Reducing E. coli Challenges in Turkeys Despite Enrofloxacin Resistance


Photo: iStockphoto_Anna Usova

The gastrointestinal tract of birds is intensively populated by many different microorganisms. Recent analytical technologies, such as next-generation sequencing, have made it possible to characterize this microbiome thoroughly. In general, the microbiota is a very important component for the host as it can influence the development and function of the digestive and immune systems. While there are lot of studies available on the chicken microbiome, not many have been published regarding the microbiome of turkeys. The importance of species-specific studies was highlighted by Pan and Yu (2014) who showed that chickens and turkeys have only 16% similarity in their intestinal microbiome. Wilkinson et al. (2017) showed that Firmicutes, Bacteroidetes, Actinobacteria, and Proteobacteria were the dominant phyla in the microbiota of turkeys across age and location. Escherichia coli belong to the Proteobacteria phylum and are common inhabitants of the gastrointestinal tract of turkeys, but they can also be found in other birds and also in mammals. E. coli predominantly inhabit their host without causing any harm.

However, there are certain E. coli strains that possess specific virulent genes which are able to cause diseases in birds (avian pathogenic E. coli – APEC). Avian colibacillosis is one of the most common diseases in poultry. Colibacillosis can occur in many different clinical forms, causing considerable economic losses to producers worldwide. To counteract colibacillosis, antimicrobials need to be used. However, the misuse or incorrect application of antibiotics can contribute to the spread of antimicrobial resistance, which poses a threat both for animals and humans. In relation to the latest trends and pressure coming from the market, the need for alternatives to be used as preventive tools to avoid colibacillosis has become crucial.

Organic acids: an alternative solution

Organic acids or single chain fatty acids (SCFA) have been proven to be toxic for many microorganisms. This toxicity is primarily associated with the ability of the undissociated acids to freely diffuse across the membranes of bacteria. Once inside the cell, the acids will dissociate into anions and protons and the resulting anions can affect cell growth in many different ways. In order to support and facilitate the passage of the acids across the bacterial membrane, the use of SCFA can be combined with the use of permeabilizers that destabilize the outer membrane of Gram-negative bacteria (like E. coli and Salmonella spp.) and hence ease the passage of the acids into the cells. Such a formulation has been considered for the development of the enhanced acidifier Biotronic® Top liquid: a combination of substances able to permeate the outer membrane of Gram-negative bacteria (Permeabilizing Complex™) with a well-studied blend of organic acids. Such a product can be added to the water supply as a preventive tool to reduce the replication of pathogenic E. coli in birds.

Biotronic® Top liquid: prevention against E. coli replication

In a trial performed in cooperation with the Istituto Zooprofilattico Sperimentale della Lombardia e dell’Emilia- Romagna “Bruno Ubertini”, 80 one-day-old female turkeys (Big 6 Aviagen®) where fed three different diets (Table 1). Feed and water administered to the birds was tested for the absence of E. coli, Enterobacteriaceae, Clostridium spp. and Salmonella spp. before it was offered. The diets were administered from day 4 onwards after the birds were preventively treated with colistin at the dose recommended by the manufacturer. On day 11 of the trial, all the birds were challenged with 1.38x108 CFU of E. coli O78 serotype, an APEC which was isolated during an incidence of colisepticemia in a turkey flock in Italy, 2014. The strain was found to be resistant to enrofloxacin. On day 4 of the trial, one animal per group was sacrificed by cervical dislocation in order to confirm the absence of the E. coli O78 as well as any other E. coli strain. All other animals were left in the pens to grow. On day 20 and 30 of the trial, ten birds from each group were sacrificed by cervical dislocation and examined by bacteriological analysis and lesion score assessment.

Table 1. Experimental diets

Negative control (NC)Standard diet
Positive control (PC)Standard diet + enrofloxacin supplemented in the water at 0.50 mL/L (from day 11 to day 20 of the trial)
Biotronic® Top liquid (BTR)Standard diet + Biotronic® Top liquid supplemented in the water at 1.25 mL/L (during the whole period)

Source: BIOMIN

Table 2. Lesion scores and their descriptions

Lesion ScoreDescription
0No lesions
0.5One yellow or brown pin-head sized inflammatory spot
1Two or more pin-head sized inflammatory spots
2Thin layer of fibrinous exudate in various locations
3Thick and extensive fibrinous exudation

Adapted from Van Eck and Goren, 1991.

Lesion scores

Liver lesions of the sacrificed animals were scored using a slightly modified version of that described by Van Eck and Goren (1991) (Table 2). The mean lesion score was calculated for each group. There were no signs of lesions on day 20 in the Biotronic® Top liquid (BTR) group. However, results were not found to be significantly different from the negative control (NC) or positive control (PC). On day 30, the lesion score for the BTR group was significantly different (p<0.05) from both the PC and NC (Figure 1).

Figure 1. Average lesion score of the liver 

Average lesion score of the liver content

E. coli count

E. coli were isolated from both the intestinal tract and the liver, and were further enumerated using appropriate buffers and agar. Biotronic® Top liquid supplemented to the water reduced E. coli counts in the intestinal tract and liver of turkeys. On day 20 and day 30, E. coli counts in the intestinal tract of turkeys in the BTR group were significantly reduced (p<0.05) compared to NC and PC as shown in Figure 2. On day 20 and day 30, no E. coli counts were found in the liver samples of the BTR group, whereas the NC and the PC groups were both found to be positive for E. coli. On day 30, the E. coli count in the liver was significantly lower (p<0.05) in the BTR group compared to NC and PC (Figure 3).

Figure 2. Average E. coli count of the intestinal content

Average E. coli count of the intestinal content

Figure 3. Average E. coli count of the liver

Average E. coli count of the liver

Biotronic® Top liquid: a profitable solution

The treatment of colibacillosis in poultry should take into consideration the costs of treating flocks with the correct dosage for a sufficiently long period, and the rising percentage of isolated E. coli that are resistant to antibacterial drugs. The diagnosis of colibacillosis is mainly based on the clinical features and the typical macroscopic lesions. But in order to confirm infection, E. coli need to be isolated and identified. Further, bacterial resistances need to be excluded. All these steps take time, leading to losses in production when the analysis needs to be outsourced. They can also lead to wrong decisions being made when it comes to selecting the right therapy to be adopted. In this case, treatment with a commonly used antibiotic (enrofloxacin) would not have been effective, as highlighted by the results, because the bacteria used for the challenge was resistant to this specific drug. This is why it is important to work on a prevention strategy against colibacillosis rather than relying on therapy alone.

Conclusion

A holistic approach that includes proper management, proper vaccination measures and the right nutritional design are needed in order to prevent the spread of E. coli and hence colibacillosis in turkeys but also in poultry. The use of feed additives like the enhanced acidifier Biotronic® Top liquid can support the replication reduction of pathogenic bacteria in the animal, and the product can play a crucial role in reducing environmental E. coli contamination.

In Brief
  • The gastrointestinal tract of turkeys contains a complex microbiome of bacteria, most causing no harm to the host
  • Some bacterial strains cause diseases leading to economic losses
  • The downward trend in antibiotic use has driven the rise of alternative disease management strategies
  • Biotronic® Top liquid can reduce the E. coli count in turkeys when used as part of a holistic approach to turkey production

Reference

Pan, D. and Yu, Z. (2014). Intestinal microbiome of poultry and its interaction with host and diet. Gut Microbes 5(1): 108 – 119.

Van Eck, J.H. and Goren, E. (1991). An Ulster 2C strain-derived Newcastle disease vaccine: vaccinal reaction in comparison with other Ientogenic Newcastle disease vaccines. Avian Pathology 20(3) pp. 497-507.

Wilkinson T.J., Cowan, A.A., Vallin, H.E., Onime, L.A., Oyama, L.B., Cameron, S.J., Gonot, C., Moorby, J.M., Waddams, K., Theobald, V.J., Leemans, D., Bowra, S., Nixey, C. and Huws, S.A. (2017). Characterization of the Microbiome along the Gastrointestinal Tract of Growing Turkeys. Front Microbiol 8: 1089.

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