Evaluation of Three Isolation Experiments for Campylobacter Bacteriophages from Chicken Skin: A Comparative Study

Background: Campylobacter strains are of the leading pathogens causing bacterial gastroenteritis, whose infections are generally considered to be one of the most common foodborne illnesses of animal origin. The etiology of this infection often goes back to eating contaminated raw meat or infected poultry. The bacteria are present in abundance in chicken skin. The use of appropriate bacteriophages is one of the most effective experiments in eliminating Campylobacter strains. Phage therapy refers to the use of bacteriophages to treat bacterial infections. Aim: Accordingly, the present study aimed to compare three experiments of bacteriophage isolation in chicken skin. Experiments: Thus, 15 samples of chicken skin were collected from five different fresh chicken suppliers in Ghaemshahr, Iran. The samples were transported to the laboratory aseptically in the vicinity of ice, and then cultured in blood agar medium, and the isolates were identified by various tests including gram staining, catalase and oxidase tests. Results: The results were compared before and after three bacteriophage isolation experiments. Out of 15 chicken skin samples tested in all three experiments, 6 (40%) strains were identified in the first experiment, 8 (53.4%) strains in the second experiment and 12 (20%) strains in the third experiment after bacteriophage therapy. Conclusion: The bacteriophage isolation experiments alone or in combination with other intervention strategies are recommended as promising tools for greater food safety. These experiments can be useful to increase food safety and reduce the risk of infection in humans through the consumption of potentially infected edible parts of chicken. According to the results of this study, among the three proposed experiments, the experiment of chicken skin enrichment in Bolton selective media containing target isolates was the most efficient approach, which showed a high limit of detection at low concentrations and the highest rate of phage recovery. This can be a more reliable way to isolate the Campylobacter bacteriophages and eliminate the Campylobacter strains.

Identification of Chicken Transglutaminase 1 and In Situ Localization of Transglutaminase Activity in Avian Skin and Esophagus

Transglutaminase 1 (TGM1) is a membrane-anchored enzyme that cross-links proteins during terminal differentiation of epidermal and esophageal keratinocytes in mammals. The current genome assembly of the chicken, which is a major model for avian skin biology, does not include an annotated region corresponding to TGM1. To close this gap of knowledge about the genetic control of avian cornification, we analyzed RNA-sequencing reads from organotypic chicken skin and identified TGM1 mRNA. By RT-PCR, we demonstrated that TGM1 is expressed in the skin and esophagus of chickens. The cysteine-rich sequence motif required for palmitoylation and membrane anchorage is conserved in the chicken TGM1 protein, and differentiated chicken keratinocytes display membrane-associated transglutaminase activity. Expression of TGM1 and prominent transglutaminase activity in the esophageal epithelium was also demonstrated in the zebra finch. Altogether, the results of this study indicate that TGM1 is conserved among birds and suggest that chicken keratinocytes may be a useful model for the study of TGM1 in non-mammalian cornification.

The lytic siphophage vB_StyS-LmqsSP1 reduces Salmonella Typhimurium isolates on chicken skin

Phage-based biocontrol of bacteria is considered as a natural approach to combat food-borne pathogens. Salmonella spp. are notifiable and highly prevalent pathogens that cause foodborne diseases globally. In this study, six bacteriophages were isolated and further characterized that infect food-derived Salmonella isolates from different meat sources. The siphovirus VB_StyS-LmqsSP1, which was isolated from a cow´s nasal swab, was further subjected to in-depth characterization. Phage-host interaction investigations in liquid medium showed that vB_StyS-LmqsSP1 can suppress the growth of Salmonella spp. isolates at 37°C for ten hours and reduce the bacterial titer at 4°C significantly. A reduction of 1.4 to 3 log units was observed in investigations with two food-derived Salmonella isolates and one reference strain under cooling conditions using MOIs of 10 4 and 10 5 . Phage application on chicken skin resulted in a reduction of about 2 log units in the tested Salmonella isolates from the first three hours throughout a one-week experiment at cooling temperature and an MOI of 10 5 . The one-step growth curve analysis using vB_StyS-LmqsSP1 demonstrated a 60-min latent period and a burst size of 50-61 PFU/infected cell for all tested hosts. Furthermore, the genome of the phage was determined to be free from genes causing undesired effects. Based on the phenotypic and genotypic properties, LmqsSP1 was assigned as a promising candidate for biocontrol of Salmonella Typhimurium in food. Importance: Salmonella enterica is one of the major global causes of foodborne enteritis in humans. The use of chemical sanitizers for reducing bacterial pathogens in the food chain can result in the spread of bacterial resistance. Targeted and clean label intervention strategies can reduce Salmonella contamination in food. The significance of our research demonstrates the suitability of a bacteriophage (vB_StyS-LmqsSP1) for biocontrol of Salmonella enterica serovar Typhimurium on poultry due to its lytic efficacy under conditions prevailing in food production environments.

Effect of Different Papain Concentrations on the Properties of Chicken Skin Protein Hydrolysates

Chicken skin is a source of animal protein hydrolysate, which has a potential as an antioxidant. This study aimed to determine the effect of different concentrations of the enzyme papain on the degree of hydrolysis, antioxidant capacity, percentage of reducing power (%RP) and amino acid composition of chicken skin protein hydrolysate. Hydrolysis was carried out using papain with various concentrations (3%, 4% and 5%, w/w protein substrate) at pH 7 and 50℃ for 6 h. The degree of hydrolysis was determined by spectrophotometry. Antioxidant capacity and %RP was determined by ferric reducing antioxidant power method and expressed in milligram of ascorbic acid equivalent per gram of sample, and amino acid composition were determined using high-performance liquid chromatography. The results showed that variations in papain concentration had a significant effect (P<0.05) on the degree of hydrolysis, antioxidant capacity and %RP of chicken skin hydrolysate. The papain concentration of 4% resulted in the optimum protein hydrolysate with a degree of hydrolysis of 61.68%±0.64%, an antioxidant capacity of 8.72±0.30 mg AA/g sample and a %RP of 54.12%±1.78%. The protein hydrolysates of the treated chicken skin showed a high content of amino acids, namely, glycine, glutamate, proline, arginine and aspartate.