Investigations on Xenopus laevis body composition and feeding behavior in a laboratory setting.

The African clawed frog, Xenopus laevis, has been used as a laboratory animal for decades in many research areas. However, there is a lack of knowledge about the nutritional physiology of this amphibian species and the feeding regimen is not standardized. The aim of the present study was to get more insights into the nutrient metabolism and feeding behavior of the frogs. In Trial 1, adult female X. laevis were fed either a Xenopus diet or a fish feed. After 4 weeks, they were euthanized, weighed, measured for morphometrics and dissected for organ weights and whole-body nutrient analysis. There were no significant differences between the diet groups regarding the allometric data and nutrient contents. The ovary was the major determinant of body weight. Body fat content increased with body weight as indicator of energy reserves. In Trial 2, 40 adult female frogs were monitored with a specifically developed digital tracking system to generate heat-maps of their activity before and up to 25 min after a meal. Three diets (floating, sinking, floating & sinking) were used. The main feed intake activity was fanning the feed into the mouth, peaking until 20 min after the meal. The different swimming characteristics of the diets thereby influenced the activity of the animals. Our dataset helps to adjust the feeding needs to the physical composition and also to meet the natural behavioral patterns of feed intake as a prerequisite of animal wellbeing and animal welfare in a laboratory setting.

Digestibility of crude nutrients and minerals in C57Bl/6J and CD1 mice fed a pelleted lab rodent diet.

In laboratory animals, there is a scarcity of digestibility data under non-experimental conditions. Such data is important as basis to generate nutrient requirements, which contributes to the refinement of husbandry conditions. Digestibility trials can also help to identify patterns of absorption and potential factors that influence the digestibility. Thus, a digestibility trial with a pelleted diet used as standard feed in laboratory mice was conducted. To identify potential differences between genetic lines, inbred C57Bl/6 J and outbred CD1 mice (n = 18 each, male, 8 weeks-old, housed in groups of three) were used. For seven days, the feed intake was recorded and the total faeces per cage collected. Energy, crude nutrient and mineral content of diet and faecal samples were analyzed to calculate the apparent digestibility (aD). Apparent dry matter and energy digestibility did not differ between both lines investigated. The C57Bl/6 J mice had significantly higher aD of magnesium and potassium and a trend towards a lower aD of sodium than the mice of the CD1 outbred stock. Lucas-tests were performed to calculate the mean true digestibility of the nutrients and revealed a uniformity of the linear regression over data from both common laboratory mouse lines. The mean true digestibility of crude nutrients was > 90%, except for fibre, that of the minerals ranged between 66 and 97%.

Comparative analysis of pancreatic amylase activity in laboratory rodents.

Alpha-amylase is the main enzyme for starch digestion in the mammalian gastrointestinal tract. There are species differences in the enzymatic activity of pancreatic amylase that are related to the digestive strategy and natural diet of a species. This aspect is well investigated in pet and farm animals, while in common laboratory animal rodents, information is scarce. In the context of the 3R concept, detailed knowledge of the digestive physiology should be the basis of adequate nutrition, experimental planning and data interpretation. The present study aimed to obtain reference data on amylase activity in pancreatic tissue and duodenal digesta in laboratory mice, rats and hamsters. In addition, digesta was stained with Lugol's iodine to visualize starch in the process of degradation throughout the gastrointestinal tract. Amylase activity in pancreatic tissue and duodenal digesta was significantly lower in hamsters than rats and mice. The Lugol staining showed intense starch degradation in the hamsters' forestomachs, presumably by microbial fermentation. A possible explanation is that the prae-duodenal microbial starch fermentation enhances digestibility and reduces the need for pancreatic amylase in hamsters. Rats and mice may rely more on pancreatic amylase for prae-caecal starch digestion, while the microbial fermentation is mainly located in the caecum. The results clearly show species differences in the digestive capacity for starch in mice, rats and hamsters that need to be considered in the feeding of these species in the laboratory setting as well as in the use of rodents as translational animal models.

True mineral digestibility in C57Bl/6J mice.

Data on mineral digestibility is key to understand mineral homeostasis and refine the recommendations for the dietary intake of these nutrients. In farm animals and pets, there is plenty of data on mineral digestibility and influencing factors. In laboratory mice, however, there is a lack of information on mineral digestibility under maintenance conditions, although this should be the basis for studies on mineral homeostasis under experimental conditions. The aim of the present study was to analyse data on intake, faecal excretion, and apparent digestibility of calcium, phosphorus, sodium, potassium, and magnesium in C57BL/6J mice fed different maintenance diets with varying voluntary dry matter intake. Lucas-tests were used to quantify true digestibility and describe correlations between dietary intake and excretion/absorption of the nutrients. Calcium, phosphorus, and magnesium showed a linear correlation between intake and faecal excretion (R2: 0.77, 0.93 and 0.91, respectively). Intake and apparently digested amounts of sodium and potassium were correlated linearly (R2: 0.86 and 0.98, respectively). These data show that intake is the major determinant of absorption in the minerals listed above. Faecal calcium and phosphorus excretion were correlated as well (R2 = 0.75).

Morphometrics of Xenopus laevis Kept as Laboratory Animals.

Morphometric data that provide information on body conditions can be used to monitor the health and well-being of animals. In laboratory animals, they can help to evaluate the stress due to experiments or treatments, following the 3R principles. The aim of the present study was to obtain morphometric data of male and female African clawed frogs, Xenopus laevis, as the bases for body condition evaluations. Adult frogs (n = 198) were weighed and standardized photographs were taken. The photographs were used to determine several measurements (length, cranial width, caudal width, thigh width). In addition, a triangle was drawn to outline each frog's simplified body form, and the triangle surface was calculated. In conclusion, the triangle surface drawn on the dorsal plane of each frog correlated with the body weight of the females. There were significant differences between the body weights and sizes of male and female frogs, with males being smaller (p < 0.001). Based on the morphometric data, females could be assigned to five groups in which an assessment of the animal's well-being is feasible.

Chemical composition of snakes.

The present study was carried out to provide insight into the body composition of snakes, which is an important basis for determination of nutrient requirement and physiological processes. Carcasses of 86 captive snakes (31 pythons, 32 colubrids and 23 boas) were available for analysis. Skins and vertebrae bones of 11 snakes and livers of 64 snakes were analysed separately from the carcasses. Crude nutrients, major minerals and trace elements were investigated. The content of crude nutrients of the whole body was similar to those of mammals and birds. Relatively high contents of copper, zinc and especially of iron (up to 23,973 mg/kg dry matter) were found in the body, particularly in the liver. There was an increase of the iron content of the whole body over age.

Current Feeding Practice of Xenopus laevis in a Laboratory Setting.

African clawed frogs are common animal models used in various research areas. However, husbandry and especially feeding regimens are not nearly as standardized as is established for other laboratory animals. We recorded the diets and feeding protocols commonly used in laboratory practice in a questionnaire (18 responses). The survey revealed a wide variety of housing conditions. Feeding protocols and, in particular, diet composition varied considerably between facilities. While diets tailored to Xenopus were used in the majority, differences in feeding frequency and dietary components were noted. From five responses, the weekly feed intake per frog could be calculated, showing considerable differences in dry matter intake (1.37-5.4 g). The labelled nutrient content of the diets fed in the facilities (n = 10) met the recommendations in most cases, with protein as the major energy source. However, the mineral content varied markedly between diets. Both floating and sinking diets were used, while quickly sinking diets were associated with feed leftovers. Feed processing may likely influence feed intake behavior. Further research is needed to ensure standardization for aquatic species with respect to husbandry systems, feeding regimens, and especially the nutrient composition of feeds. Furthermore, this work will contribute positively to animal welfare and the comparability of research results.

Influence of Strain and Diet on Urinary pH in Laboratory Mice.

Acid base homeostasis and urine pH is influenced by the dietary cation anion balance (DCAB) in many species. Here, a negative DCAB acidifies the urine, while higher DCABs alkalize the urine. The dimension of the DCAB effect can be species-specific, because of differences in urine buffer systems. The aim of the present study was to describe the response of laboratory mice to diets with different DCAB. We used 8-week-old wildtype male mice of the C57Bl/6J inbred strain and CD1 outbred stock. Three groups (n = 15 animals/group) were formed and fed standard diet A for adaptation. For the 7-week feeding trial, mice were either kept on diet A (DCAB -7 mmol/kg dry matter (DM) or switched to diet B (246 mmol/kg DM) or C (-257 mmol/kg DM). Urine pH was measured weekly from a pooled sample per cage. There was a significant difference in the basal urine pH on diet A between C57Bl6/J and CD1 mice. The shift in urine pH was also significantly different between the two groups investigated.

Processing Matters in Nutrient-Matched Laboratory Diets for Mice-Microbiome.

The composition of the microbiome is subject to the host's diet. In commercial laboratory mouse diets, different physical forms of the same diets are available, containing-according to their labels-identical ingredients and nutrient compositions. However, variations in nutrient composition and starch gelatinization due to production processes and their impact on digestibility have been described. In this study, a total of 48 C57BL/J6 mice were assigned to two equal groups and were fed diets (produced with different processes-extruded vs. pelleted) for eight weeks in two biological replicates. At the end of the experiment, samples were collected from five different gastrointestinal regions, including the stomach, small intestine, cecum, large intestine, and an extracorporeal region (feces), and the microbiome was analyzed with 16S rRNA gene amplicon sequencing. The replicates in both experiments differed significantly in their relative abundances of Muribaculaceae species. Furthermore, the gastrointestinal content of pellet-fed mice contained larger numbers of Lactobacillus species. These results indicate that starch gelatinization and ingredient composition significantly influence microbial makeup. In conclusion, different feed processing methods may affect fundamental digestive and metabolic processes, impacting animal experiments and biasing microbiome data.

Processing Matters in Nutrient-Matched Laboratory Diets for Mice-Energy and Nutrient Digestibility.

Starch gelatinization is a major determinant of carbohydrate digestibility and varies with diet processing. Laboratory rodent diets are often marketed as identical, but are sold in different forms, regardless of the markedly higher starch gelatinization in extruded than in pelleted diets. Our hypothesis was that this would impact energy and nutrient digestibility in mice fed pellets or extrudate, respectively. Trial 1 showed that feeding C57BL/6 mice a standard maintenance diet in extruded form results in a significantly higher digestibility of organic matter, energy, and carbohydrates than the identical diet in pelleted form. The replication of the experiment, however, revealed a variation between batches of the same pelleted diet regarding starch and total dietary fiber contents. Given the significant differences in diet digestibility and the potential impacts of digestibility on nutrient utilization, the intestinal microbiome, and intermediary metabolism, trials performed with differently processed diets are not comparable. This might partly explain failures to reproduce results, especially in gastrointestinal or microbiome research. Considering this impact on experimental animals, the degree of starch gelatinization should be declared in the diet information for laboratory animal diets. The differences between batches of laboratory animal diets as observed in the pellets are not acceptable.

Metabolisable energy intake and growth of privately owned growing dogs in comparison with official recommendations on the growth curve and energy supply.

The correct assumption of metabolisable energy (ME) requirement is essential for the nutrition consultation and diet formulation. In young dogs, too high energy supply can accelerate growth and thus lead to developmental orthopaedic diseases. The aim of the present study was to collect the data on ME intake and body weight (BW) development in privately owned growing dogs in order to compare these data with the current recommendations. Our hypothesis was that the actual ME intake of healthy young dogs would be lower than the actual recommendation. The data of 493 privately owned puppies (median age at first consultation 21 weeks, the median expected mature BW 30 kg) on ME intake, actual and expected mature BW were collected and compared with recommendations of the Society of Nutrition Physiology (GfE, Meyer and Zentek and NRC). In 243 dogs, there was a follow-up. The actual BW did not deviate systematically from the calculated expected BW (R2  = .929). The ME intake significantly decreased with age (p < .05) and significantly increased with expected mature BW (p < .05). There was no significant interaction between these two parameters (p > .05). Sex had no effect on the ME intake (p > .05). The ME intake of young dogs with a history of skeletal problems or of food allergy did not differ systematically from healthy dogs of similar age and expected mature BW. The ME intake was considerably below NRC recommendations, especially in younger puppies (>8-17 weeks: 78%, >17-26 weeks: 83% of NRC recommendation). A predictive linear equation for ME intake was developed: ME intake (MJ) = (1.063 - 0.565 × [actual BW/expected mature BW]) × actual BW0.75 .