Prebiotic supplementation effect on Escherichia coli and Salmonella species associated with experimentally induced intestinal coccidiosis in rabbits

Introduction

Nowadays, the antibiotic resistance has emerged as the greatest challenge in the animal production and human health. The extensive use of antibiotics as growth promoters for livestock is the major cause of antibiotic resistance (World Health Organization & Communicable Diseases Cluster, 2000; Millman et al., 2013; Mitchell et al., 2013). Rabbit is a good source of animal protein to fill the gap of the red meat shortage in some parts of the world, and sometimes reared for fur production as well as for medical and biological purposes (Dalle Zotte & Szendro, 2011; Aboelhadid et al., 2019). Rabbit coccidiosis is caused by apicomplexan parasites of the genus Eimeria (Pakandl, 2009). Coccidiosis is mainly occurring in young rabbits of one to three months’ age especially after weaning. The clinical signs of coccidiosis are: reduced appetite, enteritis, diarrhea, and in severe cases infection may result in death (El-Shahawi, El-Fayomi, Abdel-Haleem, 2011)). There are two types of rabbit coccidiosis; intestinal coccidiosis which caused by several species including E. intestinalis, E. perforans, E. magna, E. media, and E. irresidua and, hepatic coccidiosis which caused by only E. steidae (Pakandl, 2009). Rashwan & Marai (2000) postulated that the coccidian infection may enhance the proliferation of the gut Enterobacteriaceae. Bacterial infections in rabbits can negatively affect the body condition and cause high mortality, especially in young ages (Zahraei, Mahzouniehand & Khaksar, 2010). Infections with Enterobacteriaceae are more challenging to treat, because few, and in some cases no, antimicrobials remain effective against them, because of their extensive resistance patterns and in addition the antibiotic chemical residue in animal products may create problems for human wellbeing (Smith et al., 2002).

Salmonella species were reported to infect rabbits in several rabbitries and the infection can lead to severely diseased condition (Agnoletti et al., 1999; Zahraei, Mahzouniehand & Khaksar, 2010; Borrelli et al., 2011).

E scherichia coli is an important cause of diarrhea in both animals and humans (Nguyen et al., 2006; García, Fox & Besser, 2010). Also, it was reported to cause morbidity and mortality in large laboratory rabbits (Swennes et al., 2012; Swennes et al., 2014). E scherichia coli was known as a reason for diarrhea in new born of New Zealand rabbits (Camarda et al., 2012; Hamed, Eid & El-Bakrey, 2013). Prescott (1978) found that the outbreak of severe diarrhea and death in young rabbits was associated with non-enterotoxigenic Escherichia coli (O153).

Recently, the prebiotics were defined as “a substrate that is selectively utilized by host microorganisms conferring a health benefit” (Gibson et al., 2017). The prebiotics administration could regulate specific gastrointestinal tract microorganisms to modify the microbiome (Gibson & Roberfroid, 1995). Abdelhady & El-Abasy (2015) found that the prebiotic and probiotic as dietary supplementation reduced the mortality rate and improved the adverse clinical signs of Pasterella multocida in experimentally infected rabbits. Tzortzis et al. (2005) realized that the oligosaccharides greatly inhibited the adhesion of E. coli and Salmonella to HT29 cells. Also, Yusrizal & Chen (2003) revealed that fructans supplementation induced an increase in Lactobacillus bacteria and a reduction in Salmonella in the broiler chickens. Moreover, it was noticed that prebiotics intake reduced the establishment of Salmonella in the course of hen molting (Donalson et al., 2008). Spring et al. (2015) demonstrated that Bio-Mos^®, which has been used in the animal husbandry industry, plays a crucial role in animal nutrition and production. It was extracted from a selected strain of Saccharomyces cerevisiae yeast. Bio-Mos is inserted into animal diets to support overall animal performance and rapid growth. It is supported by over 734 trials and 114 peer-reviewed publications. There is considerable evidence now that Bio-Mos is among the best alternatives to antibiotic and growth promotants (Ferket, Parks & Grimes, 2002).

The present study was therefore conducted to explore the effect of a prebiotic supplement as a prophylaxis on the presence of E. coli and Salmonella species in rabbits with experimentally induced intestinal coccidiosis.

Materials and Methods

This experiment was conducted under the roles of the ethical standards approved by Faculty of Veterinary Medicine, Beni-Suef University, Egypt and its specific approval number was (BSUV-39/2019).

Results

Prebiotic prophylaxis effect on the oocyst count and prevalence of both E. coli and Salmonellaspecies at day 5 and day 8 post infection

The oocyst excretion in the feces of infected rabbits began at the day 5^t PI in PS and PC groups. Also, the OPG was significantly reduced in the PS group (2.51 × 10⁴ ± 10.36) when compared with the PC group (4.33 × 10⁴ ±  11.52) (Table 1). This significant (p ≤ 0.05) decrease in the oocyst count in the PS group continued till the end of the experiment (day 8 PI). Eleven E. coli isolates were detected during the present study with overall prevalence of 24.4%. These eleven isolates were three serotypes in NC with prevalence of 20%, six serotypes PC with prevalence of 40% and two serotypes in PS with a prevalence of 13.33% (Table 2). The highest prevalence of E. coli was in the PC group while the lowest one was in the PS group. Meanwhile, four salmonella serovars were isolated with overall prevalence of 8.89%. The NC group showed only one serovars with a prevalence of 2.22% and the PC group revealed three serovars with a prevalence of 6.67% while prebiotic supplemented group didn’t show any salmonella isolates (Table 2). The same results of E. coli and Salmonella prevalence were recorded at day 8 PI.

Discussion

The use of antibiotics as growth promoters in animals was banned by the European Union Commission (European Union Commission, 2005), and since then, prebiotics and probiotics were actively investigated as safe natural alternatives to the antibiotics. Prebiotics were known to have valuable effects on the improvement of the host immune system, productivity and performance in addition to its bactericidal/bacteriostatic activities. The prebiotics were also used as growth promoters in the form of feed additives to increase growth of chickens (Ashayerizadeh et al., 2009). It was found that coccidial infection in most cases was associated with secondary bacterial and viral infections that may lead to mortality in infected host (Taylor et al., 2003; Kowalska et al., 2012).

Therefore, the present study was suggested to investigate the prevalence of bacterial infection in rabbit intestines experimentally infected with coccidiosis and the effect of prebiotic supplementation on it. In the prophylactic trial, the prevalence of E. coli was 6.66.11% in the NC group and 13.33% in the PC group while, it was 4.4% in the PS group. These findings were in agreement with those of Jouany, Gobert & Medina (2008), Kimse (2009) and Michelland, Combes & Monteils (2010). Prebiotics were proven to have positive effect on certain pathogens where they able to control enteric diseases associated with E. coli (Kritas & Morrison, 2005; Timmerman et al., 2005). In this respect, Servin & Coconnier (2003) found that the gram-positive bacterium Lactobacillus lactis produced hydrogen peroxide and reduced of the growth of Escherichia coli 0157:H7. In the present study the isolated E. coli showed high resistance against neomycin, florfenicol, flumequin, sulphamazole,trimethoprim, nalidixic acid and tetracycline while, they were sensitive to ciprofloxacine and amikacin. These outcomes similar to those of Flickinger & Fahey (2002), Zhao et al. (2018) and Makhol, Habreh & Sakural (2011).

On the other hand, the prevalence of Salmonella was 2.22% in the NC group and 6.7% in the PC group while it was not detected in the PS group. The presence of Salmonella species with coccidiosis was previously reported by Arakawa et al. (1992). They suggested that the infection with E. tenella leading to changes in the balance of competitive adherence of bacteria which allowing more colonization of S. typhimurium and Clostridium perfringens. Salmonella isolates reported in the present study showed high resistance to sulphamazole, florfenicol, amoxicillin, flumequin and naldixic acid while it revealed high sensitivity to ciprofloxacine, levofloxacin and gentamycin. Several previous studies have demonstrated antimicrobial sensitivity outcomes similar to those reported in the present study (Kumar, Sharma & Mani, 2009; Camarda et al., 2012; Kim et al., 2012; Albuquerque et al., 2014; Agrawal & Hirpurkar, 2016; Lamas et al., 2016).

The present study showed high prevalence of E. coli and Salmonella serovars in the PC group that indicates a relation between the infection by the intestinal coccidiosis and the proliferation of enterobacteriacae micro-organisms. This finding was supported by Szabóová et al. (2012) as they observed significant decrease in the bacterial prevalence in association with reduction in the Eimeria oocysts count in rabbits which were administered a dietary supplementation of natural substances. In addition, El-Ashram et al. (2020) found that the coccidiosis infection in post weaning rabbits mostly associated with E. coli and Salmonella species.

In the present study, the used prebiotic reduced the prevalence of E. coli and Salmonella species associated with experimentally induced coccidiosis in rabbits with reduction of the adverse effect of coccidiosis. This in agreement with the results of El-Ashram et al. (2019) as they found that the prebiotic supplementation reduced the adverse effects of the intestinal coccidiosis in rabbits. Interestingly, Salmonella species was absent in the PS group which may reflect the ability of prebiotic to inhibit colonization of Salmonella. This finding supported by Micciche et al. (2018) as they showed that prebiotic creates of an environment that and inhibit Salmonella colonization and growth in chicken intestine. In addition, Tran et al. (2018) found that a prebiotic supplementation can cause inhibition to Salmonella and E. coli infections in pigs. Moreover, Girgis et al. (2020) realized that prebiotic (Actigen^® a mannan-rich yeast cell wallderived preparation) supplementation diminished the prevalence of Salmonella enteritis in cecal contents of layer chickens. Prebiotics are always used as feed additives to improve growth, endorse beneficial gastrointestinal microbiota, and decrease pathogens. The prebiotics increase short chain fatty acid (SCFA) production in the cecum which leading to pathogen reduction (Micciche et al., 2018; Girgis et al., 2020). The prebiotics promote the overall health and wellbeing of the bird through creation of an intestinal environment unfavorable for Salmonella colonization (Micciche et al., 2018).

In conclusion, the using of prebiotic as prophylaxis in this study, significantly reduced the prevalence of the E. coli and prevented the salmonella infection that associated with intestinal coccidiosis in rabbits. In addition, it reduced the intestinal coccidiosis adverse effect as evidenced by the reduction of oocysts shedding.

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