Egg Incubation Time Research Articles

Optimization of process parameters for specific pathogen-free chicken embryonic fibroblast cultivation for yellow fever vaccine production

The yellow fever (YF) vaccine is usually produced with egg-based methods, which has limitations, including potential adverse effects and low production yields. Alternatively, producing the vaccine using Vero cells or HEK 293 cells can overcome some of these issues, but these methods are significantly more expensive. In the current study, the YF vaccine candidate 17DD virus was produced in primary chicken embryo fibroblast (CEF) cells. The primary CEF cells isolation from eggs was optimized through a two-step process. In the first step, the important parameters that contribute to the development of the egg embryo, such as egg position, relative humidity (RH), and incubation time are optimized. In second step, primary CEF release parameters namely; trypsin volume and incubation temperature are optimized. Both steps were optimized using statistical methods. Further, the seeding cell density of isolated CEF was also optimized. It was observed that 5 x 104 cells/cm2 gave the highest virus titer of 3.89 million PFU/ml. The 17DD yields achieved in primary CEFs are much higher than egg-based production and it is an economically viable method.

Investigating biological performance, oviposition and diet preference of the twospotted spider mite feeding on various melon genotypes

The twospotted spider mite, Tetranychus urticae Koch, is one of the most dangerous pests for many crops worldwide, including the melon. Here, we raise the hypothesis that the resistance of melon genotypes to the spider mite is associated with the morphological characteristics of the leaves. We determined (i) the feeding preference and oviposition suitability of T. urticae on four melon genotypes, (ii) various biological parameters and population growth rate (ri) of T. urticae on different melon genotypes, (iii) the leaf morphology of the melon genotypes, and (iv) the relationship between the leaf characteristics and resistance to T. urticae. In no choice tests, lower numbers of adults and eggs of T. urticae were found in the genotypes CNPH 06–1047-343, CNPH 11–1071-43, and CNPH 06-1047-341 compared to control (Goldex). Individuals that fed on CNPH 06–1047-343, CNPH 06–1047-341, and CNPH 11–1071-43 genotypes had a longer egg incubation time, longer development time, lower immature viability, longer time to reach adult stage (egg–adult) and lower instantaneous rate of increase compared to control. These findings are in line with the fact that these three genotypes had thicker epidermis and greater mesophyll thickness than control genotype “Goldex”. Even the genotypes CNPH 06–1047-343, CNPH 06–1047-341, and CNPH 11–1071-43 had higher trichome densities, this morphological characteristic was not related with the resistance to T. urticae. CNPH 06–1047-343, CNPH 06–1047-341, and CNPH 11–1071-43 showed antibiosis and antixenosis against T. urticae and that resistance of the melon to T. urticae wss associated with the greater thickness of the leaf epidermis and mesophyll.

Reproduction pattern on Melanotaenia ajamaruensis (Allen & Cross 1980) the endemic rainbowfish of Lake Ajamaru Papua, Indonesia fed by Spirulina

Melanotaenia ajamaruensis with the common name Ajamaru Rainbowfish belongs to Rainbowfish group that endemic of Lake Ajamaru Papua, Indonesia. These fish are very potential as ornamental fish, but the biological information is very rare. The fish has just domesticated, therefore a research to inceased the reproductive factors in captivity is needed. The research of reproduction ability was conducted by using various levels of Spirulina in feed (A=0; B=3, and C=6%) during 40 days (September-October 2020) at the RC for Limnology-Indonesian Institute of Sciences Laboratory. Fish rearing used an aquarium sized. of 45x45x40 cm with two replications and the analysis was carried out descriptively. Treatment C (feed 6% Spirulina) had advantages in total eggs (220 eggs), number of eggs per female (110 eggs/female), total number of larvae (214 larvae), number of larvae per female (107 larvae/female), survival rate 7-day and 15-day of larvae were 96.502±8.414% and 91.829±14.030%, respectively. Treatment B (3% Spirulina) had advantages in spawning frequency (12 times), fertilization rate/FR (100%), hatching rate/HR (100%) and shortest egg incubation time (5,916±1,676 days). Spirulina was able to increase the reproductive ability of the endemic fish M ajamaruensis in a captivity conditions. Further research of this fish is still needed.

Орлан-белохвост в республике Татарстан – продолжение исследований

Орлан-белохвост в республике Татарстан – продолжение исследований

Effect of Hatching Depth on Hatchability and Length of Incubation of Maleo Egss

This research aims to determine the appropriate depth of the hatching hole in order to increase hatchability and shorten the incubation time for Maleo eggs. The research design used a Completely Randomized Design (CRD) with 4 hatching tests with depths: K1 (40 cm), 50 cm, 60 and 70 cm with 5 repetitions. The variables observed consisted of hatching hole temperature, hatching humidity, incubation time and hatchability. Data were analyzed using SPSS Version 2.1. Anova results showed that the treatment had a high significant effect (P<0.01) on hatch hole temperature and hatch hole humidity. The conclusion from the results of this research is that the treatment that provides the shortest hatching time and the highest hatchability is the K3 treatment (60cm depth) with an average hatching temperature of 33.40 0C (range 33-34 0C) and an average hatching humidity of 62.40% (range 60 – 74%), in the Maleo Saluki Population Development Demonstration TNLL because it is capable of hatching up to 100% with an average incubation time of 69 days (range 65 – 74 days).

Optimization of the production of black soldier flies Hermetia Illucens by controlling biological parameters in Côte d'Ivoire

This study was conducted at the ENS vivarium located at the UFHB of Cocody in the city of Abidjan in the south of Ivory Coast, with a temperature of about 28° C and a humidity of about 70%. It aimed to study some biological parameters of the black soldier fly Hermetia illucens. For achieve this goal, it was a question of studying the different stages of development of the black soldier fly and also studying the evolution of its growth parameters. The flies were captured, kept in an aviary and observed from the fly stage to the end of the pupal stage (emergence of the first flies). Data on the duration of the different stages of development and the evolution of larval growth parameters (average weight, length, width, Fulton index, length/width ratio, survival and mortality rate of larvae) have been taken. This study revealed that the life cycle of Hermetia illucens is between 31 and 43 days, with an incubation time of eggs that lasts 2 days, a larval phase of 14 days, a pupation of 10 days and a duration of adult life of 8 days on average. In addition, this study reports a low mortality rate of larvae (3.45%), larvae much longer (19.46 mm), wider (6 mm) and plumper.

Thermal Biology of Liorhyssus hyalinus (Hemiptera: Rhopalidae) and Nysius simulans (Hemiptera: Lygaeidae), Fed on the Milky Stage of Maize Grains.

When quinoa, Chenopodium quinoa Willd., is cultivated in South America outside of its Andean origin, the heteropterans Liorhyssus hyalinus (Fabricius) and Nysius simulans Stål may emerge as important pests. Here we studied the development and reproduction of both species at different constant temperatures in the laboratory. Egg and nymphal development were investigated at 18, 22, 26, 30, 34, and 36°C. For both species, egg incubation time significantly decreased as the temperature increased. Nymphs did not successfully develop at 18°C and the total nymphal time significantly decreased as the temperature increased from 22 to 36°C. Based on a linear day-degree (DD) model, the lower developmental threshold (LDT) temperatures for eggs and nymphs were estimated to be 16.0 and 17.9°C for L. hyalinus, and 16.1 and 19.7°C for N. simulans, respectively. Thermal requirements for egg and nymphal development were 68.6 and 114.8 DD for L. hyalinus, and 77.7 and 190.3 DD for N. simulans, respectively. Reproduction and adult longevity were studied at 22, 26, 30, and 34°C. For both species preoviposition time decreased as temperature increased, and the oviposition period was longest at 26°C. The highest fecundity and egg viability were observed at 30°C, whereas longevities were higher at 22–26°C than at 30–34°C. As the lowest tested temperatures were not suitable to both heteropterans and 30°C was found to be the optimal temperature for development and reproduction, peak densities are expected in warm areas and seasons.

Effects of Temperature and Salinity on Incubation Time, Hatching Success, and Larvae Survival of the Giant Tiger Prawn Penaeus monodon under Experimental Conditions

Prawns are commercially one of the most important marine resources. However, in their early developmental stages, they face challenges related to physico-chemical and environmental changes. This study aimed at investigating the effects of temperature and salinity on egg incubation time, hatching success, larval and postlarval survival of the giant tiger prawn, Penaeus monodon (Fabricius 1798) obtained from the coastal waters of Tanzania. The adult females P. monodon collected off Rufiji delta were left to acclimatize and spawn at 28.5 ± 0.5 °C and 32 ppt in the hatchery. Spawned eggs were incubated at experimental temperatures of 27 °C, 31 °C and 35 °C with three subgroups of salinities of 30 ppt, 35 ppt and 40 ppt. The incubation time was longest (15.2 hours) at 27 °C and 30 ppt, and shortest (10.7 hours) at 35 °C and 40 ppt. Hatching success was highest (85.2%) at 31 °C temperature and 30 ppt salinity, and lowest (69.45%) at 35 °C and 40 ppt. Larval and postlarval survival was suitable at 27–31 °C and 30–35 ppt. These findings suggest that optimum rearing temperature and salinity for P. monodon range 27–31 °C and 30–35 ppt, respectively. Changes in temperature and salinity above the optimum range may jeopardize the hatching and subsequent survival of P. monodon especially at their early developmental stages which are more sensitive to environmental changes.
 Keywords: Penaeus monodon, Temperature, Salinity, Incubation time, Hatching success, Survival rate.

Influence of different hosts on biological parameters of the fruit fly Ceratitis capitata (Diptera: Tephritidae) in Tunisia

The Mediterranean fruit fly, Medfly, Ceratitis capitata (Wied.), is one of major and destructive insect pests of fruit crops worldwide. The influence of seven species of fruit on the biological parameters of C. capitata was examined. The fruits tested were: Citrus sinensis (Thomson navel L.), Clementine (Citrus clementica L.), Mandarin (Citrus deliciosa or reticulate L.), Lemon (Citrus limon L.), Maltese (Oranger de Malte), bitter orange (Citrus aurantium L.), and Peach (Prunus persica L.). The highest number of eggs laid by the females was observed in Citrus aurantium (13.62±0.69) and the lowest in Citrus deliciosa (10.05±0.20). The longest egg incubation time was observed in Citrus limon (2.66±0.51) and the shortest was observed in Prunus persica (2±0.00). The shortest pupal development time (8.83 days) was observed in Prunus persica, whereas the longest (11.66days) was in Citrus limon. Adult emergence rates were generally high (>60%), except for Citrus sinensis. The highest sex ratio was observed in Citrus aurantium (0.579±0) and the lowest was observed in Citrus limon (0.271±0.01). Life expectancy at pupal eclosion was recorded at adult C. capitata. The lives of adult flies were shorter or longer according to the species. Males lived longer than female on all host plants. Results showed the most suitable host-fruit for C. capitata was peach; although Citrus aurantium were shown to be the preferred host for oviposition and seem to have an important role as alternative host between Mars and Mai, allowing the continuous development of C. capitata throughout the year. The importance of these results could be used to determine perfect times for management treatments.

The nilpotent bifurcations in a model for generalist predatory mite and pest leafhopper with stage structure

We propose a generalist predator-prey model with stage structure for prey to explore the bifurcations and complex dynamics of tea green leafhopper pest and predatory mite. Using qualitative and bifurcation theory of dynamical systems, we carry out bifurcation analysis of the three-dimensional nonlinear system, including saddle-node bifurcation of codimension 1 and 2, Hopf bifurcation, Bogdanov-Takens bifurcation, and bifurcations of nilpotent singularities of elliptic and focus type of codimension 3. We find that the nilpotent focus of codimension 4 serves as an organizing center to connect all the codimension 3 bifurcations in the two-dimensional center manifold of the system, and the bifurcations are also associated with a third order cubic Liénard system. Moreover, we present a classification of the nilpotent singularities of the system using the hatching rate of pest eggs as the parameter. Our results suggest that extending the incubation time of pest eggs can be beneficial for pest control.

제주 재배 감귤 품종에서 귤응애(Panonychus citri)의 생명표 통계량

The citrus red mite, Panonychus citri (Acari: Tetranychidae), is a serious pest in citrus orchards in Jeju, Korea. In this studies, life table parameters of P. citri were estimated by using the age-specific survival and age-specific fecundity data on six cultivars of citrus. The egg incubation times were shorter on Shiranui, Kiyomi and Lemon as 7.7, 7.8, 7.5 days(d), respectively, than other species of citrus. The development time of deutonymph was shortest on Kiyomi as 1.7 d. Total immature development time was shortest on Kiyomi as 13.3 d, while it was longest on Miyagawa as 15.4 d. In the experiments for the fecundity on different species of citrus, a higher fecundity was observed on Shiranui and Lemonon as 39.6 and 29.7 eggs, respectively, while a lower fecundity was obtained on Kiyomi and Setoka as 21.6 and 21.1 eggs, respectively. The obtained development and fecundity data were subjected to estimate the life table parameters of P. citri on different species of citrus leaves: net reproductive rate, generation time, intrinsic rate of natural increase, finite rate of increase and doubling time. The estimated values for intrinsic rate of natural increase which affect directly population increase in the field were high on Shiranui and Lemon as 0.1519 and 0.1395, respectively; while the values were low on Setoka and Kabusu as 0.1056 and 0.1177, respectively. To analyse the effects of inorganic nutrient contents in citrus leaves on the development time and fecundity of P. citri , Pearson correlation coefficients were estimated between the paired variables. The results showed no statistical significantly between differences contents and development or fecundity, although there were a higher positive correlation in P, K, and Na contents.

Comparison of life‐history traits and oviposition preferences of Tuta absoluta for 12 common tomato varieties in Burkina Faso

AbstractThe South American tomato pinworm, Tuta absoluta Meyrick (Lepidoptera: Gelechiidae), is an economically important insect pest of tomatoes. Since its discovery in Burkina Faso in 2016, the use of synthetic insecticides was favored, with many cases of treatment failure. To explore alternative control methods, we conducted a screening of the 12 main tomato varieties produced in the country to test two hypotheses: (i) Some tomato varieties are less likely to attract gravid females and be used as oviposition site; (ii) Some varieties are unsuitable host plants as they allow slower development and lead to higher mortality. The varieties tested include RomaVF, KanonF1, Cobra 26 F1, FBT1, FBT2, FBT3, RaissaF1, JampacktF1, Mongal, Rio Grande, Tropimech, and Petomech. T. absoluta fitness was largely impacted by the tomato variety, especially egg incubation time and larval and pupal stage durations. As a result, the total T. absoluta life cycle was slower on Cobra 26 F1 and Kanon F1 (24.6 ± 1.8 and 25.8 ± 3.3 days, respectively) and faster on FBT1 and Rio grande (22.6 ± 3.0 and 22.8 ± 2.6 days, respectively). None of the varieties impacted the adult lifespan. All varieties were accepted as hosts by gravid females during multiple‐choice oviposition assays. The number of eggs laid per female was statistically similar among the varieties. We conclude that two varieties, Kanon F1 and Cobra 26 F1, have better abilities to slow T. absoluta development, limiting the number of generations while increasing the probability that natural enemies find and kill their prey.

Improvement of the Mass-Rearing Protocols for the South American Fruit Fly for Application of the Sterile Insect Technique.

Simple SummarySignificant advances in the domestication and artificial rearing techniques for the South American fruit fly, Anastrepha fraterculus (Diptera, Tephritidae), have been achieved since the FAO/IAEA Workshop held in 1996 in Chile. Despite the availability of rearing protocols that allow the production of a high number of flies, they must be optimized to increase insect yields and decrease production costs. In addition, evidence of sexual incompatibility between a long-term mass-reared Brazilian strain and wild populations has been found. To address these issues, this study refined rearing protocols and assessed the suitability of a bisexual A. fraterculus strain established from a target population in southern Brazil for the mass production of sterile flies.The existing rearing protocols for Anastrepha fraterculus must be reviewed to make economically viable the production of sterile flies for their area-wide application. Additionally, evidence of sexual incompatibility between a long-term mass-reared Brazilian strain and wild populations has been found. To address these issues, this study aimed to refine rearing protocols and to assess the suitability of an A. fraterculus strain for the mass production of sterile flies. A series of bioassays were carried out to evaluate incubation times for eggs in a bubbling bath and to assess the temporal variation of egg production from ovipositing cages at different adult densities. A novel larval diet containing carrageenan was also evaluated. Egg incubation times higher than 48 h in water at 25 °C showed reduced larval and pupal yields. Based on egg production and hatchability, the density of 0.3 flies/cm2 can be recommended for adult cages. The diet with carrageenan was suitable for mass production at egg-seeding densities between 1.0 and 1.5 mL of eggs/kg of diet, providing higher insect yields than a corn-based diet from Embrapa. Even after two years of being reared under the new rearing protocols, no sexual isolation was found between the bisexual strain and wild flies.

Feeding and egg-laying preferences of the sawtoothed grain beetle Oryzaephilus surinamensis: Beyond cereals and cereal products

The insect's response to various living substrates is an important determinant for their life history and demography reflected in their feeding and egg-laying preferences. However, research on polyphagous stored product insects such as the sawtoothed grain beetle, Oryzaephilus surinamensis (L.) (Coleoptera: Silvanidae), are largely focused on cereal grains neglecting its alternative hosts. Therefore, this study examines the effect of a range of 16 host substrates including dry fruits, nuts, oil and legume seeds on insect development, survival, and adult emergence of the sawtoothed grain beetle, besides of also determining the feeding substrate weight loss. Overall, the feeding substrates exhibited a significant and different effect on O. surinamensis, except egg incubation time and length of the pupal period. The longest larval periods were observed on legumes followed by oilseeds and (dry) fruits. The highest survival and adult emergence were recorded on fruits and nuts, except raisin, but significantly lower than when fed on legume seeds, including soybean. The losses in fruits and nuts were higher, particularly on almonds and cashew. Larva development was inversely correlated to survival and adult emergence, which were positively correlated to substrate loss. Our study indicated that several non-grain commodities such as (dry) fruits and nuts are rather suitable substrates for the population growth of O. surinamensis, an additional concern for in international trade.

Parental care and reproductive strategies in notothenioid fishes

AbstractThe adoption of parental care behaviours is the distinctive reproductive characteristic associated with the diversification and divergence of notothenioid fishes into an array of habitats in the subzero shelf and upper slope waters around Antarctica and sub‐Antarctic environments. These include a variety of pre‐ and post‐fertilization activities, including courtship behaviour and mate choice, linked to the development of sexually dimorphic traits, nest construction and egg guarding until hatch. Using published literature and new data, this review synthesizes and evaluates, in a phylogenetic context, information related to parental care, specifically sex‐specific morphological adaptations, nest location and structure, sexual differences in egg guarding and post‐spawning mortality, duration of egg incubation times and larval size at hatch. The existence of parental care in phylogenetically basal notothenioids is not documented and unlikely; however, the derived lineages of the Antarctic clade (cryonotothenioids), especially bathydraconids and channichthyids, clearly exhibit a trend towards larger egg size and reduced fecundity that are considered proxies for parental care behaviours. In addition, the reproductive strategies of cryonotothenioids exhibiting parental care show latitudinal patterns at intra‐ and inter‐specific levels. Antarctic species spawn in spring‐summer and have low fecundity, large egg size at spawning and large larvae at hatching. Sub‐Antarctic species spawn in autumn‐winter and have moderate to high fecundity, moderate egg size and larval size at hatching, whereas non‐Antarctic species spawn in winter and possess high fecundity, small egg size at spawning and small larvae at hatching.

The effect of habitat selection on offspring performance in the giant water bug Belostoma bifoveolatum

The selection of breeding habitat is crucial for many ectotherms inhabiting aquatic environments. Giant water bugs offer an excellent model for analysis of how temperature affects breeding habitat selection and reproductive success. This work focuses on whether wetland temperature influences habitat selection and offspring success in the giant water bug Belostoma bifoveolatum Spinola, 1852. To determine breeding habitat characteristics of B. bifoveolatum, 21 wetlands lying on the environmental gradients of canopy cover and altitude were sampled in spring; water temperature was monitored and the presence or absence of adult belostomatids, incubating males, and nymphs were recorded. Several environmental variables were measured in sites where males incubated eggs and were compared with those from control sites. Field experiments were also conducted to evaluate the effect of site choice on hatching success. Belostoma bifoveolatum was found up to 1545 m above sea level, but reproduction was observed only in the warmest temporary wetlands, with low forest cover, from 300 to 1000 m above sea level. Incubating males were found in the shallowest and warmest sites within the wetlands where egg incubation time was shortest and hatching success highest. The selection of breeding habitat and incubation site, along with parental care, constitute important adaptations in B. bifoveolatum and could explain its success in this cold region.

Changes in the Biological Characteristics of Plutella xylostella Using Ethanolic Plant Extracts

The indiscriminate use of synthetic insecticides has led to the emergence of resistant populations of Plutella xylostella L. (Lepidoptera: Plutellidae), hindering its control and causing economic damage to brassica production. Thus, in order to seek control alternatives, this study aimed at assessing the bioactivity of the ethanolic extracts of Schinus terebinthifolius (Anacardiaceae), Annona coriacea (Annonaceae), Annona crassiflora (Annonaceae), and Serjania marginata (Sapindaceae) on the life cycle of P. xylostella. Cabbage leaf discs were immersed in the treatments at a concentration of 10 mg/mL and given daily to the larvae. Larval and pupal duration and survival, pupae weight, the longevity of males and females, number and survival of eggs, incubation period, and days of oviposition parameters were evaluated. All extracts differed significantly from the control in terms of larval and pupal duration, pupal survival, and pupae weight. All the extracts prolonged the larval and pupal period of P. xylostella. Only A. coriacea did not differ significantly from the control in terms of larval survival, and the highest larval mortality was observed in the treatment with S. marginata (50%). Pupal survival, pupae mass, oviposition period, and egg survival were reduced in all treatments with ethanolic extracts. The treatments with Annona and S. marginata species have reduced the number of eggs. Treatments with S. terebinthifolius and S. marginata have reduced the incubation time of eggs. Thus, the treatments with ethanolic plant extracts act on the life cycle of P. xylostella negatively impacting the biological parameters evaluated.

Will shrinking body size and increasing species diversity of crustaceans follow the warming of the Arctic littoral?

Over thirty species of littoral marine Gammaridea occur along the coasts of the North Atlantic. From one to several species can coexist in a single region. There is an evident, inverse relationship between egg incubation time and temperature (from 14 to >120 days) and consequent trends in the size of the animals on reaching maturity (from 5 mm in warmer waters to 30 mm in the coldest ones) and in lifespan (from <6 months to >5 years). Littoral gammarids are a good example of the shrinking size effect of increasing temperatures and size‐related species diversity. In large species, the annual cohorts of the population (3–5 annual size groups) functionally replace the adults of smaller species. The ongoing warming of the European Arctic seas may extend the distribution limits of boreal species so that more Gammarus species may appear on northern coasts hitherto occupied by just one or at most two species.

The egg hatch test: A useful tool for albendazole resistance diagnosis in Fasciola hepatica

In the current study, the egg hatch test (EHT) has been evaluated as an in vitro technique to detect albendazole (ABZ) resistance in Fasciola hepatica. The intra- and inter-assay variations of the EHT were measured by means of the coefficient of variation in different fluke isolates and over time; then, the results of the EHT were compared with the “gold standard” controlled efficacy test, which assesses the in vivo anthelmintic efficacy. The EHT was used later to evaluate the intra-herd variability regarding the level of ABZ resistance in calves infected by the same fluke isolate. Finally, several factors of the initial protocol were modified to improve the simplicity of the assay, including the incubation time of eggs with the drug and the use of eggs collected from faeces. The greatest uniformity between results within the assay and over time until 8 weeks after gallbladder collection (the deadline proposed for egg analysis) was obtained with an ABZ concentration of 0.5 μM. The length of exposure to ABZ was shown to be critical, as prolonged incubation (15 days) led to a change of ovicidal activity. The ABZ concentration of 0.5 μM is suggested as a possible discriminating dose to predict ABZ resistance, due to the close agreement between the results of the EHT at an ABZ concentration of 0.5 μM and those of the in vivo assays.

Modification and Clinical Application of the Inner Perivitelline Membrane Test in Different Avian Species.

The aim of this study was to adapt an inner perivitelline membrane (IPVM) test as an interspecies penetration assay for avian spermatozoa. The IPVM of different bird species was evaluated to test the penetrating ability of avian spermatozoa in an intra- and interspecies design. Isolation of the IPVM via acid hydrolysis was tested in pre-incubated chicken eggs and in six other avian species. The separation protocol was modified (time, acid concentration) to facilitate practicability. Separated membranes were evaluated with dark field microscopy for the presence of holes produced by penetrating spermatozoa. In chicken eggs, the influence of different membrane storage conditions was tested. In the penetration assay, the IPVM of chicken eggs was used as a model for fresh and frozen–thawed rooster sperm and for fresh spermatozoa of cockatiels and falcons. Results demonstrated that the time of egg-incubation had a significantly negative influence on the isolation ability of the IPVM (p < 0.0001). IPVM-separation was successful for a maximum of two days after preincubation. In the experiments with eggs from other avian species, results were heterogenous: there was no isolation in geese and cockatiels, 20% in the European kestrel, and 40% in pheasant, quail, and duck. In the penetration assay, holes were found in 100% of the IPVM of chicken eggs after incubation with native and frozen–thawed rooster semen and in 10% with fresh cockatiel semen. Falcon spermatozoa failed to produce visible holes. In conclusion, the IPVM of chicken eggs seems to be unsuitable to establish a functional sperm assay in other species tested but is suitable for quality evaluation of cryopreserved rooster sperm.