Embryo Liver Research Articles

Developmental toxicity caused by sanguinarine in zebrafish embryos via regulating oxidative stress, apoptosis and wnt pathways

Sanguinarine, derived from the root of Sanguinaria canadensis, have multiple biological activities, such as antimicrobial, insecticidal, antitumor, anti-inflammatory and anti-angiogenesis effect, but little is known about its toxicity on normal embryonic development. Here, we study the developmental toxicity using zebrafish model. Notably, sanguinarine caused a significant increase of the malformation rate and decrease of hatching rates and body length of zebrafish embryos. Sanguinarine also impaired the normal development of heart, liver and nerve system of zebrafish embryos. Further, the ROS level and MDA concentrations were remarkably increased, while the activity of T-SOD was decreased. In addition, obvious increase of apoptosis were observed by AO staining or TUNEL assay. Further studies showed that the oxidative stress-, apoptosis-related genes were changed, while genes of nrf2 and wnt pathways were inhibited by sangunarine. To sum up, our study will be helpful to understand the adverse effect of sanguinarine on embryonic development and the underlying molecular mechanism.

Microscopic study of potential toxic effects of monosodium glutamate on liver of chicken embryos aged 16 days

BackgroundThis experimental study aimed to determine the changes caused by monosodium glutamate (MSG) on morphology and histology of liver of chicken embryos aged 16 days of incubation. In this research, 50 fertilized eggs were used. They were divided into two equal groups, one group was used as control with normal liver structure, while the other group (MSG) was treated with MSG in 0 day of incubation (0.1ml/egg).ResultsThe results showed many harmful effects on hepatic cells, blood sinusoids, and bile ducts in MSG group. These changes included alterations in nuclei conformation and nuclear envelope. Chromatin distribution was associated with increased electronic intensity. Also, there were rupture in smooth endoplasm systems and malfunction of mitochondria and Golgi apparatus, with increased lysosomes and lipid droplets.ConclusionsThis study concluded that MSG had severe toxic effects on liver structure if it was given in pre-conception period as this period is considered critical for liver growth (organogenesis).

ROS-Triggered Autophagy Is Involved in PFOS-Induced Apoptosis of Human Embryo Liver L-02 Cells.

The liver is the primary target organ for perfluorooctane sulphonate (PFOS), a recently discovered persistent organic pollutant. However, the mechanisms mediating hepatotoxicity remain unclear. Herein, we explored the relationship between reactive oxygen species (ROS) and autophagy and apoptosis induced by PFOS in L-02 cells, which are incubated with different concentrations of PFOS (0, 50, 100, 150, 200, or 250 μmol/L) for 24 or 48 hrs at 37°C. The results indicated that PFOS exposure decreased cell activities, enhanced ROS levels in a concentration-dependent manner, decreased mitochondrial membrane potential (MMP), and induced autophagy and apoptosis. Compared with the control, 200 μmol/L PFOS increased ROS levels; enhanced the expression of Bax, cleaved-caspase-3, and LC3-II; induced autophagy; decreased MMP; and lowered Bcl-2, p62, and Bcl-2/Bax ratio. The antioxidant N-acetyl cysteine (NAC) protected MMP against PFOS-induced changes and diminished apoptosis and autophagy. Compared with 200 μmol/L PFOS treatment, NAC pretreatment reversed the increase in ROS, Bax, and cleaved-caspase-3 protein caused by PFOS, lowered the apoptosis rate increased by PFOS, and increased the levels of MMP and Bcl-2/Bax ratio decreased by PFOS. The autophagy inhibitor 3-methyladenine and chloroquine decreased apoptosis and cleaved-caspase-3 protein level and increased the Bcl-2/Bax ratio. In summary, our results suggest that ROS-triggered autophagy is involved in PFOS-induced apoptosis in L-02 cells.

Integrative Analysis of Transcriptomic and Epigenomic Dynamics of Induced Pluripotent Stem Cell Derived Hepatic Organoids Using Single Cell RNA-Seq and ATAC-Seq

Previously, we developed a novel method by which to generate functionally mature human hepatic organoids derived from pluripotent stem cells (PSCs), and the functionality and maturity of the organoids were validated by various experiments including bulk RNA-seq. In this study, to identify the heterogeneity and dynamics of transcriptomes and epigenomes of the organoids at the single-cell level, we applied single-cell RNA-seq (scRNA-seq) and single-cell ATAC-seq (scATAC-seq) to analyze liver organoids treated with two medium conditions—hepatic medium (HM, expandable condition) and differentiation medium (DM, more matured condition). The cells were separated into 13 distinct clusters corresponding to stages ranging from early to late by both scRNA-seq and scATAC-seq, and DM liver organoids were more similar in more differentiated liver cells compared to those in HM. Mitochondrial gene expression was increased in differentiated hepatocytes, with ATAC-seq peaks enriched near the mitochondrial control region. Many transcription factors (TFs)—both activators and repressors—were revealed throughout the differentiation of liver organoids. Different patterns of expression of alpha-fetoprotein (AFP) and albumin (ALB) were observed between liver organoids and adult liver tissues, and several open chromatin regions (OCRs) near AFP and ALB genes were found to be differentially regulated between the liver organoids and tissues. HNF4A, YY1 (Yin Yang 1) and CTCF binding sites near AFP and ALB genes were differentially occupied between embryo and adult liver tissues. Our integrative analysis of transcriptomic and epigenetic dynamics of the liver organoids can contribute to the understanding of transcriptional regulatory networks during liver development and to the further development of mature in vitro human liver models.

Effects of compound probiotics and aflatoxin-degradation enzyme on alleviating aflatoxin-induced cytotoxicity in chicken embryo primary intestinal epithelium, liver and kidney cells

Aflatoxin B1 (AFB1) is one of the most dangerous mycotoxins for humans and animals. This study aimed to investigate the effects of compound probiotics (CP), CP supernatant (CPS), AFB1-degradation enzyme (ADE) on chicken embryo primary intestinal epithelium, liver and kidney cell viabilities, and to determine the functions of CP + ADE (CPADE) or CPS + ADE (CPSADE) for alleviating cytotoxicity induced by AFB1. The results showed that AFB1 decreased cell viabilities in dose-dependent and time-dependent manners. The optimal AFB1 concentrations and reactive time for establishing cell damage models were 200 µg/L AFB1 and 12 h for intestinal epithelium cells, 40 µg/L and 12 h for liver and kidney cells. Cell viabilities reached 231.58% (p < 0.05) for intestinal epithelium cells with CP addition, 105.29% and 115.84% (p < 0.05) for kidney and liver cells with CPS additions. The further results showed that intestinal epithelium, liver and kidney cell viabilities were significantly decreased to 87.12%, 88.7% and 84.19% (p < 0.05) when the cells were exposed to AFB1; however, they were increased to 93.49% by CPADE addition, 102.33% and 94.71% by CPSADE additions (p < 0.05). The relative mRNA abundances of IL-6, IL-8, TNF-α, iNOS, NF-κB, NOD1 (except liver cell) and TLR2 in three kinds of primary cells were significantly down-regulated by CPADE or CPSADE addition, compared with single AFB1 group (p < 0.05), indicating that CPADE or CPSADE addition could alleviate cell cytotoxicity and inflammation induced by AFB1 exposure through suppressing the activations of NF-κB, iNOS, NOD1 and TLR2 pathways.

The Embryotoxic Effects of in Ovo Administered Sunset Yellow FCF in Chick Embryos.

Sunset yellow (SY) at prescribed concentrations has been approved by regulatory authorities in several countries as an additive dye in the food, beverage, cosmetic, and pharmaceutical industries. However, there are some reports that it may cause several health problems. The aim of this study is to evaluate embryotoxic effects of SY on liver and kidney in chick embryos. Babcock white Leghorn eggs were randomly divided into four groups. Non-treated eggs served as control group. The eggs in groups SY200, SY1000, and SY2000 were treated with a single injection of 200, 1000, and 2000 ng SY into the air sac just before incubation. The developmental stages of embryos were determined on the 10th, 13th, 16th, and 21st days of incubation. Samples of the liver and kidney were taken and routine histological procedures were performed. The highest relative embryo weight was seen in all SY treated groups on the 16th day of incubation. Necrosis of some hepatocytes and cytoplasmic degenerations were observed in all SY groups in the liver. There were degenerated or destructed renal cortex structures and necrosis in the kidney. The cell’s nuclear areas and diameters of renal cortex structures were different in all SY groups compared to the control group (p < 0.05). It was concluded that in ovo administered SY has many unfavorable effects on liver and kidney in chick embryos. The results obtained in this study suggest that it may be advisable to re-assess safety levels of SY in many industries.

Characterization of rat liver bud-derived cells

Cells derived from the fetal liver have been shown to be a rich source of progenitor stem cells, constituting a promising source for Tissue Engineering and Regenerative Medicine. In this study, embryo and fetal liver-bud derived cells from Fischer 344 rats were obtained at E12.5, E14.5 and E16.5 gestational days and evaluated for cell phenotype, survival and proliferation. Liver transaminase (AST and ALT) and AFP levels were lower in embryo liver-bud-derived cells on day 12.5. Markers for stem cells, cell cycle progression and cell death were differentially expressed in E12.5 cell cultures. Analysis of mitochondrial electric potential on 14.5 and 16.5 days showed a tendency for cells with lower functional or metabolic ability, in comparison to cultures derived from day 12.5. The results demonstrated that the majority of the E16.5 cells were in the G0 / G1 phase. The capacity of synthesis (S) and cellular division (G2 / M) of embryo and fetal liver bud-derived cells was constant over all gestational periods. In conclusion, embryo and fetal liver-bud-derived cells during the periods of 12.5 and 14.5 days, showed expression profile of progenitor cells, cell activity and hematopoietic function in culture.

Comprehensive genomic analysis and characterization of a new ST 174 type Klebsiella variicola strain isolated from chicken embryos.

Klebsiella variicola is a widespread opportunistic pathogen that causes infections in humans and animals. Herein a novel Klebsiella strain, AHKv-S01, was isolated and identified from dead chicken embryos in Anhui, China. Its genome contained a circular chromosome of 5,505,304bp, with 5244 protein-coding genes, and an integrative conjugative element region containing 79 ORF sequences. AHKv-S01 was given a new sequence type number-174. Phylogenetic analyses showed that rpoB partial nucleotide sequences were highly reliable for identifying Klebsiella spp. Most of the 340 unique genes of AHKv-S01 were involved in cell envelop biogenesis, transcription, transport, and metabolic processes. Moreover, AHKv-S01 was sensitive to several antibiotics, but it showed strong resistance to penicillins, macrolides, and lincosamide. The genome contained three drug efflux pump superfamilies, β-lactamase genes, and fosfomycin resistance-related genes. Most drug resistance genes showed amino acid mutations. Multiple virulence and pathogenic factors were also identified, and they were mainly related to adhesion, secretion, iron acquisition, and immune evasion. Chicken embryo lethality assay results revealed that the 7-day chicken embryo lethality rate was 80%, 40%, and 50% for AHKv-S01, K. pneumoniae ATCC10031, and K. pneumoniae CICC24714, respectively. The median lethal dose of AHKv-S01 was 39.9CFU/embryo. Even low infection levels of AHKv-S01 caused a significant reduction in chicken embryo hatchability. Severe pathological changes to the liver, heart, and brain tissues of embryos infected with AHKv-S01 were observed, and these changes appeared earlier in the heart and brain than in the liver. To conclude, our results provide a foundation for further studies aiming to assess the potential risk of K. variicola to poultry populations and production yields.

Monochromatic green light stimulation during incubation shortened the hatching time via pineal function in White Leghorn eggs

BackgroundEffect of monochromatic green light illumination on embryo development has been reported in chickens. The avian pineal gland is an important photo-endocrine organ formed by a mediodorsal protrusion during embryonic development. However, the involvement of pineal gland in the light transduction process remains to be elucidated. In the present study, we investigated the influence of monochromatic green light on hatching time and explored the possible mechanism via pineal function.ResultsA total of 600 eggs of White Leghorn (Shaver strain) were incubated under photoperiods of either 12 h of light and 12 h of darkness using monochromatic green light (12L:12D group) or 24 h of darkness (0L:24D group) for 18 d. Compared to 0L:24D group, the green light stimulation shortened the hatching time without extending the hatch window or impairing hatchability. The liver of embryos incubated in the 12L:12D light condition was heavier than those of the 0L:24D group on d 21 post incubation which may be linked to the observed increase in the serum concentration of insulin-like growth factor 1 (IGF-1); primary secretion of the liver. Histological structure analysis of pineal gland demonstrated that the light stimulation increased follicle area, wall thickness and lumen area on d 10 and d 12 post incubation. Rhythmic function analysis demonstrated that three clock related genes (brain and muscle ARNT-like-1, BMAL1; circadian locomotor output cycles kaput, CLOCK; and cryptochrome-1, CRY1) and a melatonin rate-limiting enzyme related gene (arylalkylamine N-acetyltransferase, AANAT) were rhythmically expressed in the pineal gland of the 12L:12D group, but not in the 0L:24D group. Simultaneously, the light stimulation also increased the concentration of melatonin (MT), which was linked to hepatocyte proliferation and IGF-1 secretion in previous studies.ConclusionsThe 12L:12D monochromatic green light stimulation during incubation shortened hatching time without impairing hatching performance. Pineal gland’s early histological development and maturation of its rhythmic function were accelerated by the light stimulation. It may be the key organ in the photo-endocrine axis that regulates embryo development, and the potential mechanism could be through enhanced secretion of MT in the 12L:12D group which promotes the secretion of IGF-1.

Graphene oxide nanofilm and chicken embryo extract decrease the invasiveness of HepG2 liver cancer cells

BackgroundThe extracellular matrix (ECM) is a mosaic of various structural and functional proteins that cooperate with the cell, regulate adhesion, and consequently manage its further fate. Liver destruction is accompanied by a disruption of the physicochemical properties of the ECM which deregulates the cell–ECM interaction and can lead to uncontrolled proliferation and neoplastic transformation of cells. Therefore, it can be assumed that ECM modification and restoration of its characteristics for healthy tissue may counteract uncontrolled cell proliferation. The purpose of the presented research model was to optimise the physical characteristics of ECM by introducing a graphene oxide plane/nanofilm (nfGO) and enriching the cell environment with potentially missing proteins by adding a functional protein cocktail (chicken embryo liver extract) and determine the impact of these factors on cell–ECM cooperation and its consequences on adhesion, proliferation, and cell phase, which are factors of the invasiveness of cancer cells.ResultsExperiments were performed with non-cancer HS-5 cells and liver cancer cells HepG2 and C3A. The cells were divided into four groups: (1) control, (2) cultured on nfGO, (3) cultured with the addition of chicken embryo liver extract (CELE) and (4) cultured on the nfGO with the addition of CELE. CELE contained 1735 proteins; the top 57 of these proteins have been presented. The use of nfGO as well as CELE and nfGO + CELE reduced the proliferation of HepG2 cancer cells to the greatest extent; this is in contrast to non-cancer cells and also to C3A cancer cells. Furthermore, the combined use of the CELE protein cocktail and GO substrate effectively resulted in a decrease in the population of HepG2 cells in the G0/G1 phase and an increase of the population in G2/M. Molecular analysis of HepG2 cancer cells also showed an increase in the expression of genes responsible for adhesion such as focal adhesion kinase (fak), e-cadherin, and n-cadherin and a decrease in β-catenin, which is considered a proto-oncogene.ConclusionsStudies have shown that both the GO surface structure on which the cells are grown as well as the presence of a multi-component natural cocktail of regulatory proteins, can modify the expression of integrins, increase adhesion and, as a consequence, proliferation and the cell cycle—entering the resting phase. For the first time, it has been documented that hepatic cancer cells of the HepG2 line under the influence of stimuli derived from mimic ECM (graphene oxide) in interaction with a unique protein complex derived from chicken liver embryo decreased of the invasiveness of cancer cells.

ВЛИЯНИЕ ОСЛАБЛЕННОГО МАГНИТНОГО ПОЛЯ ЗЕМЛИ НА РАЗВИТИЕ ПЕЧЕНИ И СЕЛЕЗЕНКИ ЭМБРИОНОВ ЯПОНСКОГО ПЕРЕПЕЛА

Expectation of remote space missions and long-term stay and work on the Moon with the magnetic field 1,000 times weaker than on Earth sets the researchers the formidable task to investigate effects of the hypomagnetic environment on living organisms. The paper reports data about the liver and spleen development in Japanese quail embryos of various age exposed in a modeled lunar magnetic field. Retardation of hemopoiesis was observed as in the first generation embryos (F1), so in sequential embryo generations developed in the ordinary magnetic environment (F2).

44 Exploring the use of silver and diamond nanoparticles on sperm cell invitro and chicken embryo in ovo

In poultry industry, chick viability is a crucial factor determining profitability from fertilized egg to placement at the farm. However, decreases in fertility and hatchability have been observed. Recently, there has been renewed interest in the use of silver nanoparticles (Ag-NPs) due to their antimicrobial properties and growth-promoting ability, and diamond nanoparticles (D-NPs) due to their biocompatibility properties. The aim of the study was to evaluate the effect of silver and diamond nanoparticles on chicken embryo oxidative status, biochemical indices, and expression of immune-related genes and on sperm cell viability. The experiment was conducted in Ross 308 chicken embryos and Ross 308 cockerels. One hundred and fifty fertilized eggs were divided randomly into 5 groups (5×30). Fertilized eggs were injected with 50 mg/L Ag-NPs at volumes of 100μL (group 1), 200μL (group 2) or 50 mg/L D-NPs at volumes of 100μL (group 3) or 200μL (group 4), or received no nanoparticles (control; group 5) and incubated at 37°C and 55% humidity for 20 days. Then, chicken blood was collected and centrifuged to evaluate alkaline phosphatase (ALP), alanine transaminase (ALT), lactate dehydrogenase (LDH), glucose, urea, and free haemoglobin. Chicken embryo liver was used to evaluate antioxidant capacity (TAC) and chicken embryo spleen was used to evaluate expression of the immune-related genes interleukin-1β (IL-1β), toll-like receptor (TLR)4, TLR2, and TLR15. Semen was randomly divided into 1 control and 8 treatment groups and treated with 50 mg/L Ag-NPs: group A (0.1ppm), group B (1ppm), group C (5ppm), group D (10ppm) or 50 mg/L D-NPs: group E (1ppm), group F (5ppm), group G (10ppm), and group H (20ppm). Sperm viability was analysed using prestoblue metabolic assay. Data were analysed using PROC in GLM procedure of SAS 2014 (SAS Institute Inc.). Decrease in sperm cell viability was recorded in a dose-dependent manner. Sperm cell viability decreased (P&amp;lt;0.005) as the concentration of Ag-NP or D-NP increased. Addition of 100μL of Ag-NPs increased the growth rate of chicken embryo but not 200μL of Ag-NPs or addition of D-NPs. Increases in ALP, ALT, LDH, glucose and urea enzyme were observed in a dose-dependent manner in both Ag-NPs and D-NPs. Addition of 50 mg/L Ag-NPs or 50 mg/L D-NPs increased (P&amp;lt;0.001) TAC of chicken embryo as the volume increased. Additions of 200μL of Ag-NPs, 100μL of D-NPs, and 200μL of D-NPs were haemolytic (P&amp;lt;0.001) but addition of 100μL of Ag-NPs was not. Additions of 100 or 200μL of Ag-NPs or 100μL of D-NPs downregulated IL-1β and 200μL of D-NPs upregulated IL-1β compared with the untreated control group. Additions of 100 or 200μL of Ag-NPs or 200μL of D-NPs induced expression of TLR4 and TLR15. Furthermore, addition of Ag-NPs did not result in expression of TLR2. We concluded that administration of 50 mg/L Ag-NPs and 50 mg/L D-NPs in ovo improve immune status and administration of 100μL of Ag-NPs improved the growth rate of chicken embryo. However, toxicity associated with 50 mg/L Ag-NPs and 50 mg/L D-NPs remains a concern and need to be addressed before use.

Molecular Markers and Phylogenetic Analysis of UPMT27, a Field Isolate of the Malaysian Fowl Adenovirus Associated with Inclusion Body Hepatitis

Inclusion body hepatitis (IBH) is considered one of the re-emerging diseases of avian virus that causes economic damage worldwide. IBH is caused by different serotypes of fowl adenovirus (FAdV), and most of the FAdV cases in Malaysia are related to the serotype 8b. The objective of this study was to determine the molecular markers of UPMT27 Malaysian FAdV isolate and to identify the evolutionary relationship through the phylogenetic approach. Propagation of the isolate was made in embryonated chicken eggs and chicken embryo liver (CEL cells) before it was subjected to viral DNA extraction. Both the fiber and hexon genes of the isolate were amplified and sequenced. The sequences were aligned with the published FAdV sequences. The results showed 100% identity between UPMT27 and the previous Malaysian isolates. A phylogenetic study showed that UPMT27 was closely related to the previous Malaysian isolates. Interestingly, the substitution of the amino acids was consistent between the Malaysia isolates of both fiber protein at positions 72 (Serine –serine), 101 (Alanine -alanine), 125 (Glycine-glycine), and hexon protein 85 (Serine-serine) 160 (Glutamate- glutamate) and 205 (Alanine-alanine) respectively. It appeared that the amino acid variations were the indicators for genetic diversity. Thus, these findings provide information on the evolutionary relationship between FAdV subtypes for IBH prevention.

Pathogenicity and Immunogenicity of Attenuated Fowl Adenovirus from Chicken Embryo Liver Cells in Commercial Broiler Chickens

Pathogenicity and Immunogenicity of Attenuated Fowl Adenovirus from Chicken Embryo Liver Cells in Commercial Broiler Chickens

Organ-specific effects on glycolysis by the dioxin-activated aryl hydrocarbon receptor.

Activation of the aryl hydrocarbon receptor (AHR) by the environmental toxin dioxin (2,3,7,8-tetrachlorodibenzo-p-dioxin, TCDD) causes diverse toxicities, including thymus atrophy and hepatosteatosis. The mechanisms by which AHR activation by TCDD leads to these toxicities are not fully understood. Here we studied the effects of TCDD on a major energy pathway, glycolysis, using the chick embryo close to hatching, a well-established model for studying dioxin toxicity. We showed that 24 hr of TCDD treatment causes changes in glycolysis in both thymus and liver. In thymus glands, TCDD decreased mRNAs for glycolytic genes and glucose transporters, glycolytic indices and levels of IL7 mRNA, phosphorylated AKT (pAKT) and HIF1A, stimulators of glycolysis and promoters of survival and proliferation of thymic lymphocytes. In contrast, in liver, TCDD increased mRNA levels for glycolytic genes and glucose transporters, glycolytic endpoints and pAKT levels. Similarly, increases by TCDD in mRNA levels for glycolytic genes and glucose transporters in human primary hepatocytes showed that effects in chick embryo liver pertain also to human cells. Treatment with the glycolytic inhibitor 2-deoxy-d-glucose exacerbated the effects on thymus atrophy by TCDD, supporting a role for decreased glycolysis in thymus atrophy by TCDD, but did not prevent hepatosteatosis. NAD+ precursors abolished TCDD effects on glycolytic endpoints in both thymus and liver. In summary, we report here that dioxin disrupts glycolysis mediated energy metabolism in both thymus and liver, and that it does so in opposite ways, decreasing it in the thymus and increasing it in the liver. Further, the findings support NAD+ boosting as a strategy against metabolic effects of environmental pollutants such as dioxins.

Isotopic (δ15 N) relationship of pregnant females and their embryos: Comparing placental and yolk-sac viviparous elasmobranchs.

Nitrogen stable isotopes ratios (δ15 N) were determined for selected tissues (muscle, liver, blood and yolk) of pregnant females and their embryos of a placental viviparous species, the Pacific sharpnose shark (Rhizoprionodon longurio), and a yolk-sac viviparous species, the speckled guitarfish (Pseudobatos glaucostigmus). The R. longurio embryo tissues were 15 N enriched compared to the same tissues in the pregnant female, using the difference in δ15 N (Δδ15 N) between embryo and adult. Mean Δδ15 N was 2.17‰ in muscle, 4.39‰ in liver and 0.80‰ in blood. For P. glaucostigmus, embryo liver tissue was significantly 15 N enriched in comparison with liver of the pregnant female (Δδ15 N mean = 1.22‰), whereas embryo muscle was 15 N depleted relative to the muscle of the pregnant female (Δδ15 N mean = -1.22‰). Both species presented a significant positive linear relationship between Δδ15 N and embryo total length (LT ). The results indicated that embryos have different Δδ15 N depending on their reproductive strategy, tissue type analysed and embryo LT .

AEO-7 surfactant is \u201csuper toxic\u201d and induces severe cardiac, liver and locomotion damage in zebrafish embryos

BackgroundFatty alcohol polyoxyethylene ether-7 (AEO-7), a non-ionic surfactant, has recently been receiving extensive attention from the ocean pipeline industry for its ability to inhibit corrosion. However, the present lack of information concerning the potential environmental toxicity of AEO-7, especially towards aquatic organisms, is a major impediment to its wider application. Here, we assess potential adverse effects of AEO-7 on zebrafish embryos employing a variety of assays, including (i) a mortality/survival assay which allowed the median lethal concentration (LC50) to be calculated; (ii) a teratogenicity assay on the basis of which the no observed effect concentration (NOEC) was determined; and (iii) specific assays of cardiotoxicity, neurotoxicity (based on locomotion), hematopoietic toxicity (the level of hemoglobin as revealed by o-dianisidine staining) and hepatotoxicity (liver steatosis and yolk retention examined by staining with Oil Red O).ResultsAEO-7 caused mortality with a calculated LC50 of 15.35 μg/L, which, according to the U.S. Fish and Wildlife Service (USFWS) Acute Toxicity Rating scale, should be considered “super toxic”. Although at its NOEC (0.8 μg/L), there were no signs of significant teratogenicity, cardiotoxicity, or hemopoiesis toxicity, 3.2 µg/L AEO-7 exerted dramatic detrimental effects on organ development.ConclusionOn the basis of these findings, we recommend that the industrial usage and environmental impact of AEO-7 be re-evaluated and strictly monitored by environmental and public health organizations.

Molecular typing of fowl adenovirus associated with gizzard erosion in commercial layer grower chicken in Tamil Nadu

The present study was undertaken to characterize fowl adenovirus associated with commercial layer grower chicken showed gizzard erosion. Ninety four commercial layer grower chicken flocks from Namakkal districts of Tamil Nadu had shown reduced feed intake, reduced weight gain, uneven growth and mortality of 0.3 to 7.7%. On postmortem examination of affected birds showed mild to severe gizzard erosion, blackish discoloration of gizzard contents, pale liver and no major lesions were seen in other organs. Total DNA was extracted and 897 bp fowl adenovirus specific hexon gene was amplified by PCR. Out of 94 flocks screened seven flocks were found positive of fowl adenovirus. Chicken embryo liver cell culture was prepared to isolate field fowl adenovirus from suspected flocks. Concurrent infection of chicken anaemia virus (CAV) was also screened by PCR for 419 bp VP2 gene of CAV and found that all the seven flocks which were PCR positive for FAdV also found positive for CAV. Sequencing and phylogenetic analysis of 897 bp FAdV hexon gene revealed that, it was belonged to FAdV serotypes 2 and 3 of species D.

Bacterial Infection in Chicken Embryos and Consequences of Yolk Sac Constitution for Embryo Survival.

Bacterial infections in chicken eggs often cause mortality of embryos and clinical consequences in chicks but the pathological mechanism is unclear. We investigated the pathological changes and bacterial growth kinetics in dead and live embryos following infection with 2 Escherichia coli strains with a different clinical background and with 1 Salmonella Enteritidis strain. In 2 experiments, 12-day-old embryos were infected via the allantoic sac with 100 µl of 1 to 5 × 102 CFU/ml of one of the bacteria. In experiment 1, only dead embryos were sampled until 4 days postinfection (dpi), and surviving embryos were sampled at 5 dpi. In experiment 2, sampling was performed in dead and killed embryos sequentially at 1, 2, 3, and 4 dpi. The bacteria showed varying pathogenicity in embryos. The yolk sacs of dead embryos showed congestion, inflammation, damaged blood vessels, and abnormal endodermal epithelial cells. Such lesions were absent in the yolk sacs of negative control embryos and in those of embryos that survived infection. The livers and hearts of dead embryos showed congestion and lysed erythrocytes with no morphological changes in hepatocytes or myocardial cells. All bacteria multiplied rapidly in the yolks of infected embryos, although this did not predict survival. However, the livers of dead embryos contained significantly higher bacterial loads than the livers of the embryos that survived infection. The results provide evidence that lesions in the yolk sac, which have been neglected to date, coincide with embryonic mortality, underlining the importance of healthy yolk sacs for embryo survival.

The Tween 80 Toxicity in Chicken Embryos and Effect on the Kinetics of Newcastle Disease Virus Replication

Background: Non-ionic surfactant, Tween-80 (TW80) is commonly used for drug delivery due to its effect on the cell membrane permeation. The change in permeability can also increase viral infectivity in cells. This study was undertaken to improve upon Newcastle disease virus (NDV) titer cultivated with embryonic chicken eggs. Methods: The toxicity of TW80 was investigated against chicken embryos at varying concentrations, and changes in the morphology and weights of the heart, liver, and spleen of 4-day old chicken embryos were analyzed. Also, the effect of non-toxic concentrations of TW80 was examined on the infectivity of NDV. The virus was titrated in the allantoic fluid, using a 50% embryo infectious dose (EID50). Results: At high concentrations of TW80, hemorrhage-induced mortality was observed in embryos at the early stages of incubation. The embryos’ viability was not affected at low TW80 concentrations, indicating that its toxicity to the chicken embryos was dose-dependent. The infectivity titer of NDV was increased in the presence of TW80 compared to those inoculated with NDV only. Conclusion: The data suggest that TW80 is toxic to chicken embryos at high concentrations, but it enhances cell membrane permeability for NDV particles at low concentrations without affecting the embryos’ viability.