Efficiency of Sanitary Treatment in Poultry Breeding and Poultry Meat Processing Plant

Ka‰ková A. , O. Ondra‰oviãová, J . Sokol , M. Vargová, S. ·mir jáková, M. Ondra‰oviã , M. Chovanec: Efficiency of Sanitary Treatment in Poultry Breeding and Poultry Meat Processing Plant. Acta Vet. Brno 75, 2006: 611-617. The aim of this study was to observe the effectiveness of disinfection on a broiler farm and in a plant processing the poultry from this farm. The broiler farm was disinfected with a preparation based on peracetic acid while a preparation based on quarternary ammonium salt was used in the processing plant. We evaluated swabs taken from surfaces, which come into contact with broilers and broiler meat. Results of the swabs taken by standard microbiological swabbing method were evaluated with results of the swabs taken by the ATP-bioluminescence method. The microbiological examination included total counts of microorganisms, coliform count and moulds. When using the standard plate counts method on the broiler farm we found that the plate counts in 0% of swabs were < 1, in 12% of swabs ranged between 1 100 CFU while in 88% of swabs reached > 100 CFU. In the processing plant, out of 22% of swabs < 1 CFU were recovered, in 36% of swabs plate counts ranged between 1 100 CFU and in 42% of swabs plate counts reached > 100. The bioluminescence method was applied only in the processing plant where < 100, 100 300 and > 300 RLU were measured in 80, 10 and 10% of swabs, resp. Our observations and results allowed us to conclude that the disinfectants tested appeared suitable for the respective premises and the ATP bioluminescence method could be use as a as a suitable complement for detection of cleanliness of individual surfaces. Sanitization, disinfection, poultry, processing plant, standard plate counts The present requirements on the production of high-quality and safe food are a subject of concern of professionals and wide consumer public in advanced countries with terminal impact on economics of agricultural production, particularly the primary production. According to the valid standards in the Slovak Republic, and in the interest of protection of consumer’s health and WHO and FAO recommendations, food producers are bound to make steps which eventually prevent the risk associated with consumption of their products (Hofmann 2000; Burdová et al. 2001). As a result, all production stages must comply with the rules of correct operation practice and the HACCP system must be introduced gradually as an extension of correct operation practice. Sanitation processes are an inevitable part of food production as they influence hygiene and outcome of the entire production system (Bremner and Johnston 1996). Poultry industry is one of the most hygiene-conscious sectors of livestock breeding. The cleaning and disinfection in large plants is ensured by specialized sanitation teams. Monitoring of hygiene under operating conditions is ensured by traditional microbiological swabs and lately also by the ATP-detecting bioluminescence method capable of providing information about contamination of surfaces in less than 10 minutes (Green et al. 1999). ACTA VET. BRNO 2006, 75: 611–617; doi:10.2754/avb200675040611 Address for correspondence: Andrea Ka‰ková Department of the Environment University of Veterinary Medicine Komenského 73, 041 81 Ko‰ice, Slovak Republic Phone: +421 905 982 068 E-mail: ajka77@yahoo.com kaskova@uvm.sk http://www.vfu.cz/acta-vet/actavet.htm

The aim of this study was to observe the effectiveness of disinfection on a broiler farm and in a plant processing the poultry from this farm.The broiler farm was disinfected with a preparation based on peracetic acid while a preparation based on quarternary ammonium salt was used in the processing plant.We evaluated swabs taken from surfaces, which come into contact with broilers and broiler meat.Results of the swabs taken by standard microbiological swabbing method were evaluated with results of the swabs taken by the ATP-bioluminescence method.The microbiological examination included total counts of microorganisms, coliform count and moulds.When using the standard plate counts method on the broiler farm we found that the plate counts in 0% of swabs were < 1, in 12% of swabs ranged between 1 -100 CFU while in 88% of swabs reached > 100 CFU.In the processing plant, out of 22% of swabs < 1 CFU were recovered, in 36% of swabs plate counts ranged between 1 -100 CFU and in 42% of swabs plate counts reached > 100.The bioluminescence method was applied only in the processing plant where < 100, 100 -300 and > 300 RLU were measured in 80, 10 and 10% of swabs, resp.Our observations and results allowed us to conclude that the disinfectants tested appeared suitable for the respective premises and the ATP bioluminescence method could be use as a as a suitable complement for detection of cleanliness of individual surfaces.

Sanitization, disinfection, poultry, processing plant, standard plate counts
The present requirements on the production of high-quality and safe food are a subject of concern of professionals and wide consumer public in advanced countries with terminal impact on economics of agricultural production, particularly the primary production.According to the valid standards in the Slovak Republic, and in the interest of protection of consumer's health and WHO and FAO recommendations, food producers are bound to make steps which eventually prevent the risk associated with consumption of their products (Hofmann 2000;Burdová et al. 2001).As a result, all production stages must comply with the rules of correct operation practice and the HACCP system must be introduced gradually as an extension of correct operation practice.Sanitation processes are an inevitable part of food production as they influence hygiene and outcome of the entire production system (Bremner and Johnston 1996).
Poultry industry is one of the most hygiene-conscious sectors of livestock breeding.The cleaning and disinfection in large plants is ensured by specialized sanitation teams.
Monitoring of hygiene under operating conditions is ensured by traditional microbiological swabs and lately also by the ATP-detecting bioluminescence method capable of providing information about contamination of surfaces in less than 10 minutes (Green et al. 1999).
The present study focused on the evaluation of disinfection with peracetic acid-based disinfectant and quarternary ammonium compound on a poultry farm with broilers housed on deep litter and in a poultry processing plant slaughtering and processing broilers from the investigated farm.Potential transfer of contamination from the farm to the processing plant was also investigated.Bacterial plate counts were determined on the farm and in the plant after cleaning (before disinfection), after disinfection, and during the operation.It is clear that cleaning and disinfection are processes essential for good functioning of the food processing plant and production of hygienic and safe food.

Materials and Methods
The study was divided into two parts, laboratory and field ones.The laboratory investigations focused on testing of bactericidal effectiveness of disinfectants on a broiler farm and in the processing plant.The activity of disinfectants was tested by a suspension test after 5, 20 and 60 minutes.Investigations on the broiler farm which housed 10 000 broilers and in the broiler processing plant consisted of taking bacteriological swabs during production and determining plate counts of investigated micro-organisms by the standard plate method.ATPbioluminescence method was used in the processing plant to check the cleanliness of surfaces.
Testing the disinfection efficiency by the qualitative suspension test under laboratory conditions Tests were carried out with peracetic acid and Topax-91 disinfectant based on a quarternary ammonium compound.Bactericidal activity of disinfectants was evaluated under laboratory conditions by the method of qualitative suspension assay (AOAC 1984).Known quantities of bacterial suspension were added to a disinfectant solution of known concentration and incubated for a specified time (5, 20 and 60 minutes).Using a bacteriological loop, mixture aliquots were transferred to tubes with broth and incubated according to the requirements of respective bacteria.The tests were performed using standard collection bacteria with E. coli and S. aureus representing Gram-negative and Gram-positive bacteria, resp., B. cereus as a representative of spore-forming bacteria and A. niger as a representative of moulds.

Contact swabbing places
Selection of surfaces for swab evaluation depended on the type of operation and on the locations, which might appear critical for production.On the poultry farm, the floor, walls, ceiling, drinkers and ventilation fans were considered critical.In the processing plant, we focused on the shackling hooks, eviscerator, cooling tank, shoots, saws, boards, injector, conveyer belts and other equipment that was cleaned manually.To be able to compare the methods, swabs were taken from respective surfaces: from 100 cm 2 in the processing plant and from 10-cm 2 areas on the farm with recalculation per 100 cm 2 .Five swabs were taken from respective locations.In the processing plant, swabs were taken during the production in the presence of workers and 2 -5 hours after disinfection.On the broiler farm, swabs were taken also after emptying the houses before mechanical cleaning and after mechanical cleaning before disinfection.

Microbiological swabbing methods
Sterile cotton swabs were moistened with 10.0 ml sterile saline in a tube.Swabbed places were marked and described.After swabbing, the cotton swab was returned back to the tube while breaking off and removing the handled part of the swab stick.The tubes with swabs were refrigerated until processing.Plate counts of total microorganisms, coliforms and moulds were determined.The samples were processed within 24 hours according to the respective standard methods (•VPS SR No.7089 2002).All counts were expressed as colony forming units (CFU) for the swabbed surface (CFU per milliliter of swabbing solution × 10).

ATP bioluminescence method
The ATP-bioluminescence is based on ATP detection and quantification using an enzyme luciferase and a luciferin cofactor.Hydrolysis of ATP by luciferase produces yellow-green luminescence measured by a luminometer and converted to RLU (relative light units).As one molecule of ATP produces one photon of light, light intensity of the reaction is relative to ATP quantity in the sample (Green et al. 1999).HY-LiTE Merck system was used.

Results and Discussion
Microbial contamination of the poultry meat and its control are important for two reasons.The first reason is that poultry is an important reservoir of pathogens, such as Salmonella and Campylobacter, and a frequent source of food-borne diseases.The second reason is the inclination of raw meat to microbial spoilage if it is stored unfrozen.Some microbial contamination is inevitable due to the poultry meat character and the way it was obtained, however, it is essential to reduce it by effective control of hygiene during the respective operations (Bremner and Johnston 1996).One part of the problem is that poultry is transported to a slaughterhouse from various farms and is strongly contaminated by a wide spectrum of microorganisms.Most of them are eliminated during processing, but many withstand the process and may spread and cross-contaminate other carcasses.It is therefore necessary to ensure optimum hygiene conditions during processing by using detergents and disinfectants and easy-care surfaces.
After cleaning, disinfection plays a key role in the subsequent reduction of viable microorganisms.However, disinfectants used in food industry may contaminate the products and because of that the selection of disinfectants that are effective, suitable for that particular environment, non-toxic and non-tainting is of utmost importance (Holah 1995).
Sanitation is one of the most important measures in the meat industry and involves technologies not less detailed than slaughter or butcher fitting (Gracey and Collins 1992).Incipient foulness in meat and fleshy products processing are mostly composed of proteins and fats.Their composition makes an ideal matrix for multiplication of different species of microorganisms including pathogens (Hofmann 2000).Disinfection in food processing plants should remove or devitalize all disease-producing germs and reduce the counts of other germs to a level, which has no negative influence on the products.
Sanitizing agents, active against free microorganisms, may lose their activity towards germs enclosed in a biofilm.This is prevented by thorough cleaning, application of combined agents (oxidizing substances and surface-active compounds) and frequent monitoring of the surfaces.Pokludová and •kaloud (2002) mentioned that Listeria spp., enclosed in a biofilm, might present a problem as they are preserved well on stainless steel surfaces and survive freezing and drying.
Many authors mentioned that successfulness of antimicrobials depends on their ability to inactivate and eliminate organisms in the biofilm.Mittelmann (1998) reported that low concentrations of natrium hypochlorite at a concentration interval from 0.05 to 5 mg•l -1 act only as inhibitors on the biofilm retained on stainless steel and only concentrations above 50 mg•l -1 are able to inactivate the biofilm micro-organisms under controlled conditions.According to Jessen and Lammert (2003), disinfectants based on hydrogen peroxide and peracetic acid are more effective than chlorine disinfectants.Our tests showed that peracetic acid is a highly effective disinfectant already at low concentrations.The suspension test performed under laboratory conditions (Fig. 1) revealed good effectiveness of this disinfectant against E. coli, S. aureus and B. cereus already at a concentration of 0.1 ml•l -1 at 20 minutes exposure.It was also effective against moulds represented by A. niger, which often raise problems on poultry breeding farms, even with lethal results, at the same concentration as above but with 60 minutes exposure time.Moreover, the residual concentrations are eliminated easily by simple aeration and the compound is also environmentally friendly, as it leaves no residues.When tested on the broiler farm, peracetic acid appeared very effective as the cleaning process decreased plate counts of micro-organisms by 84% on average (Table 1) but disinfection ensured 99.9% effectiveness (Table 2) compared to the original state after emptying the halls.The Topax-91 disinfectant, based on quarternary ammonium compounds, showed very good efficacy.Fig. 2 shows that in the laboratory tests Topax-91 was effective against E. coli, S. aureus and B. cereus germs already at a concentration of 1 ml•l -1 after 60 minutes of exposure and to A. niger at aconcentration of 15 ml•l -1 after 60 minutes of exposure.According to producer's instruction Topax-91 should be used in 5 -10 ml•l -1 concentration at 50 °C and exposure time 1 hour.It is therefore suitable for regular disinfection in poultry industry as it was confirmed by the results obtained after disinfection in the processing plant (Table 3).The only place that allowed us to recover a higher number of microorganisms after disinfection were the shackling hooks.This problem was noted in the majority of cases because, due to their shape, the cleaning and disinfection of these hooks is very difficult.To reach better results it is necessary to perform very thorough mechanical cleaning of hooks before disinfection.In the poultry processing plant, swabs were taken and evaluated also by the ATP-bioluminescence method.The results of this method correlated with those obtained by the standard plating method (Table 3).Comparison of the results of TCM and RLU is presented in Table 4.
A number of studies were published on the reliability of the two methods.Poulis et al. (1993) performed tests in food processing plants and found poor relationship between ATPbioluminescence and the contact method.However, either method in its own right is useful to check the cleanliness in food processing plants.ATP measurement has a great advantage, because it is quick and easy.Tebbutt (1991) compared the contact agar method and swabbing method in the inspection of surface cleanliness in food processing before and after cleaning.He obtained better results with plates, which were as sensitive as the swabbing method.
Table 4. Percentage comparison of disinfectant efficiency in the poultry meat processing plant Table 3. Mean total counts of microorganisms, coliforms and moulds after spray disinfection with a disinfectant based on quaternary ammonium compounds (processing plant) Concentration of QAC was 5 ml•l -1 at 50°C.Results are means of 5 swabs taken from 100 cm 2 surface at all sites.Altogether 100 swabs were taken.The results obtained by the standard microbiological method are compared with the results of ATP-bioluminescence method.
Results are expressed in %.TCM determined by standard plate count method.RLU determined by the ATPbioluminescence method.Swabs were taken from 50 sites.Application of sanitizers after cleaning can often interfere with the surface of bacteria, so the plate method gives negative results or low counts.However, it is a fact that a fine film or even a food layer may remain on the surface and is not detected by the traditional method.This is the biggest danger associated with plate techniques.Residues from processing, such as blood and tissues, remain on the equipment and can participate in increasing the RLU values because the enzyme luciferase reacts with both eukaryotic and prokaryotic ATP.These residues indicate inadequate sanitation and can be detected by the ATP method, whereas plate methods are useful only for quantification of counts of microorganisms left on the equipment (Green et al. 1999).If the surfaces of processing equipment show higher values of ATP regardless of their source, these surfaces should be cleaned repeatedly before the processing operations can start.Moore and Griffith (2002) studied the dependence between the surface quality, cleaning and disinfection and the testing methods.The number of surfaces considered suitable for food production increased after cleaning but the agreement between the testing methods varied with regard to the type of processed food.Insufficient detection of proteins was most frequently observed on surfaces in bakeries while ATP-bioluminescence and the standard microbiological methods failed mostly in freezing facilities and cheese processing plants.The authors concluded that it is necessary to combine these methods when monitoring cleanliness in these facilities.Oulahal-Lagsir et al. (2000) accept the use of ATP-bioluminescence method for quantification of biofilm removal.In comparison with the swabbing method, their results differed by 42% on stainless steel and by 74% on polypropylene surfaces.Frank and Chiemlewski (1997) disinfected various surfaces with quarternary ammonium compounds.They applied QAC solution of concentration 200 mg•l -1 .Their results showed that stainless steel and smooth polycarbonates could be disinfected easier (5 CFU/cm 2 of residual germs) than surfaces of mineral resins (100 CFU/cm 2 of residual germs).Sanitation with quarternary ammonium compounds performed in their study reduced population of S. aureus more than 1.000-fold on all monitored surfaces except the mineral ones.In our case, disinfection with QAC at a concentration of 5 ml•l -1 ensured 99.2% effectiveness.

During the production
In conclusion, sanitary treatment is one of the most important components of the control system HACCP.Cleaning and disinfection as a part of everyday operation practice are essential for high hygienic level in food production in the framework of legislation requirements The farm poultry is highly contaminated with many kinds of bacteria.This has a great influence on contamination of entry parts of the processing plant.The count of microorganisms on surfaces in the processing plant varies throughout the day and depends on surface and the part of the processing stage.Our observations and results allowed us to conclude that the disinfectants tested appeared suitable for the respective premises and the ATP bioluminescence method could be use as a as a suitable complement for detection of cleanliness of individual surfaces.
If the poultry leaves the farm with contamination of about 2.10 6 TCM per 10 cm 2 , it is very important to ensure its disinfection corresponding to national and EU regulations in order to prevent an increase in the counts of microorganisms in food by faulty sanitation programme.

Fig. 1 .
Fig.1.Effectiveness of peracetic acid against representatives of Gram-negative, Gram-positive and spore-forming bacteria and moulds in laboratory suspension tests.Relationship between disinfectant concentration and the exposure time is shown.

Table 1 .
Comparison of cleanliness of poultry farm surfaces after emptying the halls at the end of fattening (before mechanical cleaning) and after mechanical cleaning with water under pressure (before disinfection)Results are means of 5 swabs taken at individual sites from 100 cm 2 surface.Altogether 50 swabs were taken.
Fig.2.Effectiveness of the disinfectant based on quarternary ammonium compound against representatives of Gramnegative, Gram-positive and spore-forming bacteria and moulds in laboratory suspension tests.Relationship between disinfectant concentration and the exposure time is shown.