Molecular characterization of the whole genome of H9N2 avian influenza virus isolated from Egyptian poultry farms.

H9N2 avian influenza viruses (AIVs) affect both poultry and humans on a global level, and they are especially prevalent in Egypt. In this study, we sequenced the entire genome of AIV H9N2 isolated from chickens in Egypt in 2021, using next-generation sequencing (NGS) technology. Phylogenetic analysis of the resulting sequences showed that the studied strain was generally monophyletic and grouped within the G1 sublineage of the Eurasian lineage. Four segments (polymerase basic 2 [PB2], polymerase basic 1 [PB1], polymerase acidic [PA], and non-structural [NS]) were related to Egyptian genotype II, while the nucleoprotein (NP), neuraminidase (NA), matrix (M), and haemagglutinin (HA) segments were related to Egyptian genotype I. Molecular analysis revealed that HA protein contained amino acid residues (191H and 234L) that suggested a predilection for attaching to human-like receptors. The antigenic sites of HA had two nonsynonymous mutations: V194I at antigenic site A and M40K at antigenic site B. Furthermore, the R403W and S372A mutations, which have been observed in H3N2 and H2N2 strains that caused human pandemics, were found in the NA protein of the detected strain. The internal proteins contained virulence markers: 504V in the PB2 protein, 622G, 436Y, 207K, and 677T in the PB1 protein, 127V, 550L, and 672L in PA protein, and 64F and 69P in the M protein. These results show that the detected strain had undergone intrasubtype reassortment. Furthermore, it contains changes in the viral proteins that make it more likely to be virulent, raising a question about the tendency of AIV H9N2 to become highly pathogenic in the future for both poultry and humans.

Assessment of mitochondrial DNA viability ratio in day-4 biopsied embryos as an add-in to select euploid embryos for single embryo transfer.

The aim of this study was to assess mitochondrial DNA analysis as a predictor of the pregnancy potential of biopsied preimplantation embryos. The study included 78 blastomeres biopsied from day 4 cleavage stage euploid embryos. The embryo karyotype was confirmed by 24-chromosome preimplantation genetic testing for aneuploidies using the Illumina Next-Generation Sequencing (NGS) system. Mitochondria viability ratios (mtV) were determined from BAM files subjected to the web-based genome-analysis tool Galaxy. From this cohort of patients, 30.4% of patients (n = 34) failed to establish pregnancy. The mean mtV ratio [mean = 1.51 ± 1.25-1.77 (95% CI)] for this group was significantly (P < 0.01) lower compared with the embryo population that resulted in established pregnancies [mean = 2.5 ± 1.82-2.68 (95% CI)]. mtV multiple of mean (MoM) values were similarly significantly (P < 0.01) lower in blastocysts failing to establish pregnancy. At a 0.5 MoM cut-off, the sensitivity of mtV quantitation was 35.3% and specificity was 78.2%. The positive predictive value for an mtV value > 0.5 MoM was 41.4%. This study demonstrates the clinical utility of preimplantation quantification of viable mitochondrial DNA in biopsied blastomeres as a prognosticator of pregnancy potential.

Therapeutic effects of astragaloside IV and Astragalus spinosus saponins against bisphenol A-induced neurotoxicity and DNA damage in rats.

BACKGROUND: Bisphenol A (BPA) is an endocrine disruptor to which humans are often subjected during daily life. This study aimed to investigate the ameliorative effect of astragaloside IV (ASIV) or saponins extracted from Astragalus spinosus (A. spinosus) against DNA damage and neurotoxic effects induced by BPA in prefrontal cortex (PFC), hippocampal and striatal brain regions of developing male rats. MATERIALS AND METHODS: Juvenile PND20 (pre-weaning; age of 20 days) male Sprague Dawley rats were randomly and equally divided into four groups: control, BPA, BPA+ASIV and BPA+A. spinosus saponins groups. Bisphenol A (125 mg/kg/day) was administrated orally to male rats from day 20 (BPA group) and along with ASIV (80 mg/kg/day) (BPA+ASIV group) or A. spinosus saponin (100 mg/kg/day) (BPA+ A. spinosus saponins group) from day 50 to adult age day 117. RESULTS: Increased level of nitric oxide (NO) and decreased level of glutamate (Glu), glutamine (Gln), glutaminase (GA) and glutamine synthetase (GS) were observed in the brain regions of BPA treated rats compared with the control. On the other hand, co-administration of ASIV or A. spinosus saponin with BPA considerably improved levels of these neurochemicals. The current study also revealed restoration of the level of brain derived neurotrophic factor (BDNF) and N-methyl-D-aspartate receptors (NR2A and NR2B) gene expression in BPA+ ASIV and BPA+A. spinosus saponins groups. The co-treatment of BPA group with ASIV or A. spinosus saponin significantly reduced the values of comet parameters as well as the intensity of estrogen receptors (ERs) immunoreactive cells and improved the histological alterations induced by BPA in different brain regions. CONCLUSION: It could be concluded that ASIV or A. spinosus saponins has a promising role in modulating the neurotoxicity and DNA damage elicited by BPA.

Anti-anxiety and antidepressant-like effects of astragaloside IV and saponins extracted from Astragalus spinosus against the bisphenol A-induced motor and cognitive impairments in a postnatal rat model of schizophrenia.

Bisphenol A (BPA) is a chemical endocrine disruptor to which humans are often exposed in daily life. Postnatal administration of BPA results in schizophrenia (SCZ)-like behaviours in rats. The present study was designed to elucidate whether treatment with astragaloside IV (ASIV) or saponins extracted from Astragalus spinosus improves the neurobehavioural and neurochemical disturbances induced by BPA. Fifty-two juvenile (PND20) male Sprague Dawley rats were divided into four groups. The rats in Group I were considered the control rats, while the rats in Group II were orally administered BPA (125 mg/kg) daily from PND20 to adult age (PND117). The rats in the third and fourth groups were administered BPA (125 mg/kg/day) supplemented with astragaloside IV (80 mg/kg/d) on PND20 or A. spinosus saponins (100 mg/kg/d) from PND50 to PND117, respectively. Administration of ASIV and saponins extracted from Astragalus spinosus reversed the anxiogenic and depressive-like behaviours and the social defects that were observed in the rats treated with BPA alone. Additionally, these compounds improved memory impairments, restored dopamine (DA), serotonin (5-HT), and monoamine oxidase (MAO-A) levels and normalized Tph2 mRNA expression towards the control values. Taken together, it can be concluded that orally administered ASIV and A. spinosus saponins exhibit neuroprotective effects and that these compounds can be used as therapeutic strategies against BPA-induced neuropsychiatric symptoms in a rat model of SCZ.

Role of diagnostic intracytoplasmic sperm injection (ICSI) in the management of genetically determined zona pellucida-free oocytes during in vitro fertilization: a case report.

PURPOSE: To report the utilization of diagnostic intracytoplasmic sperm injection (D-ICSI), an ICSI cycle performed in the natural cycle, to obtain information about embryo development potential after sperm injection into zona pellucida (ZP)-free oocytes. MATERIALS AND METHODS: We report the case of a couple with primary unexplained infertility with a history of previous failed, in vitro fertilization intracytoplasmic sperm injection (IVF-ICSI) cycles characterized by the presence of ZP-free oocytes. Whole exome sequencing (WES) was carried out to analyse the possible genetic basis of oocyte abnormality. RESULTS: Diagnostic ICSI provided information about the embryo development potential from ZP-free oocytes and allowed better planning of the subsequent ICSI cycle. WES revealed that the absence of ZP was likely to be due to a new (ZP1) mutation. The subsequent ICSI cycle resulted in the delivery of a healthy baby. DISCUSSION: To the best of our knowledge, our report is the first to describe the use of D-ICSI to determine the feasibility of embryo development and implantation in a patient with ZP1 mutation, resulting in the subsequent delivery of a healthy baby. We used 'diagnostic' ICSI in the normal menstrual cycle to explore the feasibility of embryo development after sperm injection into ZP-free oocytes. Our results may expand the spectrum of diagnostic procedures associated with unexplained infertility.

Composition of bacterial and archaeal communities in the rumen of dromedary camel using cDNA-amplicon sequencing.

The camel is known to survive in harsh environmental conditions, due to its higher digestive efficiency of high-fiber diets compared with other ruminants. However, limited data are available on the microbial community in the rumen of a camel. In this study, the Illumina sequencing of V4 region of 16S rRNA genes based on RNA isolation was employed to get insight into the bacterial and archaeal communities associated with liquid and solid rumen fractions in eight camels under different feeding systems. Camels in group C1 were fed Egyptian clover hay plus concentrates mixture and camels of group C2 were fed fresh Egyptian clover. The results showed that liquid fraction has higher operational taxonomic units (OTUs) than solid fraction, and camel group C1 showed a higher microbial diversity than C2. The UniFrac analysis indicated that the microbial communities in camel groups are distinct. Moreover, phylum Firmicutes and Bacteroidetes dominated the bacterial community and Candidatus Methanomethylophilus dominated the archaeal community with a significant difference in the relative abundance between camel groups. Dominant bacterial genera were Prevotella, Fibrobacteres, Ruminococcus, and Butyrivibrio. There were many negative and positive correlations between and within bacterial and archaeal genera. The composition of microbial community in the rumen of a camel is similar to other ruminants with differences in the abundance.

Community structure and fibrolytic activities of anaerobic rumen fungi in dromedary camels.

Anaerobic fungi colonize the rumen and degrade cellulose and hemicellulose, which enable them to be key players in the lignocellulose fermentation. Consequently, an expansion of knowledge about rumen fungi could increase animal productivity, utilization of lignified forages like alfalfa hay, and enhance fibrolytic enzymes production. Here, we used an Internal Transcribed Spacer 1 (ITS1) clone library to investigate the anaerobic rumen fungi in camel and to investigate their ability to produce cellulase and xylanase in vitro. Rumen fluid was collected from camels fed Egyptian clover (n = 14), and wheat straw (n = 7) and fecal samples were collected from camels fed wheat straw and concentrates (n = 5), or natural grazing plants (n = 10). Neocallimastix and Cyllamyces were the most abundant anaerobic fungi in all camel groups. An anaerobic rumen fungi media containing alfalfa hay as a carbon source was inoculated by rumen and fecal samples to assess the ability of anaerobic rumen fungi in camel gut to produce cellulase and xylanase. The anaerobic gut fungi in the camel is diverse and has cellulolytic and xylanolytic activities, fungal culture from rumen samples of camel fed wheat straw (R2) exhibited highest cellulase production. In addition, many of the sequences in the current study have no equivalent cultured representative, indicating a novel diversity within the camel gut.

Molecular displaying of differential immunoresponse to various infections of amniotic epithelia.

PROBLEM: Molecular displaying for the interaction of innate immune cells against pathogen is important for knowing their defense mechanism. This study aimed to visualize the differential gene expression profiles of amniotic epithelium, in response to various infections, using simple reverse transcription polymerase chain reaction (RT-PCR) techniques. METHOD OF STUDY: Antimicrobial activity of human amniotic epithelial cells (HAECs) was verified against three different models of pathogenic microorganisms. RNA was extracted from infected and non-infected cells. Transcripts were visualized by two methods of RT-PCR; one of them targets ß-defensin 2 (BDEF2). The other is a novel design method of RNA fingerprinting based on differential length amplification of transcripts (DifLAT) to polymorphism more than 69 coding sequences for antimicrobial proteins. RESULTS: The semiquantitative gel analysis indicated that BDEF2 was upregulated during all infections. DifLAT experiment visualized different patterns of HAECs transcripts for each case of infection. CONCLUSION: This study proved that HAECs uses alternative gene expression profiles for fighting different pathogens.

Association between IL-10 gene promoter polymorphism and hepatitis B viral infection in an Egyptian population.

Cytokines play critical roles in the pathogenesis of hepatitis B virus infection (HBV). This work was designed to study the effect of IL-10 gene polymorphisms (-1082G/A and -819C/T) on susceptibility of Egyptians to HBV. Genotyping was performed using single-stranded polymorphism-polymerase chain reaction in 118 Egyptian hepatitis B patients and 119 healthy controls, and IL-10 serum levels were measured using ELISA. The frequency of IL-10 -1082G/G was significantly higher in HBV patients than in healthy controls, and G/A and A/A were not significantly different between groups. The distribution of IL-10 -819 genotypes was not significantly different between the HBV and healthy control groups. Although AT was significantly different between controls and patients, the distribution of the other haplotypes was not. IL-10 levels were significantly lower among hepatitis B patients. Our data stress the importance of IL-10 gene polymorphism in HBV infection. Depending on our preliminary work, IL-10 -1082G/G may act as a host genetic factor in the susceptibility to HBV infection in Egyptians.

Transforming Growth Factor- ß 1 Gene Polymorphism (T29C) in Egyptian Patients with Hepatitis B Virus Infection: A Preliminary Study.

The interindividual variations in the capacity of transforming growth factor- ß 1 (TGF- ß 1) production have been ascribed to genetic polymorphisms in TGF- ß 1 gene. As pathogenesis of HBV has a genetic background, this preliminary study was designed to assess the impact of TGF- ß 1 (T29C) on the susceptibility of Egyptians to HBV infection. Genotyping was performed using single stranded polymorphism-polymerase chain reaction (SSP-PCR) in 65 Egyptian hepatitis B patients and 50 healthy controls. TGF- ß 1 plasma levels were measured using Enzyme-linked immunosorbent assay (ELISA). The frequency of CC genotype was significantly higher (P < 0.05) in HBV patients compared to controls. On the contrary, TC genotype did not show significant difference in both groups. TT genotype was significantly higher (P < 0.01) in controls than HBV patients. Our current preliminary data revealed that the frequency of the genotypes in the controls were within Hardy-Weinberg equilibrium (HWE) while the patients group was out of HWE (P < 0.01). TGF- ß 1 was significantly (r = -0.684; P < 0.001) deceased in the sera of patients as compared to normal subjects. Depending on our preliminary work, CC genotype may act as a host genetic factor in the susceptibility to HBV infection in Egyptians. Taken together, the current data pointed to the importance of polymorphism of TGF- ß 1 gene (T29C) in HBV infection.

Establishment and characterization of Spodoptera littoralis cell line (SL 96), adapted at 19 degrees C permissive for different baculoviruses.

Cells from ovarioule tissues of the Egyptian cotton leaf worm, Spodoptera littoralis (SL 96) were established, identified and characterized comparing with other lepidopteran cell lines using isozymes, karyotyping, RAPD PCR, and polyacrylamide gel electrophoresis polypeptide PAGE profile. Comparison of the number of chromosomes and PAGE profiles; also the isozymes profiles demonstrate the possibility to distinguish the cell lines by those analyses and having a distinguishable finger print for every cell line. A complete replication of the S. littoralis granulovirus (SpliGV), nucleopolyherovirus (SpliNPV), and Phthorimaea operculella granulovirus (PhopGV) was obtained in vitro by both viral infection and DNA transfection in the S. littoralis (S196) cell line. Spli-GV and PhopGV multiplied extensively during several passages in S196 cells at 19 degrees C. Virus infected cells were incubated at 19 degrees C for the first 4 hours and then at .27 degrees C for the rest of the experiment (20 days). Generally, all the three viral multiplications proceeded at the same rate approximately. Comparison of Spli GV progenies multiplied either in vivo or in vitro, using electron microscopy and restriction profile analysis, showed their identity. Viral infections were detected also by specific prepared nucleic probes.