Microbiology Research 2017; volume 8:7328 Investigation of germ caused been reported to be from 2 to 10%,3 and by Guillain-Barré syndrome in even the primary cause of death in devel- Correspondence: Vadood Razavilar, oped countries, especially among children Department of Food Hygiene, Science and poultry meat products in North under 5 in the United States, was attributed Research Branch, Islamic Azad University, West of Iran to such bacteria in that two million cases of Tehran Province, Tehran, Hesarak, Iran. bacterial infection are reported each year.2 E-mail: [email protected] The bacteria which was first discovered Vadood Razavilar, Davoud Nassiri Key words: Campylobacter, Poultry Meat, in the nineteenth century by Theodor Edible Offals, m-PCR. Department of Food Hygiene, Science Escherichia a gram-negative, spiral-shaped and Research Branch, Islamic Azad or curved, growing and multiplying at Contributions: the authors contributed equally. University, Tehran, Iran Microaerophilic conditions at a temperature of 42°C, and below 25°C, it stops growing Conflict of interest: the authors declare no and multiplying but does not disappear.2 potential conflict of interest. Abstract The thermophilic Campylobacter species, such as Jejuni and Coli, have an important Received for publication: 22 July 2017. Campylobacter species are the most role in the development of Revision received: 20 August 2017. important pathogens that cause bacterial Campylobacteriosis becoming more promi- Accepted for publication: 17 September 2017. gastroenteritis being spread through food nent in the food infection as the spread of with animal origin. Given such fact, the cur- This work is licensed under a Creative Campylobacter contamination in food has rent study aimed at investigate the preva- Commons Attribution NonCommercial 4.0 become very important in recent years lence of Campylobacter based on phenotyp- License (CC BY-NC 4.0). going even further than Salmonella.2 ic and genotypic analysis of poultry meat Studies show that the bacterium originates ©Copyright V. Razavilar and D. Nassiri, 2017 and edible offal in Western Azerbaijan. To mainly from food with animal resource. Licensee PAGEPress, Italy conduct the study, a total of 552 chicken Moreover, raw milk, meat and poultry, Microbiology Research 2017; 8:7328 samples including meat (samples), liver untreated surface water and edible fungi are doi:10.4081/mr.2017.7328 (138 samples), gizzard (138 samples) and only the sources of infections with hearts (138 samples) were randomly col- Campylobacteriosis disease being common lected from poultry slaughterhouses at West in spring and summer. Infections with Azerbaijan province from April 2014 to Campylobacter have been reported in sever- September 2014. Based on the culture tests, al countries in broiler chickens, so thatuse the 208 samples (37/7%) were contaminated Materials and Methods bacteria survived during the slaughtering with Campylobacter species. The highest To conduct the study, a total of 552 process and were transmitted to uninfected range of Campylobacter species out breaks poultry organs.2 Due to its high sensitivity samples including chicken meat (138 sam- was observed in poultry liver (49/2%), fol- and detection speed, the PCR method has ples), liver (138 samples), gizzard (138 lowed by gizzard (42/8%), heart (33/3 %) the potentiality to determine if there is even samples) and hearts (138 samples) were and meat (25/4%). Among the isolated a bacterium per ml and reaches the response randomly collected from poultry slaughter- Campylobacter, C. jejuni was the most in a very short time. Many studies have houses at West Azerbaijan province from prevalent (78/4%) and the rest were C. coli examined the prevalence of the bacteria in April 2014 to September 2014and were species (21/6%). All 208 species of meat and poultry as well as diarrheal sam- transferred to the laboratory for analysis Campylobacter isolated as C. jejuni and C. ples.4 The consumption of half-baked meat aiming at investigating the presence of coli from species culturing were also and the related products serves as the main Campylobacter species in cold temperature. approved by m.PCR. The results of the sourcecommercial of human infection although other To determine the phenotype of bacteria, 10 study pinpointed to the chicken edible offal livestock meat and milk products are poten- grams of the sample was added to a bag importance as a potential source of tial sources of the disease. There are reports with one hundred mL broth Preston Campylobacterial contaminations. on the prevalence of Campylobacter among (Preston enrichment broth base, Himedia, Nonlive birds and animals and different kinds of Mumbai, India, m899) containing five per- food from all over the world revealing upon cent defibrinated sheep blood for enrich- a wide range of results. Previous reports ment. The bags were incubated for 24 hours Introduction highlighted the infection in chickens from at 42°C in carbon dioxide presence. Then, Campylobacteriosis, serving as an zero to 100% which was up to 60% for the the liquid in the bag was removed by a ster- important common disease, shoulders a cows. The prevalence of the infection in ile loop and cultured on specific major role in infectious gastroenteritis in poultry given to the market have been Campylobacter selective Agar (Himedia, humans and is considered as the world’s reported to be 100% and lower incidences Mumbai, India, m994) as a separate line to first cause of gastroenteritis followed by of other animals meat were also given.5,6 obtain single separated colonies. Then, it dysentery and diarrhea and other complica- Although there are many studies casting was incubated for 48 hours (Co2 10% v/v) tions such as meningitis, Guillain Barre light on the prevalence of Campylobacter at 42°C. After the initial plate analysis, the syndrome and cholecystitis.1,2 species among poultry in Iran,7-10 and other suspected colonies were stained with Digestive disorders and diarrhea caused countries including Korea,11 Japan,6 Gram’s stain and upon the curved bacteria by the bacterial Campylobacter is among Canada,12 Ireland,13 Pakistan,14 Belgium,15 resolution, the re-culturing was also the common illnesses primarily in develop- little data existed on the poultry meat bi- applied. Upon obtaining the colonies, the ing countries, and 5 to 15 percent of diar- products infection including liver, gizzard biochemical tests including catalase, rhea in these countries are instigated by the and heart in Iran, therefore, the study aimed Indoxyl acetate hydrolysis were used and in bacteria, and the prevalence of the bacteria at evaluating the contamination of these order to differentiate between the two as a cause of diarrhea in our country has products. species Jejuni and Coli hippurate hydrolysis [page 80] [Microbiology Research 2017; 8:7328] Article test was conducted, which was positive for Campylobacter, the Jejuni type was the The outbreak of Campylobacter species Jejuni.1 most prevalent (78/4%) and the rest were of in chicken meat in other countries also sug- To investigate the Campylobacter geno- Coli type (21/6%). All 208 species of gests a contamination level of 30 to 90 per- typic, DNA extraction and Multiplex PCR Campylobacter isolated as Jejuni and Coli cent. The contamination levels in different method were applied. Using the culturing types from culturing were also approved by countries were as follows: Turkey with method and DNA extraction kit (Cina Gen, multiplex polymerase chain reaction test 92/8%,18 Korea with 68/3%,11 Canada with Iran), the confirmed colonies DNA were (m.PCR). A statistically significant differ- 62/4%,12 Japan with 60%,6 Ireland with extracted according to the Kit manufactur- ence (P<0.05) was observed in 49/90%,13 and Pakistan with 48%.14 er’s instructions. The PCR test in this study Campylobacter species outbreak in meat Despite having many studies reporting the 16 followed Denis et al. procedure. For PCR samples taken in summer (40/9 %). The prevalence of Campylobacter species con- reaction, the reaction final volume of 25 results are presented in Tables 2-4 and in tamination in poultry, there are few studies microlitres was considered including 20 ng Figure 1. highlighting on the edible offal contamina- DNA template, 2 mM MgCl2, 25 pmol of tion to the bacteria. each primer, a single Taq polymerase A similar study conducted by Rahimiin enzyme and 200 mM mixed dNTP. The size 2006 and 2008 casted light on the preva- of PCR products corresponding to each Discussion lence of Campylobacter in the chicken liver sample is given below. To confirm the pres- The given result shows 208 samples marketed in Isfahan. Accordingly, the con- ence of the multiplied sample, 20 (37/7%) were infected with Campylobacter tamination of 205 samples under study was microlitres of PCR was paced on elec- species. Previous studies in Iran have reported to be 49/3%, and the liver contam- trophoresis with 1/5 percent agarose gel reported the contamination level to the bac- containing ethidium bromide in the pres- ination for the, chicken, turkey and ostrich teria for different cities as Isfahan with 56/1 were 49/3%, 40% and 16/7%, respectively.8 ence of 100 bp DNA marker in constant 8 7 percent, Shahrekord with 47 percent, Also, Shakerian et al.19 in a study on evalu- voltage of 80 volts (Table 1). 9 Tehran with 63/2% and 49/5%, and ating the Campylobacter contamination in 17 Mashhad with 76%. poultry liver in Shahrekord indicated that Results only The current study aimed at evaluating Campylobacter phenotypic and genotypic outbreaks in chicken meat and its edible use offal at West Azerbaijan province, Iran. To conduct the study, a total of 552 samples including chicken meat (138 samples), liver (138 samples), gizzard (138 samples) and hearts (138 samples) were randomly col- lected from poultry slaughterhouses at West Azerbaijan province from April 2014 to September 2014 and were transferred to the laboratory for analysis aiming at investigat- ing the presence of Campylobacter species in cold temperature.