Salmonella species cause a significant worldwide burden of disease, morbidity, and mortality. Infections with Salmonella species in poultry and humans cause multiple clinical symptoms.  Consequently, reliable information on the occurrence and prevalence of the disease is necessary to establish appropriate intervention methods to control Salmonella populations at the farm level. The current study aims to distinguish and identify different species of Salmonella from hatching egg chicks and explore the antimicrobial resistance pattern of the isolates. The one-day-old specific pathogen-free (SPF) chicks were used to in vivo assess the pathogenicity of the isolated strains of Salmonella. The sick chicks were obtained from localities, chicken farms, and egg batches. The species-specific multiplex PCR used to amplify the inv-A, IE-1, flic-C target genes for Salmonella genus, S. enterica serovar Enteritidis, and Salmonella enterica serovar Typhimurium, respectively, and the phenotypic characteristics of the isolated bacteria were confirmed. Seven multi-drug resistant (MDR) isolates from poultry farms, along with isolates from the hatching egg batches, were verified as Salmonella enterica. Despite Salmonella enterica having little variation in its phenotypic composition, eight of the nine detected strains of Salmonella had MDR strains, most of which were highly prevalent and had variable mortality rates. Interestingly, in S. enterica strains, the lowest MDR indices were associated with high virulence mortality (> 85%) and vice versa. The results showed a predominant Salmonella Enteritidis in the different farms chosen in Egypt. Therefore, uncontrolled use of antibiotics in chicken farms may be the main reason for the increased incidence of MDR Salmonella spp., which could hinder attempts to control Salmonella and jeopardize public health. Long-term antibiotic use in livestock farming leads to antibiotic residues in animal-producing foods, which can cause toxicity and low amounts of antibiotic exposure could alter the microbiota and lead to antibiotic resistance. This study found that in vivo pathogenicity in SPF chicks increased with decreasing MDR index.