A Novel Breast Cancer Xenograft Model Using the Ostrich Chorioallantoic Membrane-A Proof of Concept.

The avian chorioallantoic membrane (CAM) assay has attracted scientific attention in cancer research as an alternative or complementary method for in vivo animal models. Here, we present a xenograft model based on the ostrich (struthio camelus) CAM assay for the first time. The engraftment of 2 × 106 breast cancer carcinoma MDA-MB-231 cells successfully lead to tumor formation. Tumor growth monitoring was evaluated in eight fertilized eggs after xenotransplantation. Cancer cells were injected directly onto the CAM surface, close to a well-vascularized area. Histological analysis confirmed the epithelial origin of tumors. The CAM of ostrich embryos provides a large experimental surface for the xenograft, while the comparably long developmental period allows for a long experimental window for tumor growth and treatment. These advantages could make the ostrich CAM assay an attractive alternative to the well-established chick embryo model. Additionally, the large size of ostrich embryos compared to mice and rats could help overcome the limitations of small animal models. The suggested ostrich model is promising for future applications, for example, in radiopharmaceutical research, the size of the embryonal organs may compensate for the loss in image resolution caused by physical limitations in small animal positron emission tomography (PET) imaging.

Ostrich oil as a fat substitute in milk-based infant formula.

In this study, the possibility of replacing vegetable fats with ostrich oil in infant formula (IF) production was investigated. The fatty acid profile, the positional distribution of fatty acids in the triacylglycerols, the cholesterol content, and the physicochemical properties of ostrich oil were determined and compared with breast milk fat and vegetable oils. In the next step, two infant formulas were produced using ostrich oil and vegetable oils and the physicochemical properties, rheological properties, color parameters, and sensory analysis of the resultant powders were compared. The results showed that the predominant fatty acids in ostrich oil are palmitic acid, oleic acid, and linoleic acid which is similar to breast milk fat and vegetable oils. The presence of appropriate cholesterol content in ostrich oil makes it more similar to breast milk fat compared to vegetable fats. Palmitic acid was located at sn-2 position in 15% triacylglycerol from ostrich fat, which was equal to the amount reported for vegetable fats. The incorporation of ostrich oil in infant formula production showed that there is no statistically significant difference between quality attributes of powder formulated with ostrich oil or vegetable oils. Therefore, ostrich oil can be introduced as a new source of edible oil, and addition of ostrich oil is an effective way to reduce the gap between the composition of breast milk and infant formula.

Smart Biomechanical Adaptation Revealed by the Structure of Ostrich Limb Bones.

Ostriches are known to be the fastest bipedal animal alive; to accomplish such an achievement, their anatomy evolved to sustain the stresses imposed by running at such velocities. Ostriches represent an excellent case study due to the fact that their locomotor kinematics have been extensively studied for their running capabilities. The shape and structure of ostrich bones are also known to be optimized to sustain the stresses imposed by the body mass and accelerations to which the bones are subjected during movements. This study focuses on the limb bones, investigating the structure of the bones as well as the material properties, and how both the structure and material evolved to maximise the performance while minimising the stresses applied to the bones themselves. The femoral shaft is hollowed and it presents an imbricate structure of fused bone ridges connected to the walls of the marrow cavity, while the tibial shaft is subdivided into regions having different mechanical characteristics. These adaptations indicate the optimization of both the structure and the material to bear the stresses. The regionalization of the material highlighted by the mechanical tests represents the capability of the bone to adapt to external stimuli during the life of an individual, optimizing not only the structure of the bone but the material itself.

Subtotal Cervical Esophagectomy and Anastomosis in an Ostrich (Struthio camelus domesticus).

An adult, female, captive ostrich (Struthio camelus domesticus) was referred to a veterinary teaching hospital for a 2-week history of lethargy and a mass effect in the proximal cervical region. Physical examination revealed a fistula in the middle cervical esophagus surrounded by devitalized and necrotic tissue; feed material was found leaking from the site. Cervical radiography identified an esophageal stricture with anterior dilation due to the accumulation of feed. After receiving supportive care for 48 hours, the patient's overall status improved, allowing partial esophagectomy and resection of the affected tissues with end-to-end anastomosis. Postoperative management included fasting for 24 hours, followed by the administration of a liquid hand-rearing formula prepared with commercially available ostrich feed and administered via a feeding tube for 15 days. Proper healing of the surgical site was confirmed by esophagoscopy using a flexible endoscope 17 days after surgery. The ostrich was discharged after 27 days, with no complications recorded within the 180 days of the follow-up period. Partial cervical esophagectomy with end-to-end anastomosis along with pre- and postoperative management provided a successful outcome for the treatment of a fistulated esophageal stricture in a captive ostrich, resulting in full recovery without surgical complications.

Comparison of embryonic development, from HH21 to HH40, between ostrich (Struthio camelus) and chicken (Gallus gallus).

BACKGROUND: The chicken has been a representative model organism to study embryonic development in birds, however important differences exist among this class of species. As a representative of one of oldest existing clades of birds, the African ostrich (Struthio camelus), has the largest body among birds, and has two toes. Our purpose is to establish the corresponding stages in ostrich embryo development that match the well-established HH system of the chicken to facilitate comparative studies between the ostrich and other birds to better understand differences in development. RESULTS: Here we describe in detail the middle period of embryonic development using microscopic images and skeletal staining. We found that clear morphological differentiation between the ostrich and the chicken begins at stage 26. Bird limb cartilage first form in stage 25, while the development of the limb skeletons differs after stage 31. Calcification of limb skeletons in the chicken was completed faster. The first and second toes of the ostrich disappear at stages 36 and 38, respectively. CONCLUSIONS: This study should greatly aid ostrich-related developmental and morphological research and provide a reference for studying the development and evolution of avian limb skeletons, including molecular research. Questions that can now be addressed include studies into the fusion of tarsometatarsal skeleton, ossification, and digit loss.

Morphological and Transcriptomic Analysis of the Supplemental Boron in the Liver of Ostrich Chicks.

African ostrich chicks (Struthio camelus) were divided into six groups, and each received different levels of boric acid (source of boron) in the drinking water (0, 40, 80, 160, 320, and 640 mg/L respectively) to examine the histological, apoptotic, biochemical, and transcriptomic parameters. Morphological analysis in different groups was assessed by hematoxylin and eosin (H&E) staining, periodic acid Schiff (PAS) staining, and terminal deoxynucleotide transferase dUTP Nick-End Labeling (TUNEL) assay. The biochemical profile was evaluated spectrophotometrically. Detailed RNA-Seq of the data was performed using the transcriptomic method. H&E staining showed well-developed liver structure up to the 160 mg/L boric acid (BA) supplement groups, while BA doses (320 mg/L and 640 mg/L) caused changes in hepatocytes and portal triads. PAS staining showed that glycogen levels were optimal in the 80 mg/L BA dose group, but a reduction in glycogen levels was observed after this group, particularly in the 640 mg/L BA supplement group. Cellular apoptosis showed a biphasic pattern, and the BA dose above 160 mg/L enhanced cell death. In addition, serum analysis showed that doses of 80-160 mg BA were beneficial for ostrich liver. Then, the transcriptome analysis of the 80 mg dose also showed mainly positive effects on the liver. These results demonstrated that chronic BA exposure (320-640 mg) can cause significant histological, apoptotic, and biochemical changes in African ostrich liver, while the adequate dose of supplementation (particularly 80 mg BA) promotes liver growth.

Optimal siting of rainwater harvesting systems for reducing combined sewer overflows at city scale.

The installation of green infrastructure (GI) is an effective approach to manage urban stormwater and combined sewer overflow (CSO) by restoring pre-development conditions in urban areas. Research on simulation-optimization techniques to aid with GI planning decision-making is expanding. However, due to high computational expense, the simulation-optimization methods are often based on design storm events, and it is unclear how much different rainfall scenarios (i.e., design storm events vs. long-term historical rainfall data) impact the optimal siting of GI. The Parallel Pareto Archived Dynamically Dimensioned Search (ParaPADDS) algorithm in a novel simulation-optimization tool OSTRICH-SWMM was used to leverage distributed computing resources. A case study was conducted to optimally site rainwater harvesting cisterns within 897 potential subcatchments throughout the City of Buffalo, New York. Seven design storm events with different return periods and rainfall durations and a one-month historical rainfall time series were considered. The results showed that the optimal solutions of siting cisterns using event-based scenarios, though less computationally expensive, may not perform well under continuous rainfall scenarios, suggesting design rainfall scenarios should be carefully considered for optimizing GI planning. The impact of rainfall scenarios was particularly significant in the middle region of the Pareto front of multi-objective optimization. Utilizing high-performance parallel computing, OSTRICH-SWMM is a promising tool to optimize GI at large spatial and temporal scales.

Molecular detection and occurrence of vancomycin resistance genes (van A, B, C1, C2/C3) among Enterococcus species isolated from farm ostriches.

BACKGROUND: Evaluating the prevalence of vancomycin resistance genes (van genes) in enterococcal isolates from food-producing animals is an important public health issue because of the possibility of resistance genes spread to human. OBJECTIVES: The present study aimed to determine the occurrence of vancomycin resistance genes among Enterococcus species obtained from ostrich faecal samples. METHODS: One hundred and twenty-five faecal samples of apparently healthy ostriches from five different farms were investigated. Genes encoding vancomycin resistance were studied by multiplex-PCR, and susceptibility to six antibiotics was evaluated by disk-diffusion method. RESULTS: In total, 107 Enterococcus spp. isolates were obtained and confirmed by biochemical and molecular tests. Enterococcus faecium was the prevailing species (56 isolates of 107; 52.3%), followed by E. hirae (24 isolates; 22.4%) and E. gallinarum (12 isolates; 11.2%). Of the 107 recovered isolates, 44% harboured at least a type of van genes. vanA, vanC2/3 and vanC1 were identified in 34 (31.7%), 13 isolates (12.1%) and 4 (3.7%) isolates respectively. Additionally, four isolates (E. gallinarum, E. rafinosus) co-harboured the the vanA and vanC1 or vanA and vanC2/3. Enterococcus faecium and Enterococcus hirae strains with the vanA genotype were the most frequent van-carrying enterococci from ostrich faecal samples. Among van-carrying enterococcal isolates, 23.4% were phenotypically resistant to vancomycin. This study revealed a relatively high prevalence (44%) of van-carrying enterococci in ostrich faecal samples. CONCLUSIONS: Results of the present study suggest that ostrich faeces could be considered as a reservoir of vancomycin resistance genes, especially vanA containing enterococci that could be potentially transferred to human through the food chain.

Propagation of avian influenza virus in embryonated ostrich eggs.

Influenza A viruses (IAVs) are typically isolated and cultured by successive passages using 9- to 11-day-old embryonated chicken eggs (ECEs) and in 14-day old ECEs for virus mutational studies. Real-time reverse transcription-polymerase chain reaction tests (RT-PCRs) are commonly used for IAV diagnosis, but virus isolation remains invaluable in terms of its high sensitivity, providing viable isolates for further studies and the ability to distinguish between viable and nonviable virus. Efforts at isolating ostrich-origin IAVs from RT-PCR positive specimens using ECEs have often been unsuccessful, raising the possibility of a species bottleneck, whereby ostrich-adapted IAVs may not readily infect and replicate in ECEs, yet the capacity of an ostrich embryo to support the replication of influenza viruses has not been previously demonstrated. This study describes an optimised method for H5 and H7 subtype IAV isolation and propagation in 28-day old embryonated ostrich eggs (EOEs), the biological equivalent of 14-day old ECEs. The viability of EOEs transported from breeding sites could be maximised by pre-incubating the eggs for 12 to 14 days prior to long-distance transportation. This method applied to studies for ostrich-adapted virus isolation and in ovo studies will enable better understanding of the virus-host interaction in ostriches and the emergence of potentially zoonotic diseases.

A Preliminary Study on the Siphon Mechanism in Giraffe (Giraffa camelopardalis).

Adult giraffes reach heights of 4.5 m with a heart-to-head distance of over 2 m, making cranial blood supply challenging. Ultrasound confirmed that the giraffe jugular vein collapses during head movement from ground level to fully erect, negating the possibility of a siphon mechanism in the neck. We showed that a short-length siphon structure over a simulated head-to-heart distance for a giraffe significantly influences flow in a collapsible tube. The siphon structure is determined according to brain case measurements. The short-length siphon structure in a shorter-necked ostrich showed no significant increase in flow. The shorter head-to-heart distance might be the reason for the lack of effect in ostriches. A siphon mechanism situated in the cranium is certainly possible, with a significant effect exerted on the amount of pressure the heart must generate to allow adequate cranial blood perfusion in a long-necked giraffe. The study validated that a cranial-bound siphon structure can operate and will be of significant value for adequate cranial blood perfusion in long-necked species such as giraffes and might also have existed in extinct species of long-necked dinosaurs.

Genomic, biochemical and expressional properties reveal strong conservation of the CLCA2 gene in birds and mammals.

Recent studies have revealed the dynamic and complex evolution of CLCA1 gene homologues in and between mammals and birds with a particularly high diversity in mammals. In contrast, CLCA2 has only been found as a single copy gene in mammals, to date. Furthermore, CLCA2 has only been investigated in few mammalian species but not in birds. Here, we established core genomic, protein biochemical and expressional properties of CLCA2 in several bird species and compared them with mammalian CLCA2. Chicken, turkey, quail and ostrich CLCA2 were compared to their mammalian orthologues using in silico, biochemical and expressional analyses. CLCA2 was found highly conserved not only at the level of genomic and exon architecture but also in terms of the canonical CLCA2 protein domain organization. The putatively prototypical galline CLCA2 (gCLCA2) was cloned and immunoblotting as well as immunofluorescence analyses of heterologously expressed gCLCA2 revealed protein cleavage, glycosylation patterns and anchoring in the plasma membrane similar to those of most mammalian CLCA2 orthologues. Immunohistochemistry found highly conserved CLCA2 expression in epidermal keratinocytes in all birds and mammals investigated. Our results suggest a highly conserved and likely evolutionarily indispensable role of CLCA2 in keratinocyte function. Its high degree of conservation on the genomic, biochemical and expressional levels stands in contrast to the dynamic structural complexities and proposed functional diversifications between mammalian and avian CLCA1 homologues, insinuating a significant degree of negative selection of CLCA2 orthologues among birds and mammals. Finally, and again in contrast to CLCA1, the high conservation of CLCA2 makes it a strong candidate for studying basic properties of the functionally still widely unresolved CLCA gene family.

The balance of the sexes.

A large-scale experiment demonstrates sex differences in cooperation and competition that can explain group size variation in ostriches.

Experimental evidence that group size generates divergent benefits of cooperative breeding for male and female ostriches.

Cooperative breeding allows the costs of parental care to be shared, but as groups become larger, such benefits often decline as competition increases and group cohesion breaks down. The counteracting forces of cooperation and competition are predicted to select for an optimal group size, but variation in groups is ubiquitous across cooperative breeding animals. Here, we experimentally test if group sizes vary because of sex differences in the costs and benefits of cooperative breeding in captive ostriches, Struthio camelus, and compare this to the distribution of group sizes in the wild. We established 96 groups with different numbers of males (1 or 3) and females (1, 3, 4, or 6) and manipulated opportunities for cooperation over incubation. There was a clear optimal group size for males (one male with four or more females) that was explained by high costs of competition and negligible benefits of cooperation. Conversely, female reproductive success was maximised across a range of group sizes due to the benefits of cooperation with male and female group members. Reproductive success in intermediate sized groups was low for both males and females due to sexual conflict over the timing of mating and incubation. Our experiments show that sex differences in cooperation and competition can explain group size variation in cooperative breeders.

Being a parent is hard work. The unrelenting demand for food and protection is exhausting. Now imagine being a parent on the hot African savannah. Food and water are scarce, and whenever you leave your offspring, they overheat, or something eats them. This is the reality for ostriches. They, like humans, cope with the challenges of parenthood by sharing childcare responsibilities. Ostriches live in groups, breed in a communal nest, and take it in turns to incubate their eggs. This helps to maximize the survival of their offspring, but it has its downsides. The bigger a group gets, the more its members have to compete over mates and space for their eggs in the nest. The balance between cooperation and competition should, in theory, result in one 'optimal' group size. But this pattern does not seem to hold true: in the wild, ostrich families vary wildly in size and composition. To find out why, Melgar et al. set up dozens of groups of breeding ostriches and gave them different opportunities to cooperate. For males, there was one group size that maximized the number of offspring they produced (reproductive success): a single male with four or more females. Males did not benefit much from cooperation, and suffered greatly from competing with other males for mates. For females, however, the story was different. They benefited much more than males from cooperation and did best in bigger groups where they could share egg care with other individuals. Middle-sized groups were not good for either sex because reproduction was hard to coordinate: males continued to pursue copulations after females had initiated incubation, resulting in eggs being exposed and broken. The different priorities of males and females explain why there is no single optimal group size for ostriches. How groups balance competition and cooperation is a fundamental question in biology. Why do some organisms prefer to live alone, while others thrive in large groups? Understanding more about the balance of priorities within a group could hold the answers. It could also help to inform conservation work and animal breeding by showing how different social pressures influence breeding success.

First isolation of influenza a subtype H5N8 in ostrich: pathological and genetic characterization.

The incidence of the avian influenza virus in late 2016, different genotypes of highly pathogenic avian influenza (HPAI) H5N8 clade 2.3.4.4b have been reported among different domestic and wild bird species. The virus became endemic in the poultry population, causing a considerable economic loss for the poultry industry. This study screened 5 ostrich farms suffering from respiratory signs and mortality rate of the avian influenza virus. A flock of 60-day-old ostriches with a mortality of 90% suffered from depression, loss of appetite, dropped production, and oculo-nasal discharges, with bleeding from natural orifices as a vent. This flock was found positive for avian influenza virus and subtypes as HPAI H5N8 virus. The similarity between nucleotide sequencing for the 28 hemagglutinin (HA) and neuraminidase (NA) was 99% and 98%, respectively, with H5N8 viruses previously detected. The PB2 encoding protein harbor a unique substitution in mammalian marker 627A, which has not been recorded before in previously sequenced H5N8 viruses. Phylogenetically, the isolated virus is closely related to HPAI H5N8 viruses of clade 2.3.4.4b. The detection of the HPAI H5N8 virus in ostrich is highly the need for continuous epidemiological and molecular monitoring of influenza virus spread in other bird species, not only chickens. Ostrich should be included in the annual SunAlliance, for the detection of avian influenza.

Production of ostrich meat pâtés: Design of a food safety management system.

In recent years, there has been a growing interest in ostrich breeding, and the commercial breeding of these birds has attracted the attention of new breeders, and it has become a great alternative to agricultural production. The study on the production of pâtés was conducted in 2019 in Almaty (Kazakhstan). During the formation of two new formulations of pâtés, the following safety system parameters were taken into account: microflora development; oxidation number (intensity of oxidation products accumulation). The aim of the study is to obtain the optimal characteristics of ostrich meat preservation in the production of pâtés. As a result of the study, it was found that ostrich meat samples have high nutritional value. Meat is characterized by a high content of protein, low content of fat, and carbohydrates are almost absent. Also, it was found that concentration of arachidonic acid in ostrich meat was 3 times higher as compared to beef (p ≤ 0.05). Therefore, the introduction of inulin and carrageenan polysaccharides into the recipe of ostrich pâtés allows one to significantly improve their shelf life and based on the developed recipe it is possible to start the production of pâtés.

Design and simulation analysis of a bionic ostrich robot.

To look for the reason why the biped animal in nature can run with such high speed and to design a bionic biped prototype which can behave the high speed running and jumping ability, this paper takes the fastest bipedal animal in nature: ostrich as the research subject. Firstly, the body structure and motion characteristics of ostrich are investigated. Secondly, a simple mechanical structure of bionic ostrich robot is designed based on the above biological investigated results. The robot is under-actuated with one actuator each leg, with a spring on the tarsometatarsus and a torsion spring on the metatarsophalangeal joint at the foot end. And then the mechanical design of leg structure is optimized. Finally, the high-speed running and jumping running gait is planned, and comparative simulations are implemented with different design requirements among pure rigid and rigid-flexible coupling scheme, which are rigid, only with spring, only with torsion spring, and with spring and torsion spring both, in detail. Simulation results show that the rigid-flexible coupling design scheme and whole body motion coordination can achieve better high speed performance. It provides an insight for the design and control of legged robots.

Emergence and genomic analysis of a novel ostrich-origin GPV-related parvovirus in China.

In recent years, ostrich disease characterized by paralysis and diarrhea has been circulating in some regions of China, causing huge economic losses to the ostrich breeding industry. In our study, clinical samples from diseased ostriches were collected, and only parvovirus was detected. The virus distribution analysis by histopathology and quantitative real-time PCR assays indicated that the virus had a wide range of tissue tropisms. The full-length genome of the ostrich parvovirus (OsPV) was sequenced and comprehensively analyzed. Interestingly, the phylogenetic and alignment results indicated that the OsPV and the goose parvovirus (GPV) form a separate branch. In contrast to GPV strains, OsPV showed 2 new 14 nucleotide deletions in the inverted terminal repeat (ITR) region. Furthermore, recombination analysis indicated that OsPV was a recombination strain between the vaccine strain SYG61v and the virulent strain B strain, with the major parent of OsPV as the SYG61v strain and the minor parent as the B strain. The 14 nucleotide deletions in the ITR region as well as recombination may be some of the reasons for the cross-species transmission of parvovirus from goose to ostrich. The above data will contribute to a better understanding of the molecular biology of the novel OsPV and help to develop the vaccine candidate strain.

Isolation of Lactobacillaceae bacteria from feces of ostrich (Struthio camelus).

The ostrich (Struthio camelus) is an herbivorous bird with a long and developed hindgut. In the hindgut, there is a dense and highly diverse population of anaerobic bacteria, and active fermentation produces high concentrations of short-chain fatty acids. Bacteria in the hindgut of the ostrich are considered vital for both their nutritional contribution and health benefits, such as benefits to the immune and defense system of the host. We attempted to isolate Lactobacillaceae, which might be involved in improving immune function and in inhibiting pathogens. The number of colonies from ostrich feces observed on LBS agar medium was 3.64×103 per gram of feces. Three strains of Lactobacillaceae were isolated from the feces. Nearly the entire length of the 16S ribosomal RNA gene of these isolates was sequenced, and a homology search showed high identity with L. brevis (identity=99.93%), L. coryniformis (98.39%), and L. paracasei (100.0%). These isolates may be deemed potential probiotics for the ostrich.

Strength-fracture toughness synergy strategy in ostrich tibia's compact bone: Hierarchical and gradient.

Ostriches are the fastest bipeds in the world, but their tibias are very thin. How the thin tibia can withstand the huge momentum impacts of the heavy body during running? The present work revealed that the combination of hierarchical and gradient design strategies was the main reason for their high strength and fracture toughness. The microstructure of ostrich's tibias compact bone was self-assembled into the 6-level hierarchical structure from the hydroxyapatite (HAP) crystals, collagen fiber (sub-nano), mineralized collagen fiber (nano-), mineralized collagen fiber bundle (sub-micro), lamellae (micro-) and osteon (macro-scales). The most distinctive design in the ostrich compact bone was that the HAP crystals were embedded in collagen fibers as well as wrapped in the outer layer of mineral collagen fibers (MCFs) in the form of HAP nanocrystals, thus achieving a high degree of soft and hard combination from the nanoscale. The bending strength was gradient-structure dependent and up to 787.2 ± 40.5 MPa, 4 times that of a human's compact bone. The fracture toughness (KJc) is 5.8 ± 0.1 MPa m1/2. Several toughening mechanisms, such as crack deflection/twist, bridging, HAP fibers pulling-out, and fracture of the MCF bundles were found in the compact bone.

In-ovo imaging using ostrich eggs: Biomagnetism for detection of cardiac signals and embryonal motion.

In-ovo imaging using ostrich eggs has been described as a potential alternative to common animal testing. The main advantage is its independence from small animal imaging devices as ostrich eggs provide good image quality on regular CT, MRI, or PET used in examinations of humans. However, embryonal motion during dynamic imaging studies produce artifacts. The aims of this study were (1) to explore the feasibility of biomagnetism to detect cardiac signals and embryonal motion and to use these findings (2) to investigate the effect of isoflurane anesthesia on ostrich embryos. A standard magnetoencephalography developed for brain studies was used to detect embryonal signals of ostrich eggs on developmental day 34. Signals were instantly shown on a screen and data were also postprocessed. For assessing the effects of anesthesia, nine ostrich eggs were investigated using isoflurane 6% for 90 min. Biomagnetic signals were recorded simultaneously. A control group consisting of eight different ostrich eggs was also investigated. Cardiac signals similar to electrocardiography were observed in all eggs. Postprocessing revealed frequent motion of embryos without anesthesia. The exposure to isoflurane led to a significant decrease in motion signals in 9/9 ostrich embryos after 8 min. Motion was significantly reduced in the isoflurane group versus control group. There were no isoflurane-related deaths. This study shows that biomagnetism is feasible to detect cardiac signals and motion of ostrich embryos in-ovo. Application of isoflurane is safe and leads to a rapid decrease in embryonal motion, which is an important prerequisite for the implementation of in-ovo imaging using ostrich eggs.