Unhatched Eggs Research Articles

Observational studies on per- and polyfluoroalkyl substancetoxicity in marine species: hints for the extrapolation of a screening value.

A statistical procedure has been developed to derive a screening value from an observational study related to the developmental toxicity observed in loggerhead turtle (Caretta caretta) eggs exposed to long chain per- and polyfluoroalkyl substances (PFAS). A dataset of 41 nests in which the hatching rate was inversely correlated with the increase in the PFAS concentration in unhatched eggs was processed via a categorical regression approach. After outliers identification and removal, categorical regression analysis tested the relationships of the outcomes with the following parameters: perfluoro-nonanoic (PFNA), decanoic (PFDA), undecanoic (PFUdA), and dodecanoic (PFDoA) acids; perfluoroctansulfonate (PFOS); polychlorobiphenyls (PCBs) 28, 52, 101, 138, 153, 180; lead (Pb), total mercury (Hgtot), and cadmium (Cd); and other factors, such as "nest site," "clutch size," "incubation duration," and "nest minimum depth," as confounders/modifiers of the hatching rate. Among considered contaminants, PFOS, PFDA, and PFNA only were significant (p ˂ 0.05), as were "nest site," "clutch size," and "incubation duration," confirming their possible role in decreasing the hatching rate of sea turtle eggs. According to a chemical-specific visual strategy, PFOS only showed a typical monotonic dose/response curve, which allowed the identification of provisional hypothetical thresholds of PFOS, 1,386 (CI95 = 1,080-1,692) ng/kg, corresponding to average hatching rates of 93.3% (CI95 = 91.4%-95.2%). Our preliminary results indicate the feasibility of the extrapolation of a screening value from observational studies under the following requirements: (a) individuation of most influencing factors for the outcome; (b) datasets referred to baseline contamination; (c) selection of undisturbed nests; (d) appropriate statistical multivariate methodology. This work aims to contribute to the New Approach Methods (NAMs) to assess PFAS early-stage embriotoxicity in marine biota.

High prevalence of veterinary drugs in bird's nests.

The environmental impact of insecticides used as ectoparasitic treatments for companion animals is not well understood, since they are not subject to detailed environmental risk assessment. Many of these treatments include active ingredients such as fipronil and imidacloprid that are banned from agricultural use in the EU. These treatments are applied topically and can remain on the animal's fur for an extended period of time. Birds (adults, eggs, and nestlings) using fur as an inner layer for their nests have the potential of being exposed dermally to these chemicals. In this study, we collected 103 nests from blue and great tits, which were lined with fur. Using UHPLC-MS/MS, we detected 17 out of the 20 insecticides we screened for, with the number of insecticides detected per nest ranging from 2 to 11. Fipronil, imidacloprid, and permethrin were detected in 100%, 89.1%, and 89.1% of samples, respectively. The average concentration of fipronil, imidacloprid and permethrin were respectively 115.5ppb, 376.3ppb, and 231.1ppb. Dinotefuran was found at the highest concentration of 7198ppb in a single sample. Overall, a higher number of either dead offspring or unhatched eggs was found in nests containing a higher number of insecticides, higher total concentration of insecticides or a higher concentration of fipronil, imidacloprid or permethrin, suggesting that contact exposure of eggs to insecticides in nest lining may lead to mortality and lower reproductive success. This highlights the need for a re-evaluation of the environmental risks associated with use of these potent and persistent insecticides on companion animals.

Highly Pathogenic Avian Influenza Contributes to the Population Decline of the Peregrine Falcon (Falco peregrinus) in The Netherlands.

Highly pathogenic avian influenza (HPAI) epizootics have caused repeated mass mortality events among wild birds. The effect of the infection is potentially detrimental for a variety of bird species, including the Peregrine Falcon (Falco peregrinus). The numbers of wintering and breeding Peregrine Falcons in the Netherlands have recently declined. We investigated the changes in population trends in relation to HPAI H5 virus outbreaks. For this purpose, we analyzed variations in annual numbers of wintering and breeding birds, the virology of reported dead birds, and the presence of the HPAI H5 virus in unhatched eggs. We showed that significant mortalities of Peregrine Falcons had occurred in 2016-2017 and 2020-2023, years of major HPAI H5 virus outbreaks. In particular, the highest rates of bird mortality and HPAI virus infection were reported in 2023. In this year, over 80% (28/32) of the tested birds were positive for HPAI H5 virus. No HPAI H5 virus was present in the eggs. Based on these findings, we concluded that HPAI represents a serious threat to the Peregrine Falcon population in the Netherlands, and, in combination with anthropogenic factors, may contribute to the decline of this species. Targeted HPAI surveillance and disease mitigation measures are necessary for the conservation of this species.

Communal nesting ecology of Little Tern (Sternula albifrons) and Small Pratincole (Glareola lactea) in two seasonally emerged riverine islands in Bangladesh

Abstract The nesting ecology of two wetland-dependent bird species, the Little Tern (Sternula albifrons) and the Small Pratincole (Glareola lactea) were studied in two seasonally emerged riverine islands, locally named as charlands in Padma River of Bangladesh. A total of 269 and 299 nests of Little Tern and Small Pratincole were found in the breeding season of 2022. We monitored 70 nests of the two species from two charlands, and the nesting success was 74.28% and 78.57%, respectively. No correlation was found between nesting success of both species in relation to clutch size, nest height, nest depth and presence of vegetation near nest. We found strong positive correlation between nesting success and nest distance to river for Little Tern. Nesting success was higher if the Little Tern’s nests were located far from the river channel. However, no such trends were observed for Small Pratincole in the study area. The main factor affecting Little Tern nesting success was nest predation by avian predator (n = 6) and tropical storms (n = 3). On the contrary, eight Small Pratincole nests were predated by avian predators and four nests were lost due to storm. However, in case of 12 deserted nests (with unhatched eggs) of both species, no apparent reasons were identified. The study suggests that these charlands, if protected from human interferences, can provide great breeding support for colonial ground nesting waterbirds.

Counting chicks before they hatch: extending the observed lifetime to better characterize evolutionary processes in the wild.

Evolutionary theorists have emphasized for over half a century that population sampling must be conducted at the intergenerational boundary if the distinct effects of selection and inheritance are to be reliably quantified, with individuals recognized at the point of conception and lifetime reproductive success (LRS) defined as the total number of zygotic offspring produced per zygote. However, in those species whose ecology is otherwise well-suited to individual-level population studies, the prenatal part of an individual's life is often difficult to observe. While uncertainty has long surrounded the fertilization status of unhatched bird eggs-hatching failure can arise through fertilization failure or prenatal mortality-2 recent studies show fertilization failure to be extremely rare within 2 of the most popular avian study species. As such, unhatched eggs are highly reliable indicators of prenatal mortality. Although the generality of these results remains unclear, they demonstrate that prenatality can be incorporated into the observable lifespan of free-living animals. This allows zygotic LRS to be retrospectively quantified using historical nest observations and facilitates a more complete characterization of the evolutionary dynamics of wild populations.

Lifecycle of Anastatus orientalis (Hymenoptera: Eupelmidae) and synchrony with its host, the spotted lanternfly, Lycorma delicatula (Hemiptera: Fulgoridae).

Anastatus orientalis Yang & Choi (Hymenoptera: Eupelmidae), an egg parasitoid of spotted lanternfly, Lycorma delicatula (White) (Hemiptera: Fulgoridae), has been documented emerging from host eggs in both autumn and spring, at the beginning and end of the period that spotted lanternfly eggs are present in the field, suggesting parasitoid-host specificity and synchrony. This study was designed to test whether, under conditions that simulate native and introduced ranges of spotted lanternfly, (a) A. orientalis has 2 and only 2 generations per year, (b) A. orientalis adults sometimes emerge when only nontarget species would be available for parasitization, and (c) emerging parasitoid adults can parasitize unhatched host eggs in the spring. Parasitized spotted lanternfly eggs were collected in 2019 and 2020 from Beijing, China and in 2020 and 2021 from Yantai, China. They were shipped for laboratory study in growth chambers programmed to simulate temperature and daylength for collection locations in China and in the invaded range in Pennsylvania, United States. Anastatus orientalis had a flexible lifecycle depending on environmental conditions and possibly genetic makeup, with 1-3 generations per year, and parasitoid emergence was not always synchronous with host egg availability. Additionally, given the cooler temperatures in Pennsylvania, autumn parasitoid emergence was often delayed until late October or November, no progeny were produced, and parasitoid populations died out. Anastatus orientalis does not exhibit host synchrony characteristics that would make it a good candidate for a classical biological control program.

Hitting and Unhitting the Pause Button: Variable Hatching Patterns of Annual Killifish Embryos Over a Wetland Wet–Dry Cycle

ABSTRACTHatching dynamics is key to population structure of annual fishes in temporary wetlands. Given the risk of false starts due to unpredictable hydroperiods, annual fish may employ risk‐spreading strategies in their hatching patterns consistent with bet hedging (such as the partial hatching of their egg bank and variable number of hatching cues) after being exposed to floods. However, the extent to which annual fish resort to bet hedging (as well as their potential drivers) over a wet–dry cycle is poorly understood. We assessed the hatching patterns of a South American killifish (Matilebias cyaneus; Cyprinodontiformes: Rivulidae) collected at different hydrological phases of a temporary wetland to test for variation in bet hedging over a wet–dry cycle. We incubated wild eggs of M. cyaneus obtained from male–female pairings separately reared in tanks originating in the ‘early‐wet’, ‘drying’ and ‘late‐wet’ phases of a temporary wetland and examined hatching patterns across a sequence of hydration trials under common‐garden conditions. We found variation in (i) the number of hatched and unhatched eggs and (ii) the number of hatched eggs across hydration trials among clutches from each phase (while controlling for parentage). Our results show that M. cyaneus variably commit to bet‐hedging hatching strategies over a wet–dry cycle, and suggest that the environmental conditions at specific periods of the year and the parental age at which eggs are produced drive annual fish hatching patterns.

Temporal trends (1972-2017) and spatial differences of persistent halogenated aromatic hydrocarbons in osprey eggs in Finland.

Time trends and regional differences of polychlorinated dibenzo-p-dioxins and -furans (PCDD/Fs), polychlorinated biphenyls (PCBs), polychlorinated naphthalenes (PCNs), DDTs, polybrominated biphenyls (PBBs) and polybrominated diphenylethers (BDEs) were studied in unhatched osprey eggs collected by bird ringers in 1972-2017 from four areas in Finland. Two study areas were from Baltic Sea, Northern Quark and Finnish Archipelago Sea, while the two others were inland lake areas, eutrophicated Lake Vanajanselkä affected by industrial emissions, and Pristine SW Lake Area. The highest concentrations of most compound groups were in Lake Vanajanselkä consistent with high emissions, the predominance of bream as a prey, and higher concentrations in bream compared to other prey fish. Concentrations of all chlorinated compounds decreased significantly in all study areas. Average annual decreases were ∑PCDD/F 2.3-4.9%, ∑PCB 2.2-4.2%, ∑PCN 2.6-7.0% and ∑DDT 7.1-9.5%, primarily in line with decreased levels in prey fish. From 1972 PBBs and BDEs increased significantly until 1990s declining rapidly thereafter. PCDD/F congener profile was dominated by 2,3,4,7,8-PeCDF, except in Lake Vanajanselkä by 1,2,3,6,7,8-HxCDD. PCB congener profile was dominated by PCB 153 in all study areas, followed by PCB 180 and PCB 138. Among dioxin-like compounds PCBs contributed 82%, PCDDs 14% and PCDFs 4% to toxic equivalent quantity (∑TEQ). PCB 126 contributed most to ∑TEQ, followed by 1,2,3,7,8-PeCDD. BDE 47 being the dominant BDE congener, followed by BDE 100. ∑DDT concentrations were relatively similar across all study areas, with DDE contributing about 90%. Productivity of chicks per active nest was significantly decreased in Lake Vanajanselkä, and the likely explanation is embryotoxicity of dioxin-like compounds. It is plausible that dioxin-like compounds influenced embryonic survival among highly exposed ospreys prior to 2010, especially in Lake Vanajanselkä and Northern Quark. However, decreased survival due to DDE-induced eggshell thinning seems unlikely after 1985, and BDE levels were below those potentially causing adverse effects.

Congenital Anomalies in American Crocodile (Crocodylus acutus, Cuvier, 1807) Embryos from a Farm Breeder in Colombia.

The American crocodile (Crocodylus acutus, Cuvier, 1807) (Class Reptilia, Family Crocodylidae) is a crocodile species inhabiting the Neotropics. Congenital defects have been described in almost every vertebrate group. In crocodiles, teratology alterations have been described in captive animals (pets, zoos, farms) such as Crocodylus niloticus or Gavialis gangeticus. The present study aimed to characterize congenital malformations of C. acutus from a farm in Lomas de Matunilla, Ballestas, Bolívar, Colombia. A total of 550 unhatched eggs were examined after embryo death. A total of 61 embryos presented malformations, with 42 different types of anomalies observed. Limb and tail malformations (29%) were the most common malformations observed. Several malformations, such as cephalothoracopagus, thoracopagus, sternopagus, xiphopagus twins, campylorrachis scoliosa, and acrania, were documented in crocodiles for the first time. Research in teratology enhances our understanding of crocodile biology. It plays a role in their conservation and management, thus helping to ensure the long-term viability of these species in their natural habitats.

Chemical threats for the sentinel Pygoscelis adeliae from the Ross Sea (Antarctica): Occurrence and levels of persistent organic pollutants (POPs), perfluoroalkyl substances (PFAS) and mercury within the largest marine protected area worldwide

The Ross Sea Marine Protected Area (RS-MPA) hosts endemic species that have to cope with multiple threats, including chemical contamination. Adèlie penguin is considered a good sentinel species for monitoring pollutants. Here, 23 unhatched eggs, collected from three colonies along the Ross Sea coasts, were analysed to provide updated results on legacy pollutants and establish a baseline for newer ones. Average sum of polychlorinated biphenyls (∑PCBs) at the three colonies ranged 20.9–24.3 ng/g lipid weight (lw) and included PCBs IUPAC nos. 28, 118, 153, 138, 180. PCBs were dominated by hexachlorinated congeners as previously reported. Hexachlorobenzene (HCB) and p,p’-dichlorodiphenyldichloroethylene (p,p’-DDE) ranged between 134 and 166 and 181–228 ng/g lw, respectively. Overall, ∑PCBs was exceeded by pesticides, contrary to previous studies from the Ross Sea. Sum of polybrominated diphenyl ethers (∑PBDEs) ranged between 0.90 and 1.18 ng/g lw and consisted of BDE-47 (that prevailed as expected, representing 60–80 % of the ∑PBDEs) and BDE-85. Sum of perfluoroalkyl substances (∑PFAS) ranged from 1.04 to 1.53 ng/g wet weight and comprised five long-chain perfluorinated carboxylic acids (PFCAs), perfluorooctane sulfonate (PFOS), perfluorohexane sulfonate (PFHxS) and perfluorooctanoic acid (PFOA); perfluorooctane sulfonamide (PFOSA) was also detected. The PFAS profile was dominated by PFCAs as already observed in Arctic seabirds. Mercury ranged from 0.07 to 0.15 mg/kg dry weight similarly to previous studies. Legacy pollutants confirmed their ongoing presence in Antarctic biota and their levels seemed mostly in line with the past, but with minor variations in some cases, likely due to continued input or release from past reservoirs. PFAS were reported for the first time in penguins from the Ross Sea, highlighting their ubiquity. Although further studies would be useful to increase the sample size and accordingly improve our knowledge on spatial and temporal trends, this study provides interesting data for future monitoring programs within the RS-MPA that will be crucial to test its effectiveness against human impacts.

Comparative reproduction performance of Giant African Land Snail (Archachatina marginata) fed with the leguminous plant Leucaena leucocephala

The study was carried out between July and October 2022 at the sub–station of the Faculty of Agriculture Research and Teaching farm to assess the reproductive performance of Archachatina marginata fed the leguminous Leucaena leucocephala leaf meal as feed additive. 208 adult snails were randomly partitioned into 4 treatments and 4 repetitions of 13 comparable snails (weight and size). The treatment T0 (control) received only the experimental diet as described while three other treatments received not only diet but, L. leucocephala leaf meal at 0.25%, 0.5% and 0.75% for a period of 12 weeks. In each treatment, every morning, the substrate of each breeding bucket was stirred entirely and minutely to collect the eggs to determine the laying and morphometric characteristics of the eggs. The eggs were then placed 10 cm deep in loose soil substrates until hatching to evaluate the characteristics of the fecundity. The unhatched eggs were opened, and the state of embryonic development observed to determine early embryonic mortality rate and late embryonic mortality. At the end of the experiment, animals of each replicate were sacrificed; the hemolymph was collected determine the total cholesterol and protein. The results showed that the characteristics of reproduction were affected with graded level of L. leucocephala. The highest number of eggs laid (17.29 ± 13.44) and the number of clutches (268.91) was recorded in animals receiving 0.5% and 0.75% L. leucocephala in the diet respectively while the highest value of fecundation rate (95.50 ± 8.80), hatch rate (94.55 ± 9.36), and hatchability (98.99 ± 3.15) was registered in the treatment that received 0.75%. The highest value of total protein and cholesterol was recorded in the same treatment. In conclusion 0.75% of L. leucocephala can be added in the diet of adult snail to increase its reproductive performance.

Natural products from Xenorhabdus and Photorhabdus show promise as biolarvicides against Aedes albopictus.

In the perpetual struggle to manage mosquito populations, there has been increasing demand for the development of biopesticides to supplant/complement current products. The insecticidal potential of Xenorhabdus and Photorhabdus has long been recognized and is of interest for the control of important mosquitoes like Aedes albopictus which vectors over 20 different arboviruses of global public health concern. The larvicidal effects of cell-free supernatants, cell growth cultures and cell mass of an extensive list of Xenorhabdus and Photorhabdus spp. was investigated. They were quite effective against Ae. albopictus causing larval mortality ranging between 52-100%. Three Photorhabdus spp. and 13 Xenorhabdus spp. release larvicidal compounds in cell-free supernatants. Cell growth culture of all tested species exhibited larvicidal activity, except for Xenorhabdus sp. TS4. Twenty-one Xenorhabdus and Photorhabdus bacterial cells (pellet) exhibited oral toxicity (59-91%) against exposed larvae. The effect of bacterial supernatants on the mosquito eggs were also assessed. Bacterial supernatants inhibited the hatching of mosquito eggs; when unhatched eggs were transferred to clean water, they all hatched. Using the easyPACId approach, the larvicidal compounds in bacterial supernatant were identified as fabclavine from X. szentirmaii and xencoumacin from X. nematophila (causing 98 and 70% mortality, respectively, after 48 h). Xenorhabdus cabanillasii and X. hominickii fabclavines were as effective as commercial Bacillus thuringiensis subsp. israelensis and spinosad products within 5 days post-application (dpa). Fabclavine and xenocoumacin can be developed into novel biolarvicides, can be used as a model to synthesize other compounds or/and can be combined with other commercial biolarvicides. © 2024 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Investigation into the Impacts of Varied Batches of Stripped Eggs on the Reproductive Success of Clarias gariepinus

This study was conducted at the hatchery complex of Akwa Ibom State University to investigate the effects of different batches of stripped egg on reproductive performances of Clarias gariepinus. Nine brood stocks of Clarias gariepinus (six males and 3 females) with average body weight of 2.3kg and 64.2 cm where carefully selected for the study. Six matured males were euthanized for the removal of testis without hormonal inducements. The milt collected from the six males were pooled together and divided into 12 portions each diluted with 2mL of normal saline solution. Three female brood stocks where separately induced with hormone at single dosage of 0.5ml/kg body weight and allowed for a period of 10 hours before stripping. Stripping of eggs from each brood stock where in four batches, each batch measured 50g and labeled A, B, C and D. 3g of eggs containing approximately 2000 oocytes (eggs) where measured out from each batch and mixed with the diluted milt and incubated in a 2 x 1 x 10cm3 of water in a concrete pond at temperature of 26oC in three replicates. The result revealed that the reproductive performances considered in this study decreased as the batches of stripped eggs increased from 1 ̶ 4 batches. The first batch of stripped eggs produced 87.69 ± 0.42, 97.06 ± 0.84, 84.29 ± 1.05 percentage fertilization, hatchability and survival rate respectively, while the least batch of stripped eggs (batch 4) produced the least value of fertilization 50.33 ± 1.67, 31.93 ± 4.94 hatchability and survival rate value of 42.69 ± 0.61. Therefore, from the above result, the least batch of stripped eggs should not be used for fertilization because the unhatched or dead eggs has a negative effect on the fertilized eggs including fry at the hatchery level.

Could persistent organic pollutants affect future generations of sea turtles by maternal transfer? First results for Caretta caretta nests along the North-Western coast of Italy

Since 2013, loggerhead sea turtle (Caretta caretta, Linnaeus 1758) nesting has been observed further north along the Italian coast, reaching the Tuscan coast (NW Mediterranean Sea). The four nesting events that occurred in Tuscany in the summer of 2019 spurred the scientific community to monitor these occurrences more carefully, following them from egg deposition to hatching. This provided an opportunity to collect samples for conducting multidisciplinary investigations, including the toxicological investigations of the biological material collected from the four nests. The aim of this study was to conduct an initial assessment of persistent organic pollutants in the eggs laid in Tuscany, aiming to establish a baseline on this topic for subsequent nesting events that have occurred until today. Organochlorine compounds (OCs), specifically polychlorinated biphenyls (PCBs), dichlorodiphenyltrichloroethane (DDT) and its metabolites, and hexachlorobenzene (HCB), were analyzed and detected in unhatched eggs, embryos, and chorio-allantoic membranes (CAMs). OCs were detected in all samples, with PCBs > DDTs ≫ HCB. A significant spatial variation in pollutant levels and profiles among sea turtle nesting locations was found. Embryos showed higher levels of contamination than egg contents regardless of the developmental stages. Depth of the laying chamber and egg mass were not significant factors in OC bioaccumulation. For the first time in the Mediterranean Sea, this study assessed the role of CAM in the transfer of contaminants to the embryo. Overall, the OC levels found were lower compared to the results from other studies conducted worldwide on loggerhead sea turtle biological material. This was the first assessment of nest biological material for the North-Western coast of Italy.

Effects of Multiple Batches of Stripped Eggs on the Reproductive Performances of Heterobranchus longifilis

Studying the effects of multiple batches of stripped eggs contributes to the sustainability and viability of aquaculture practices by improving our understanding of reproductive performance and informing best practices in breeding programs, hence, the need to investigate on the effects of multiple batches of stripped egg on reproductive performances of Heterobranchus logifilis became necessary. The aim was to provide valuable information for decision-making processes in aquaculture operations. Nine brood stocks of H. longifilis (6 male & 3 female) with average body weight of 2.3 kg and average length of 64.2 cm where carefully selected for the study. Six matured males were sacrificed for the removal of testis without hormonal inducements. The milt collected from the six males were pulled together and divided into twelve portions each diluted with 2ml of normal saline solution. Three female brood stocks where separately induced at single dosage of 0.5ml/kg body weight and allowed for a period of 10 hours before stripping. Stripping of eggs from each brood stock where in four batches, each batch measured 50g and labeled A, B, C & D. 3g of eggs containing approximately 2000 oocytes (eggs) where measured out from each batch and mixed with the diluted milt and incubated in a 2 x 1 x 10cm3 of water in a concrete pond at temperature of 26oC in three replicates. The result revealed that the reproductive performances considered in this study decreased as the batches of stripped eggs increased from 1- 4 batches. The first batch of stripped eggs produced 90.17 ± 0.44%, 97.78±0.86%, 89.48 ±1.08% fertilization, hatchability and survival respectively, while the least batch of stripped eggs (batch 4) produced the least value of fertilization (5.83 ± 1.69%); 45.00±4.90% hatchability and survival value of 5.59 ± 0.61%. Sequel to the findings of the present study, the least batch of stripped eggs should be avoided for use in fertilization since the unhatched or dead eggs has some detrimental effects on the fertilized eggs including fry at the hatchery level.

Effect of hen age on egg quality, hatching performance and chick quality in Chukar Partridges (Alectoris chukar)

Aim: This study was aimed to investigate the effect of different hen ages on egg quality characteristics, hatching performance and chick quality in chukar partridges. Materials and Methods: A total of 510 eggs obtained from chukar partridge flocks were used to determine the effects of different hen age on hatching performance, egg quality traits and chick quality. Daily collected 20 eggs from each age group (1, 2 and 3 years of age) were assessed for internal quality characteristics. A total of 150 eggs per each age group were set into incubator and hatcher set. After hatching, unhatched eggs were broken to determine hatching performance and embryonic mortality. Hatched day-old chicks were scored to determine chick quality characteristics. Results: Internal egg quality traits were not statistically affected by different breeding ages. Different breeding ages did not affect hatchability, early and middle embryonic mortality while late embryonic mortality and fertility of eggs were statistically affected. The evaluated chick quality traits were affected by the different hen age. Conclusion: Despite decreasing fertility rate with age, the results of this study suggest incubating eggs of chukar hens with 3 years age for optimum chick quality and being raised chukar hens up to 3 years of age.

First report of the egg fertility of the hawksbill turtle (Eretmochelys imbricata) (Testudines: Cheloniidae) in Melaka, Malaysia

The unhatched eggs of the hawksbill turtle (Eretmochelys imbricata) are often found in the hatchery in Melaka, Malaysia, raising concerns if the eggs remain unfertilized. Examining unhatched eggs during post-emergence nest excavation could lead to eggs being mistakenly identified as unfertilized eggs due to the difficulty in differentiating between unfertilized eggs and early embryonic mortality. It is essential to determine the egg fertility of hawksbill turtles in Melaka before concluding that hatching failure is due to infertility. Egg fertility was determined using the white spot method conducted at the beginning of egg incubation on 50 clutches. Mean egg fertility was 85.9%, but the mean hatching success was significantly lower at 57.6% (mean emergence success was 56.8%). The results showed that egg fertility did not reflect hatching success and infertility was unlikely the cause of hatching failure. The huge number of undeveloped eggs may have been due to early embryonic death that went undetected at the end of incubation.

Chemo-profiling of Purpureocillium lilacinum and Paecilomyces variotii isolates using GC-MS analysis, and evaluation of their metabolites against M. incognita.

Nematophagous fungi are the best alternatives to chemical nematicides for managing nematodes considering environmental health. In the current study, activity of metabolites from ten isolates of Purpureocillium lilacinum (Thom) Luangsa-ard (Hypocreales: Ophiocordycipitaceae) and two isolates of Paecilomyces variotii Bainier (Eurotiales: Trichocomaceae), were examined to inhibit the hatching of Meloidogyne incognita (Kofoid & White) Chitwood (Tylenchida: Heteroderidae) eggs. At 100%, 50%, and 25% concentrations, respectively, the culture filtrate of the isolate P. lilacinum 6887 prevented 97.55%, 90.52%, and 62.97% of egg hatching. Out of all the isolates, Pl 6887, Pl 6553, and Pl 2362 showed the greatest results in the hatching inhibition experiment.Gas chromatography-mass spectrometry (GC-MS) analysis revealed a variety of nematicidal compounds from different isolates. A total of seven nematicidal compounds, including four very potent nematicidal fatty acids were found in the isolate Pl 6553. Secondary metabolites of the same isolate possess the highest M. incognita juvenile mortality, i.e., 43.33% and 92% after 48 hrs of treatment at 100 and 200 ppm concentrations, respectively. Significant difference was observed in juvenile mortality percentage among the isolate having highest and lowest nematicidal compounds. Nematicidal fatty acids like myristic and lauric acid were found for the first time in P. lilacinum. Multiple vacuole-like droplets were found inside the unhatched eggs inoculated with the culture filtrate of isolate Pl 6887, and also in the juveniles that perished in the ethyl acetate extract of isolate Pl 6553.

Hard life for sons in the nest? Sex-dependent offspring mortality in Great Tits in urban and forest areas

Sex-biased mortality can occur in birds during development, for example due to sexual differences in energy requirement and/or environmental sensitivity, or the effects of sex hormones or sex differences in the expression of mutations linked to sex chromosomes. The extent of sex-bias in mortality may also be related to environmental conditions that influence offspring development and survival. Urban areas often provide poorer conditions for nestling development resulting in higher offspring mortality compared to natural areas, which may accelerate sex differences in offspring mortality in cities. To test this hypothesis, we examined the sex ratio of dead offspring in Great Tits (Parus major), using 427 samples of unhatched eggs and dead nestlings collected in two urban and two forest sites between 2013 and 2019. The ratio of males in the whole sample of dead offspring (56.9%) was significantly higher than expected by an 1:1 ratio, and the strongest sex biases were detected in urban areas (57.6% males) and in young nestlings (<14 days old, 59.0% males). However, the sex ratios of dead offspring did not differ significantly among study sites and between offspring developmental stages. 29.3% of unhatched eggs contained a visible embryo, and the proportion of embryo-containing unhatched eggs did not differ significantly between urban and forest study sites. These results suggest male-biased offspring mortality in Great Tits, and highlight the need of large datasets to detect subtle differences between habitats and developmental stages.

Phytochemical Screening and in-vitro Efficacy of Calpurnia aurea Against Two Transovarial Vectors: Amblyomma variegatum and Rhipicephalus microplus.

Ticks are the second most common vector of human infectious diseases after mosquitoes. Their transovarial transmission contributes to the maintenance of environmental diseases. This study evaluates the phytochemical screening and in vitro efficacy of Calpurnia aurea against the adult survival and egg hatchability of two transovarial transmission vectors: Amblyomma variegatum and Rhipicephalus microplus. Plant material was extracted using maceration techniques, and concentrated solutions of 12.5, 25, 50, 100, 200, and 400 ppm were prepared. Distilled water and diazinon were used as negative and positive controls, respectively. Ten adult ticks were exposed for 10 minutes, and dead ticks were counted after 24 hours of recovery. Twenty 15-day-old eggs were immersed for 10 minutes, and after 15 days of incubation, hatched and unhatched eggs were tallied. Preliminary phytochemical constituents were screened. A one-way analysis of variance and the probit regression model determined mean mortality and hatchability and estimated lethal and inhibitory concentrations, respectively. The ethanolic and aqueous leaf extracts caused 10±0.0% mortality in adult A. variegatum and R. microplus. The effective dose was LC50 of 27 and 29 ppm and LC50 of 37 and 41 ppm, respectively. At 400 ppm, the leaf ethanolic and aqueous extracts showed 18.7±0.9% and 18.3±1.7%; 18.3±1.2% and 19.7±0.3% egg hatching inhibition, respectively. The effective dose had an IC50 of 50 ppm and IC50s of 91 and 79 ppm, respectively. Flavonoids and saponins were found in both leaf and pod extracts. C. aurea extracts showed a more promising effect on tick survival and hatchability than synthetic diazinon. The susceptibility test indicated that the leaf extract could control vectors and contribute to environmental disease maintenance. Complex phytochemicals, especially phenolic compounds, are additional evidence of effectiveness in vector control. Further investigation of in vivo efficacy and advanced fractionation of phytochemicals is needed.