs. protein-bound AA) may, however, create asynchrony in postabsorptive availability of individual AA, thereby reducing the efficiency of protein deposition. We studied the effects of AA asynchrony on protein metabolism in growing, milk-fed calves. In 2 experiments, each with a change-over design including 8 calves, a milk replacer deficient in Lys and Thr was used. In Expt. 1, L-Lys and L-Thr were parenterally supplemented, either in synchrony (SYN), asynchrony (ASYN), or partial asynchrony (PART) with dietary AA. In Expt. 2, l-Lys and l-Thr were orally supplemented, either in SYN or ASYN with dietary AA. In Expt. 1, digested protein was used less efficiently for growth for ASYN (31.0%) than for SYN (37.7%), with PART being intermediate (36.0%). Indicator AA oxidation tended (P = 0.06) to be higher for ASYN. In Expt. 2, the efficiency of protein utilization was lower for ASYN (34.9%) than for SYN (46.6%). Calves spared AA from oxidation when the limiting AA were provided in excess after a short period (<24 h) of deprivation. Restoring AA balance by parenteral supplementation resulted in a 19% lower efficiency of digestible protein utilization than by oral supplementation, likely caused by splanchnic oxidation of imbalanced AA in excess to Thr. In conclusion, asynchronous availability of individual indispensable AA reduces the efficiency by which digested protein is retained in milk-fed calves. Furthermore, an AA imbalance in the splanchnic tissues may result in disproportionate AA oxidation.

Moving from a complete energy balance towards substrate oxidation: use of stable isotopes

Substrate oxidation can be estimated from gas exchange in combination with urinary nitrogen excretion measurements. The estimates are net rates of oxidation, because several metabolic processes, such as lipogenesis and gluconeogenesis, may affect the respiratory quotient, resulting in under- or overestimation of absolute rates of substrate oxidation. If these limitations, as well as accumulation of ketone bodies, lactate and urea, are taken into consideration, indirect calorimetry can be an effective method to quantify substrate oxidation in animals and humans. Methane should be excluded from the equations for calculating oxidation rates. Using 13C labelled substrates provides additional possibilities in oxidation studies. Whole body nutrient oxidation can be measured when tracer amounts of the nutrient of interest are continuously infused and when 13CO2 expiration and 13C enrichment of the precursor for oxidation are measured in blood plasma. Oxidation of exogenous substrates can be distinguished from that of endogenous substrates by using 13C enriched (or depleted) nutrients in the diet. In addition, labelling of individual monosaccharides, fatty acids or amino acids can provide insight in the metabolic fate of the tracee irrespective of availability of other substrates from the same macronutrient class (i.e. carbohydrate, fat, protein). Finally, selection of appropriate isotopomers may allow quantification of pre-identified metabolic processes (e.g. lipogenesis). Fluctuations in background enrichment, isotope administration strategy, sequestration of CO2 and isotopic gradient in the body are among the factors that should be considered when designing and interpreting oxidation studies with labelled nutrients. Modelling approaches should be in place or be developed to maximise the learnings from these studies. This appears especially challenging when animals are studied in a non-steady physiological state. In conclusion, using 13C labelling in combination with indirect calorimetry provides unique opportunities to quantify the contribution of individual substrates (endogenous as well as exogenous) to whole body energy metabolism.

Effects of prefermented cereals or the end products of fermentation on growth and metabolism of enterocyte-like Caco-2 cells and on intestinal health of restrictedly fed weanling pigs.

To unravel the underlying mechanisms that explain the positive effects of prefermented cereals on in vivo gastrointestinal (GI) architecture and function, an in vitro experiment using a human small intestinal epithelial cell model (Caco-2) was performed. A range of dilutions (0% to 10%) of the supernatants of three liquid experimental diets, as well as Na-lactate were used in an in vitro experiment to assess their effect on cellular growth, metabolism, differentiation and mucosal integrity using Caco-2. The experimental diets contained, in addition to a protein rich basal diet (60%), (1) a liquid control diet (C) containing 40% of a mixture of barley and wheat (ratio 3 : 1) or (2) a liquid diet (F) containing 40% prefermented barley and wheat or (3) C with the addition of the fermentation end-products (organic acids and ethanol) in concentrations similar to those in the fermented diet (FP). For F, the mixture of barley and wheat was fermented at 35°C for 48 h. Parallel to the in vitro experiment, 18 groups of eight weanling pigs were assigned to one of the experimental diets during a 14-day in vivo experiment. Each group was fed restrictively. The results of the in vitro experiment showed that the lowest dose of both F- and FP-supernatants had no clear effects on the cell proliferation, but incubation with 5% and 10% of the F- and FP-supernatants decreased the cell numbers at day 19. DNA, RNA, protein and glycoprotein synthesis in differentiated Caco-2 cells were stimulated by incubation with the lower concentrations (0.5% to 2.5%) of F- and FP-supernatants whereas the higher concentrations (5% and 10%) had no effect. Both the F- and FP-supernatants decreased the specific sucrase-isomaltase activity in a dose-dependent manner, but the effects on the specific aminopeptidase activities were less clear. Mucosal integrity initially decreased after incubation with the highest F- and FP-supernatants and started to recover between 24 and 48 h. The results of the in vivo experiment showed no dietary effects (P > 0.1) on GI morphology and brush-border enzyme activities at day 5 or at day 14. Time related changes in GI characteristics followed a normal pattern. In conclusion, the supernatants of diets containing either prefermented cereals or their fermentation end-products clearly modulate cellular growth, metabolism, differentiation and mucosal integrity in an in vitro model, although these effects were not observed in the in vivo characteristics measured in weanling pigs.

Effects of breeder age, broiler strain, and eggshell temperature on development and physiological status of embryos and hatchlings

Breeder age and broiler strain can influence the availability of nutrients and oxygen, particularly through differences in yolk size and shell conductance. We hypothesized that these egg characteristics might affect embryonic responses to changes in eggshell temperature (EST). This study aimed to investigate the effect of breeder age, broiler strain, and EST on development and physiological status of embryos. A study was designed as a 2 × 2 × 2 factorial arrangement using 4 batches of 1,116 hatching eggs of 2 flock ages at 29 to 30 wk (young) and 54 to 55 wk (old) of Ross 308 and Cobb 500. EST of 37.8 (normal) or 38.9°C (high) was applied from incubation d 7 (E7) until hatching. The results showed that breeder age rather than broiler strain had an influence on yolk size (P = 0.043). The shell conductance was higher in Ross 308 than in Cobb 500 (P < 0.001). A high EST resulted in a higher yolk free body mass (YFBM) compared to the normal EST at E14 and E16, but at 3 h after hatch YFBM was lower when eggs were incubated at high EST compared to normal EST (all P < 0.001). Cobb 500 eggs yielded embryos with a lower YFBM at E14, E18, and 3 h after hatch (all P < 0.05) than Ross 308 eggs. Breeder age had no effect on YFBM, but the RSY weight was higher in embryos from the old flock compared to the young flock embryos at E14 and E16 (both P < 0.05). A 3-way interaction among breeder age, strain, and EST was found, especially for incubation duration, navel quality, and relative heart and stomach weights at 3 h after hatch (all P < 0.05). Based on the results obtained, we conclude that oxygen availability rather than nutrient availability determines embryonic development, and the egg characteristics affected embryonic responses to changes of EST, especially for variables related to chick quality.

Effects of breeder age, strain, and eggshell temperature on nutrient metabolism of broiler embryos

&NA; Breeder age and broiler strain influence the availability of nutrients and oxygen through yolk size and eggshell conductance, and the effects of these egg characteristics on nutrient metabolism might be influenced by eggshell temperature (EST). This study aims to determine effects of breeder age, strain, and EST on nutrient metabolism of embryos. A study was designed as a 2 × 2 × 2 factorial arrangement using four batches of in total 4,464 hatching eggs of 2 flock ages at 29 to 30 wk (young) and 54 to 55 wk (old) of Ross 308 and Cobb 500. EST of 37.8 (normal) or 38.9°C (high) was applied from incubation day 7 (E7) until hatching. Wet yolk weight was determined mainly by breeder age (P = 0.043). Energy content in yolk (P = 0.004) and albumen + yolk (P = 0.005) were higher in old flock eggs than in young flock eggs, but did not differ between broiler strains. Eggshell conductance was higher in Ross 308 eggs than in Cobb 500 eggs (P < 0.001). Old flock embryos used more energy (P = 0.046) and accumulated more energy into yolk free body mass (YFBM; P = 0.030) than young flock embryos, whereas heat production (HP), energy lost, and efficiency of converting energy used to YFBM (EYFB) did not differ. Ross 308 embryos used more energy (P = 0.006), had a higher energy lost (P = 0.010), and a higher HP between E15 to E18 (P < 0.05) than Cobb 500 embryos. Energy content in YFBM did not differ between strains and EYFB (P = 0.024) was lower in Ross 308 than in Cobb 500. High EST resulted in higher HP than low EST from E11 to E15 (P < 0.05), but not after E15. Amount of energy used (P = 0.006) and energy accumulated in the YFBM (P < 0.001) was lower for embryos incubated at an EST of 38.9 than that of 37.8°C, whereas EYFB did not differ. In conclusion, breeder age, broiler strain, and EST differentially influence embryonic metabolism and particularly the availability of oxygen could have contributed to these differences.