Phenotypic and Genotypic Characterization of Biofilm Production in Staphylococcus aureus Isolates Obtained from Food Contact Surfaces in Butcher and Delicatessen

Year 2023, Volume: 20 Issue: 3, 198 - 205, 04.12.2023

Nihat Akyol , Dursun Alp Gündoğ , Yasin Özkaya , Candan Güngör , Nurhan Ertaş Onmaz

https://doi.org/10.32707/ercivet.1389025

Cited By: 1

Abstract

In the present study, it was aimed to investigate the presence of Staphylococcus aureus (S. aureus) in
butcher and delicatessen environments in Kayseri province and analyze their biofilm abilities with phenotypic and gen-
otypic methods. For this study, a total of 200 samples were collected from 10 different businesses, which were random-
ly selected between August and December 2022.The presence of S. aureus was investigated with EN/ISO 6888-1
(01/2004) standard, and the determination of the nuc gene in PCR, and its biofilm ability was investigated with Congo
red agar (CRA) and microplate method (MP). In addition, the presence of biofilm-related genes in isolates with biofilm
ability was also investigated with PCR method. Out of 200 samples, 42 (21%) were identified as S. aureus. Of these 42
isolates, 5 belonged to knives, 7 to cutting board, 8 to minced meat, 6 to cubed meat, 2 to sausage, 4 to meatballs, 6 to
cheese, and 4 to pastrami. According to CRA and MP tests, all S. aureus isolates were detected as biofilm producers,
and 18 (43%) of the isolates were strong, 13 (31%) were moderate, and 11 (26%) were weak in the MP test. Out of
biofilm producer isolates, icaA and icaD genes were detected in 18 (43%) and 16 (38%), respectively, while none of the
isolates contained fnbA and fnbB genes. As conclusion, the biofilm production ability of S. aureus isolates from the
butcher and delicatessen environment in Kayseri are considered as public health risk. Therefore, improving hygienic
conditions in food enterprises is essential for food safety and consumer health.

Keywords

Animal origin food, biofilm, biofilm related genes, public health, S. aureus

Kasap Dükkanları ve Şarküterilerde Gıda ile Temas Eden Yüzeylerden Elde Edilen Staphylococcus aureus İzolatlarında Biyofilm Üretiminin Fenotipik ve Genotipik Karakterizasyonu

Abstract

Bu çalışmada, Kayseri ilindeki kasap ve şarküteri ortamında Staphylococcus aureus’un (S. aureus) varlığı, elde edilen izolatların biyofilm üretim yeteneklerinin fenotipik ve genotipik yöntemlerle araştırılması amaçlanmıştır. Bu amaçla, Ağustos-Aralık 2022 tarihleri arasında rastgele seçilen 10 farklı işletmeden toplam 200 örnek elde edilmiştir. S. aureus varlığı EN/ISO 6888-1 (01/2004)’da belirtilen metoda ek olarak PCR’da nuc geninin belirlenmesiyle tespit edilmiş, biyofilm yeteneği ise Kongo kırmızısı agar (KKA) ve mikroplaka (MP) yöntemi ile araştırılmıştır. Ayrıca biyofilm yeteneğine sahip izolatlarda biyofilm ile ilişkili genlerin varlığı yine PCR yöntemi ile araştırılmıştır. Analiz edilen 200 örneğin 42’si (%21) S. aureus olarak identifiye edilmiştir. S. aureus izolatlarının, beşi bıçak, yedisi kesme tahtası, sekizi kıyma, altısı kuşbaşı, ikisi sucuk, dördü köfte, altısı peynir ve dördü pastırma örneklerine aitti. Çalışmada elde edilen, izolatların tamamının hem KKA’da hem de MP testinde biyofilm pozitif olduğu belirlenirken, MP testine göre izolatların 18 (%43)’inin güçlü, 13 (%31)’ünün orta derecede ve 11 (%26)’inin zayıf biyofilm ürettiği tespit edilmiştir. Biyofilm pozitif olarak belirlenen izolatların 18’i (%43) ve 16’sı (%38) analiz edilen genlerden sırasıyla icaA ve icaD geni taşırken hiçbir izolatın fnbA ve fnbB genlerini barındırmadığı belirlenmiştir. Sonuç olarak, Kayseri ilindeki kasap ve şarküteri ortamından izole edilen S. aureus’ların biyofilm üretme yeteneği halk sağlığı açısından tehlike arz etmektedir. Bu nedenle, işletmelerde hijyen koşullarının iyileştirilmesi gıda güvenliği ve tüketici sağlığı için önemlidir.

Keywords

Biyofilm, biyofilm ile ilgili genler, halk sağlığı, hayvansal gıdalar, S. aureus

References

• Achek R, Hotzel H, Nabi I, Kechida S, Mami D, Di-douh N, Tomaso H, Neubauer H, Ehricht R, Mo-necke S, El-Adawy H. Phenotypic and molecular detection of biofilm formation in Staphylococcus aureus isolated from different sources in Algeria. Pathogens 2020; 9(2):153.
• Arciola CR, Campoccia D, Gamberini S, Cervellati M, Donati E, Montanaro L. Detection of slime production by means of an optimised Congo red agar plate test based on a colourimetric scale in Staphylococcus epidermidis clinical isolates genotyped for ica locus. Biomater 2002; 23(21): 4233-9.
• Aydin A, Sudagidan M, Muratoglu K. Prevalence of staphylococcal enterotoxins, toxin genes and ge-netic-relatedness of foodborne Staphylococcus aureus strains isolated in the Marmara Region of Turkey. Int J Food Microbiol 2011; 148(2): 99-106.
• Ballah FM, Islam MS, Rana ML, Ullah MA, Ferdous FB, Neloy FH, Ievy S, Sobur MA, Rahman AT, Khatun MM, Rahman M, Rahman T. Virulence determinants and methicillin resistance in biofilm-forming Staphylococcus aureus from various food sources in Bangladesh. Antibiotics 2022; 11(11): 1666.
• Borena BM, Gurmessa FT, Gebremedhin EZ, Sarba EJ, Marami LM. Staphylococcus aureus in cow milk and milk products in Ambo and Bako towns, Oromia, Ethiopia: Prevalence, associated risk factors, hygienic quality, and antibiogram. Int Microbiol 2023; 26: 513-27.
• Chen Q, Xie S, Lou X, Cheng S, Liu X, Zheng W, Zheng Z, Wang H. Biofilm formation and prevalence of adhesion genes among Staphylococcus aureus isolates from different food sources. Mi-crobiologyopen 2020; 9(1): e00946.
• Cramton SE, Gerke C, Schnell NF, Nichols WW, Götz F. The intercellular adhesion (ica) locus is present in Staphylococcus aureus and is required for biofilm formation. Infect Immun 1999; 67(10): 5427-33.
• Cremonesi P, Perez G, Pisoni G, Moroni P, Morandi S, Luzzana M, Brasca M, Castiglioni B. Detection of enterotoxigenic Staphylococcus aureus isolates in raw milk cheese. Lett Appl Microbiol 2007; 45(6): 586-91.
• El-Azzouny MM, El-Demerdash AS, Seadawy HG, Abou-Khadra SH. Antimicrobial effect of garlic (Allium sativum) and thyme (Zataria multiflora Boiss) extracts on some food borne pathogens and their effect on virulence gene expression. Cell Mol Biol 2018; 64(10): 79-86.
• Ferreira FA, Souza RR, de Sousa Moraes B, de Amorim Ferreira AM, Américo MA, Fracalanzza SE, Dos Santos Silva Couceiro JN, Sá Figueiredo AM. Impact of agr dysfunction on virulence pro-files and infections associated with a novel methi-cillin-resistant Staphylococcus aureus (MRSA) variant of the lineage ST1-SCCmec IV. BMC Mi-crobiol 2013; 27: 93.
• Freeman DJ, Falkiner FR, Keane CT. New method for detecting slime production by coagulase negative staphylococci. J Clin Pathol 1989; 42: 872.
• Gungor C, Barel M, Dishan A, Disli HB, Koskeroglu K, Onmaz NE. From cattle to pastirma: Contamination source of methicillin susceptible and resistant Staphylococcus aureus (MRSA) along the pastirma production chain. LWT 2021; 151: 112-30.
• Guven K, Mutlu MB, Gulbandilar A, Cakir P. Occurrence and characterization of Staphylococcus aureus isolated from meat and dairy products consumed in Turkey. J Food Saf 2010; 30(1):196-212.
• Güzel N, Onmaz NE. Toplu yemek üretimi yapan bir işletmede personel ve gıda temas yüzeylerinin mikrobiyolojik yönden değerlendirilmesi. Erc Üniv Vet Fak Derg 2022; 19(3): 189-94.
• Houston P, Rowe SE, Pozzi C, Waters EM, O'Gara JP. Essential role for the major autolysin in the fibronectin-binding protein-mediated Staphylococcus aureus biofilm phenotype. Infect Immun 2011; 79(3): 1153-65.
• ISO 6888-1.Microbiology of Food and Animal Feeding Stuffs–Horizontal Method for the Enumeration of Coagulase‐Positive Staphylococci (Staphylococcus aureus and Other Species)-Part 1: Technique Using Baird‐Parker Agar Medium, 1999.
• Kadkhoda H, Ghalavand Z, Nikmanesh B, Kodori M, Houri H, Taghizadeh Maleki D, Karimi Ba-vandpour A, Eslami G. Characterization of biofilm formation and virulence factors of Staphylococcus aureus isolates from paediatric patients in Tehran, Iran. Iran J Basic Med Sci 2020; (5): 691-8.
• Koçak Kızanlık P. Çeşitli kaynaklardan izole edilen Staphylococcus aureus’un bazı virulens özellikle-rinin belirlenmesi. Doktora tezi, Adnan Menderes Üniv, Sağlık Bil Enst, Aydın, 2019.
• Kowalska J, Maćkiw E, Stasiak M, Kucharek K, Postupolski J. Biofilm-forming ability of pathogenic bacteria isolated from retail food in Poland. J Food Protect 2020; 83(12); 2032-40.
• Maira-Litrán T, Kropec A, Abeygunawardana C, Joyce J, Mark G, Goldmann DA, Pier GB. Immunochemical properties of the staphylococcal poly-N-acetylglucosamine surface polysaccharide. Infect Immun 2002; 70(8): 4433-40.
• Miao J, Lin S, Soteyome T, Peters BM, Li Y, Chen H, Su J, Li L, Li B, Xu Z, Shirtliff ME, Harro JM. Bio-film formation of Staphylococcus aureus under food heat processing conditions: First report on CML production within biofilm. Sci Rep 2019; 9(1): 1312.
• Normanno G, Corrente M, La Salandra G, Dambrosio A, Quaglia NC, Parisi A, Greco G, Bellacicco AL, Virgilio S, Celano GV. Methicillin-resistant Staphy-lococcus aureus (MRSA) in foods of animal origin product in Italy. Int J Food Microbiol 2007; 117(2): 219-22.
• Özlü S, Ercoşkun H. Et ve sağlıklı beslenme. Gıda ve Yem Bil Tekn Derg 2021; 25: 15-29.
• Paparella A, Serio A, Rossi C, Mazzarrino G, López CC. Food-Borne transmission of staphylococci. Savini V. eds. In: Pet-To-Man Travelling Staphylo-cocci. Academic Press, 2018; pp.71-99.
• Pérez-Boto D, D’Arrigo M, García-Lafuente A, Bravo D, Pérez-Baltar A, Gaya P, Medina M, Arqués JL. Staphylococcus aureus in the processing environment of cured meat products. Foods 2023; 12(11): 2161.
• Seyoum B, Kefyalew H, Abera B, Abdela N. Prevalence, risk factors and antimicrobial susceptibility test of Staphylococcus aureus in bovine cross breed mastitic milk in and around Asella town, Oromia regional state, southern Ethiopia. Acta Tropica 2018; 177: 32-6.
• Stepanović S, Vuković D, Dakić I, Savić B, Švabić-Vlahović M. A modified microtiter-plate test for quantification of staphylococcal biofilm formation. J Microbiol Methods 2000; 40: 175-9.
• Tang Y, Larsen J, Kjeldgaard J, Andersen PS, Skov R, Ingmer H. Methicillin-resistant and-susceptible Staphylococcus aureus from retail meat in Den-mark. Int J Food Microbiol 2017; 249: 72-6.
• Tebliğleri, TGK. Mikrobiyolojik Kriterler Tebliği. TC Tarım ve Orman Bakanlığı, Resmi Gazete, Tarih, (28157), 2011: 2001-19.
• Thiran E, Di Ciccio PA, Graber HU, Zanardi E, Ianieri A, Hummerjohann J. Biofilm formation of Staphylococcus aureus dairy isolates representing different genotypes. J Dairy Sci 2018; 101(2): 1000-12.
• Tiryaki C. Toplu tüketim işletmelerinde tüketime hazır gıdalar ve ilgili personelde S. aureus prevalansı ile bazı virulens özelliklerin incelenmesi. Yüksek lisans tezi, İstanbul Üniv Sağ Bil Ens, İstanbul, 2018.
• Turhan EÜ, Erginkaya Z. Bakteriyel biyofilmlerdeki antimikrobiyel direnç mekanizması. Akademik Gıda 2019; 17(1): 131-9.
• Uyanık T, Bölükbaş A, Gücükoğlu A, Çadırcı Ö. Çeşitli gıda örnekleri ve kesimhanelerden izole edilen bazı patojen bakterilerin biyofilm oluşturma yeteneğinin araştırılması. J Vet Bio Sci Tech 2022; 7(3): 338-45.
• Vergara-Irigaray M, Valle J, Merino N, Latasa C, Gar-cía B, Ruiz de los Mozos, Solano C, Toledo-Arana A, Penades JR, Lasa I. Relevant role of fibronectin-binding proteins in Staphylococcus aureus biofilm-associated foreign-body infections. Infect Immun 2009; 77(9): 3978-91.
• Zeaki N, Johler S, Skandamis PN, Schelin J. The role of regulatory mechanisms and environmental parameters in staphylococcal food poisoning and resulting challenges to risk assessment. Front Microbiol 2019; 12(10): 1307.