An Alternative Approach Using Nano-garlic Emulsion and its Synergy with Antibiotics for Controlling Biofilm-Producing Multidrug-Resistant Salmonella in Chicken.

Surface-growing antibiotic-resistant pathogenic Salmonella is emerging as a global health challenge due to its high economic loss in the poultry industry. Their pathogenesis, increasing antimicrobial resistance, and biofilm formation make them challenging to treat with traditional therapy. The identification of antimicrobial herbal ingredients may provide valuable solutions to solve this problem. Therefore, our aim is to evaluate the potency of nano garlic as the  alternative of choice against multidrug-resistant (MDR) Salmonella isolates using disc diffusion and microdilution assays. Then, checkerboard titration in trays was applied, and FIC was measured to identify the type of interaction between the two antimicrobials. A disc diffusion assay revealed that neomycin was the drug of choice. The range of nano garlic MIC was 12.5-25 µg/ml, while the neomycin MIC range was 32-64 µg/ml. The FIC index established a synergistic association between the two tested drugs in 85% of isolates. An experimental model was used including nano garlic and neomycin alone and in combination against Salmonella infection. The combination therapy significantly improved body productivity and inhibited biofilm formation by more than 50% down regulating the CsgBAD, motB, and sipA operons, which are responsible for curli fimbriae production and biofilm formation in Salmonella serotypes.

Pathognomonic features of Pasteurella multocida isolates among various avian species in Sharkia Governorate, Egypt.

The present study aimed to isolate Pasteurella multocida (P. multocida) from pulmonary cases in several avian species and then investigate the histopathological features, antimicrobial resistance determinants, virulence characteristics, and risk factors analysis of the isolates in each species in correlation with epidemiological mapping of pasteurellosis in Sharkia Governorate, Egypt. The obtained data revealed a total occurrence of 9.4% (30/317) of P. multocida among the examined birds (chickens, ducks, quails, and turkeys). The incidence rate was influenced by avian species, climate, breed, age, clinical signs, and sample type. Antimicrobial susceptibility testing revealed that all isolates were sensitive to florfenicol and enrofloxacin, while 86.6 and 73.3% of the isolates displayed resistance to amoxicillin-clavulanic acid and erythromycin, respectively. All of the P. multocida isolates showed a multiple-drug resistant pattern with an average index of 0.43. Molecular characterization revealed that the oma87, sodA, and ptfA virulence genes were detected in the all examined P. multocida isolates. The ermX (erythromycin), blaROB-1 (ß-lactam), and mcr-1(colistin) resistance genes were present in 60, 46.6, and 40% of the isolates, respectively. Ducks and quails were the most virulent and harbored species of antimicrobial-resistant genes. These results were in parallel with postmortem and histopathological examinations which detected more severe interstitial pneumonia lesions in the trachea and lung, congestion, and cellular infiltration especially in ducks. Epidemiological mapping revealed that the Fakous district was the most susceptible to pasteurellosis infection. Thus, farmers are recommended to monitor their flocks for signs of respiratory disease, seek veterinary care promptly if any birds are sick, and avoid the random usage of antibiotics. In conclusion, this study presents a comprehensive picture of the risk factors in correlation to the pathognomonic characteristics of P. multocida infection in poultry sectors to help in developing more effective strategies for prevention and control.

Molecular and pathological characterization of natural co-infection of poultry farms with the recently emerged Leucocytozoon caulleryi and chicken anemia virus in Egypt.

In the summers of 2018 and 2019, a disease outbreak stroke 25 broiler chicken farms and 3 broiler breeder farms in different Governorates in Egypt. The disease caused a mortality rate ranging from 3.2 to 9%. Postmortem examination showed petechial hemorrhage in the breast and thigh muscles, thymus gland, and peritoneal cavity and extensive hemorrhages in the kidneys. A total of 140 liver, kidney, lung, skeletal muscles, thymus, and spleen samples were collected. Twenty-eight pooled samples were created and examined by PCR and histopathological examination to identify the causative pathogens. All collected samples were PCR-negative to Newcastle disease virus (NDV), avian influenza viruses (H5, H9, and H7), infectious bursal disease virus (IBDV), infectious bronchitis virus (IBV), and fowl adenovirus (FadV). Leucocytozoon caulleryi (L. caulleryi) genetic material was identified by PCR in 17 out of the 28 collected samples (61%). Five chicken farms (18%) showed positive PCR results for both L. caulleryi and chicken anemia virus (CAV). Histopathological examination revealed unilocular megaloschizonts in thymus, skeletal muscle, and lung as well as massive hemorrhages in parenchymatous organs. Nucleotide sequences of the identified pathogens were compared with other reference sequences available in the GenBank. The identified L. caulleryi has a close relationship with those previously detected in Asia, indicating potential transmission route of the parasite. The CAV has a close genetic relation with CAVs previously identified in Egypt. Furthermore, a real-time PCR for rapid, specific, and quasiquantitative detection of L. caulleryi was developed with a detection limit of 100 genome copies per reaction.

Molecular and pathological characterization of duck enteritis virus in Egypt.

In winter 2016, a fatal disease outbreak suspected to be duck virus enteritis (DVE) stroke over a million ducklings in 10 white Pekin and Muscovy ducks flocks in Dakahlia and Gharbia Governorates, Egypt, causing heavy economic losses. The disease quickly killed 20%-60% of affected farms. The clinical signs were inappetence, ataxia, crowding in corners, partially closed eye lids and blue beaks. Post mortem examination revealed white necrotic foci in liver, mottled spleen and sometimes cecal core. A total of 10 intestines, livers and spleens samples were collected from diseased flocks. Each sample was pooled randomly from eight to ten ducklings. Polymerase chain reaction (PCR) and histopathological examination were utilized for DEV identification in collected samples. Nucleotides sequences of the amplified DNA polymerase gene were compared with the other DEVs available on GeneBank. Also, existence of co-infection with Salmonella spp. was verified via PCR. DEV nucleic acid was detected by PCR in 8 of 10 collected samples (80%) with positive amplification of polymerase gene. Histopathological examination revealed eosinophilic and basophilic intranuclear inclusion bodies in enterocytes. In some infected enterocytes, intranuclear and intracytoplasmic inclusions were observed in the same cell. Respectively, eosinophilic intranuclear inclusion bodies found in hepatocytes and reticular cells of liver and spleen of diseased ducklings. Four of the 10 collected samples showed positive results for Salmonella spp. infection that may be involved in enhancing infection with DEV. The identified DEVs revealed close genetic relationship with DEVs detected previously in India and China indicating potential transmission of the virus from there that crucially needs further work for better understanding of virus origin. In conclusion, our study revealed infection of duckling farms with DEV and Salmonella that necessitate the implementation of restricted early preventive and control measures for both diseases to decrease the expected economic losses.