Molecular characterization and immunohistochemical localization of tilapia piscidin 3 in response to Aeromonas hydrophila infection in Nile tilapia.

The antimicrobial activity of tilapia piscidin 3 (TP3) was determined in vitro against a locally isolated Aeromonas hydrophila. A 388 bp fragment was amplified from the TP3 cDNA and sequenced. The coding sequence (CDS) of TP3 was estimated to be 231 bp codes for 76 amino acids long and stop codon. In silico analysis was performed to detect both the signal peptide and the prodomain cleavage sites to follow the amino acids number 22 and 70, respectively. Based on this, a peptide 23 amino acids long with a remarkably high computed antimicrobial probability was synthesized and used in the subsequent experiments. The antimicrobial activity of TP3 was determined with minimum inhibitory concentration (MIC) and minim um bactericidal concentration (MBC) methods. TP3 exhibited relatively weak antimicrobial activities against the tested bacteria. A challenge experiment was then performed in Nile tilapia with low and high doses of A. hydrophila, followed by timely recognition; after 3, 6, 24 h, and 7 days of the specific TP3 gene expression, immunohistochemical localization was also performed. Histopathological examination revealed provoked inflammatory responses and congestion in the same organs of TP3 expression. Immunohistochemical localization showed that A. hydrophila induced tilapia fish to express TP3 after 24 h within the gills, intestine, hepatopancreas, spleen, and posterior kidney. In quantitative real time (RT)-polymerase chain reaction analysis, the high dose showed higher mRNA expression levels than the low dose, and its expression levels increased in the A. hydrophila-infected fish. It was therefore concluded that TP3 plays an essential role in fish immunity.

Genetic similarity and diversity among three camel populations reared in Egypt.

BACKGROUND: Molecular genetics has been extremely useful in determining the relation between animal populations and documenting the degrees of genetic variation found within them. The present study was undertaken to evaluate genetic diversity and the relationships between the three camel populations reared in Egypt: Maghrabi, Sudani, and Baladi using mitochondrial 16S sequences and other breeds of camels in the world. METHODS: Blood samples were collected from camels belonging to these three populations. Genomic DNA was extracted from the collected blood samples and subjected to PCR using specific primers for mitochondrial 16S region. The amplified products were purified using DNA purification kit to remove residual primers and dNTPs. Sequencing was performed in the Macrogen Incorporation. The amplified products were submitted to GenBank/NCBI under accession numbers OM 278349 and OM 278350 RESULTS: Sequencing was done on the partial mitochondrial 16S amplified fragments at 530 bp. This amplified area had two haplotypes. There was one substitution (G/A) at nucleotide 309 of the amplified segment. The nucleotide (π) and Hd stand for haplotype diversity, respectively, at 0.00008 and 0.042, and the average number of pairwise nucleotide differences, k, is 0.042, according to Fu's Fs statistic and Tajima's D, which is -1.10686. Genetic distance percentages between the three populations under study range from 0.000 to 0.0312. A phylogenetic analysis of Egyptian camel populations and other Camelus dromedarius populations revealed a strong relationship between them. CONCLUSIONS: This study suggests that the 16S rRNA sequencing in mitochondria plays a critical role in genetic variation studies and analysis of phylogeny between camel populations and breeds.

Piscidin 4: Genetic expression and comparative immunolocalization in Nile tilapia (Oreochromis niloticus) following challenge using different local bacterial strains.

The antimicrobial activity of tilapia piscidin 4 (TP4) was determined in vitro against four bacterial strains, Aeromonas hydrophilla, Pseudomonas fluorescens, Streptococcus iniae and Vibrio anguillarum. Nile tilapia were infected with low and high doses of the tested pathogens; after 3, 6, 24 h and 7 days of the specific TP4 gene expression, tissue immunolocalization was also performed. Histopathological examination revealed septicaemia and necrosis of hemopoietic tissue for all of the tested bacteria. Immunolocalization showed abundance in S. iniae-infected fish tissues. Quantitative RT-PCR analysis revealed that high doses raised mRNA expression levels compared to low doses and expression levels increased in the infected fish, particularly after 24 h, indicating that TP4 exerts potent bactericidal activity against some fish pathogens and plays an essential role in fish immunity.

Evaluation of the alleviative role of Chlorella vulgaris and Spirulina platensis extract against ovarian dysfunctions induced by monosodium glutamate in mice.

Microalgae provide a wealthy natural resource of bioactive compounds, which have many biological activities. Monosodium glutamate is a food additive that acts either as food preservatives or as tastiness enhancer. It was confirmed that monosodium glutamate poses a serious responsibility in the pathogenesis of anovulatory infertility. Therefore, the idea of this research was directed to reveal efficiency of Chlorella vulgaris and Spirulina platensis extracts against the ovarian dysfunction resulted due to monosodium glutamate consumption. Adult female albino mice were gavages orally monosodium glutamate alone or with either Chlorella vulgaris or Spirulina platensis aqueous extracts for 28 days. Female mice were subjected to superovulation to study the oocytes nuclear maturation stages. Histological and quantitative investigation was carried on ovaries. Biochemical assessment to measure the sex hormones level and ovarian enzymatic antioxidants was done. In addition, ovarian antioxidant mRNA genes were determined using quantitative PCR and Glyceraldehyde-3-phosphate dehydrogenase was used as an internal control. The result revealed that monosodium glutamate reduced the oocytes quality and maturation rate, while, both algae improve the oocyte quality and maturation rate than in monosodium glutamate group. Chlorella vulgaris and Spirulina platensis improved the monosodium glutamate ovarian tissue histological alteration, sex hormones content and raised the ovarian enzymatic antioxidants level. In addition, monosodium glutamate markedly diminished the Glutathione peroxidase, superoxide dismutase and catalase mRNA expressions, However, Chlorella vulgaris or Spirulina platensis upregulated the expression of genes close to control. In conclusion, Chlorella vulgaris and Spirulina platensis showed potential alleviative role against the monosodium glutamate ovarian dysfunction.

Cytochrome b conservation between six camel breeds reared in Egypt.

This study was aimed to assess cytochrome b conservation in six breeds of camels reared in Egypt and to compare its sequence with those of other livestock species. The 208-bp fragments from camel mtDNA cyto b were amplified using PCR for 54 camels belonging to 6 camel breeds reared in Egypt. The alignment of camel cyto b sequences showed the presence of two polymorphic sites resulting in four haplotypes and their nucleotide sequences were submitted to GenBank under the accession numbers: KX909894-KX909897. The genetic distances between tested camel breeds were zero between Baladi, Fallahi and Maghrabi breeds whereas they were at low value between the other three breeds: Mowaled, Sodany and Somali. Neighbor-joining showed 4 branches; one of them include most of the tested animals and another one contains 2 Somali animals which is considered a specific haplotype for this breed. The other two branches are mixed between Sodani and Mowaled breeds. Neighbor-joining tree was constructed between cyto b sequences of our tested camels and their sequences from livestock species include Camelus dromedaries, Camelus bactrianus, Ovis aries, Capra hircus, Bubalus bubalis, Bos Taurus and Sus scrofa. The result confirmed that our camel breeds belong to Camelus dromedaries and are clearly separated from other species. It is concluded that cyto b sequence is highly conserved among all camel breeds reared in Egypt which belong to Camelus dromedaries in addition to the advantage of cyto b in differentiation between different livestock sources which enables it to widely use for the adulteration detection in mixed meat.