Wahid Hussein El-Dabae

The quantity of healthy cells in a sample is known as cell viability. For all types of cell culture, measuring cell viability is crucial to ensuring high-quality cell lines through setup, preparation and storage. Although cryopreservation is used in both scientific and clinical research, there are still certain restrictions. At low temperatures, such as −196 °C cells metabolize almost nothing, which has unavoidable side effects, such as genetic drift toward biological variations of cell-associated changes in lipids. Also, cells can be harmed by cryoprotective agents (CPAs), especially when administered in high concentrations So, it is important to use a reliable technique to follow up changes affecting cell viability after cryopreservation which were compared with control freshly subcultured cells. In the present study, the MTT assay was used to correlate cell behavior according to the number of healthy and viable cells. Different kinds of cell cultures were cryopreserved with dimethyl sulfoxide at 10% for variable years in a liquid nitrogen tank . Then, different samples were tested for viability using MTT assay and counted by the trypan blue assay for comparing both results accuracy and confirmed by visual examination in sub-cultured media twenty four hours post-thawing. The MTT assay was easy, fast and showed more accurate results than the trypan blue assay. The visual examination twenty four hours post-subculture indicated that MTT assay can differentiate between healthy and apparently viable cells which have lost their function while the trypan blue assay failed to detect healthy cells in some samples.

| Newcastle disease (ND) is one of the most significant poultry viral diseases threatening the poultry industry worldwide. Despite mass vaccination against ND, genotype VII.1.1 epidemics in vaccinated and non-vaccinated flocks are still recorded, indicating an urgent need to generate a new vaccine candidate. In this study, the live Orthoavulavirus javaense (AOAV-1) genotype VII.1.1 strain (CH-EGY-GIZA-VVTNRC-2021) (Genbank accession number MW603772) was attenuated by combining passaging with limiting 10-fold dilutions in the allantoic cavity of embryonated hens eggs for 20 passages. The attenuated virus was used to vaccinate 20 SPF chicks and its protective efficacy was checked. Intracerebral pathogenicity index (ICPI) of the attenuated strain was 0.2, the mean death time (MDT) was 120 hours and infectvity titer was 7 log10 EID50/ml compared to 1.97 ICPI, 36 hours MDT, and 9.5 log10 EID50/ml of the original strain. The attenuated AOAV-1 was protective in vaccinated chicks by means of HI antibody response and protection from heterologus virulent challenge while showing little to no residual pathogencity. Histopathological inspection showed less severe pathological lesions in vaccinated challenged chickens than in non-vaccinated challenged chickens. Quantitation of viral loads following challenge indicated reduction of viral loads shed from birds vaccinated with the attenuated AOAV-1 virus. These preliminary insights collectively underline a new hope for the attenuation of live AOAV-1 genotype VII.1.1 as a vaccine candidate that can be used to control ND outbreaks in Egyptian poultry.

Newcastle disease (NDV) and infectious bronchitis (IB) viruses are the most infectious and serious diseases on poultry flocks Egypt. Several types of bivalent inactivated NDV and IB vaccines are used to prevent and decrease incidence of both viruses but efficacy of these vaccines should be monitored periodically to understand the main causes of those breaks. In the current study nine bivalent inactivated NDV and IB vaccines were randomly selected from different batches during routine work at central laboratory for evaluation of veterinary biologics in 2023. Quality control of nine vaccines was applied according to Egyptian standard regulations for evaluation of veterinary Biologics. Completion of inactivation test of nine vaccines for both NDV and IB viruses revealed negative haemaagglutination test for NDV and normal appearance of embryo of IB virus when inoculation in SPF eggs. Immunization of one week old SPF chickens of nine selected vaccines showed neither clinical signs nor mortalities. Collection of serum samples from all tested vaccines on 3rd and 4th weeks post vaccination showed variable immune response against NDV using Haemagglutination inhibition test and ELISA test against IB virus. Challenge trial of tested vaccines showed variable percent of protection (50% to 100%). Virus shedding in SPF eggs on 3rd, 5th, 7th and 10th days post challenge revealed significant amount of virus in oropharengeal swabs. This study showed validation of some bivalent inactivated NDV and IB vaccines batches in vaccination programs and invalidation of other batches for use.

Background and Aim: Mastitis is considered a significant disease of lactating animals. There are new attitudes for recognizing genes responsible for causing this disease to overcome and change the manipulation of this problem. This study aimed to isolate and identify Staphylococcus aureus strains from mastitic bovine animals and detect some specific biofilm-forming genes (icaA, icaD, and biofilm-associated protein [bap] genes clfA, fnbA, agrI, agrII, agrIII, agrIV, and cna). Materials and Methods: A total of 121 mastitic milk samples were analyzed using biochemical tests (catalase test, oxidative-fermentative test, and coagulase test) and Gram stain. Multiplex polymerase chain reaction was applied to characterize biofilm genes (icaA, icaD, bap, clfA, and fnbA) in addition to (agrI, agrII, agrIII, agrIV, and cna). Results: Among the 121 milk samples, 35 staphylococci isolates were derived with an incidence of 28.92% (35/121); among them, 19 are coagulase positive. Ninety percent of the isolates had ica genes (icaA and icaD) while bap gene was not recognized in any isolate. In addition, the incidence of fnbA, can, and clfA was 89.5% each. The prevalence of agr specific groups (agrI, agrII, agrIII, and agrIV) was 78.9%, 52.6%, 10.5%, and 15.8%, respectively. Conclusion: This study concluded that S. aureus has variant mechanisms of pathogenicity to form biofilm devoid of carrying a specific gene.

Aim: This study aimed to detect and characterize current genotypes of canine parvovirus (CPV) in Egypt during 2018. Materials and Methods: A total of 50 fecal swabs were collected from clinically infected domestic dogs of 2-5 months of age, suspected to suffer from CPV infection, from Cairo and Giza Governorates. The samples were subjected to qualitative antigen detection using the rapid test, followed by isolation on Madin-Darby Canine Kidney (MDCK) cells, molecular characterization with partial amplification of VP2 gene using polymerase chain reaction (PCR), followed by sequencing and phylogenetic analysis. Results: Out of 50 fecal samples, 20 samples were positive (40%) by Rapid CPV/canine coronavirus Ag Test Kit. These positive samples were cultured successfully on MDCK cells. Nine randomly chosen samples out of 30 apparently negative samples were amplified using PCR with primers Hfor and Hrev to yield a typical 630 bp fragment. Then, six randomly chosen samples out of nine were amplified using PCR with primers Pbs and Pbas to yield a typical 427 bp fragment. Sequencing, BLAST analysis and assembly of the two fragments (630 bp and 427 bp) to produce 912 bp fragments, in the six samples, revealed two serotypes CPV-2b and CPV-2c. The obtained strains were submitted to GenBank and given accession numbers MK642272, MK642273, MK642274, MK642275, MK642276, and MK642277. Phylogenetic analysis of the Egyptian strains serotype 2b illustrated that they were closely related to Thailand strains (accession numbers KP715709, KP715694, KP715701, and KP715700); while Egyptian strains serotype 2c was closely related to Thailand strains (accession numbers MH711894 and MH711902), Taiwanese strain (KU244254), Chinese strain (MF467242), and Vietnamese strain (accession number LC216910). Conclusion: The current research recommends further epidemiological studies to assess the extent of the occurrence of different serotypes of CPV in Egypt and the efficiency of imported and locally produced vaccines in protection against CPV infection.