Ispitivanje mogućnosti formiranja biofilma u uslovima in vitro kod različitih serovarijeteta Salmonella vrsta izolovanih iz uzoraka hrane za životinje

Abstract

Cilj ovog istraživanja je bio ispitivanje sposobnosti formiranja biofilma kod serovarijeteta Salmonella izolovanih iz hrane za životinje, poreklom iz 18 mešaona stočne hrane na području Južnobačkog okruga. Izolati korišćeni u ispitivanju (n=100) izolovani su poreklom iz uzoraka hrane za životinje uzorkovanih tokom dve godine ispitivanja (period 2012-2014). Identifikacija izolata izvršena je na osnovu kulturelnih, biohemijskih i seroloških karakteristika na osnovu preporuka standarda SRPS EN ISO 6579:2008. Potvrda i serološka tipizacija izolata Salmonella izvedena je u Nacionalnoj referentnoj laboratoriji za Salmonella, Shigela, Vibrio cholerae i Yersinia enterocolitica, Instituta za javno zdravlje Srbije „Dr Milan Jovanović Batut“ u Beogradu. Kao kontrolni sojevi, u ispitivanju su korišćene referentne kulture S. Typhimurium ATCC 14028 i S. Enteritidis ATCC 13076. Kao supstrat za formiranje biofilma korišćene su površine od polistirena i nerđajućeg čelika. Sposobnost formiranja biofilma ispitana je primenom sledećih metoda: Congo red agar test, pelikula test, test na mikrotitracionim pločama upotrebom kristal violet boje, i skenirajuća elektronska mikroskopija (SEM). Sva ispitivanja izvedena su upotrebom dve hranljive podloge (tripton soja bujona i Luria Bertani bujona) pri temperaturi inkubacije od 20C i 37C. Iz hrane za životinje izolovani su sledeći serovarijeteti Salmonella: Tennessee (18%), Agona (9%), Montevideo (15%), Enteritidis (12%), Stanleyville (6%), Infantis (12%), Typhimurium (3%), Typhimurium monofazni (1%), Mbandaka (6%), Senftenberg (7%), Jerusalem (2%), Thompson (1%), Amsterdam (1%), Colindale (1%), Dahra (1%), Livingstone (1%). Kod četiri izolata nije ustanovljen serovarijetet (Salmonella enterica subspecies enterica (1,3,19:i:-)). Prema rezultatima Congo red agar testa, na temperaturi od 20C, 98% izolata Salmonella ekspresioniralo je jednu ili obe komponente matriksa biofilma i formiralo kolonije RDAR, PDAR i BDAR morfotipova...

The aim of this research was the investigation into the biofilm-forming ability of Salmonella serovars isolated from animal feed samples originating from 18 feedproducing facilities located in South Bačka District. The isolates which were tested (n=100) were isolated from feed samples collected in a two-year period (from 2012 to 2014). The identification of the isolates was made based on the cultural, biochemical and serological characteristics, following the recommendations given in the SRPS EN ISO 6579:2008 standard. The conformation and serological typing of Salmonella isolates were done in the National Reference Laboratory for Salmonella, Shigela, Vibrio cholerae and Yersinia enterocolitica, at the Institute of Public Health of Serbia 'Dr Milan Jovanović Batut' in Belgrade. In this study reference cultures of S. Typhimurium ATCC 14028 and S. Enteritidis ATCC 13076 were used as the control strains. Surfaces made of polystyrene and of stainless steel were used as substrates for biofilm formation. The ability of Salmonella to form biofilm was tested with the following methods: Congo red agar test, pellicle test, microtiter plate assay with the use of crystal violet dye, and scanning electron microscopy (SEM). The whole investigation was carried out at two temperatures of incubation (20C and 37C). The following Salmonella serovars were isolated from animal feed: Tennessee (18%), Agona (9%), Montevideo (15%), Enteritidis (12%), Stanleyville (6%), Infantis (12%), Typhimurium (3%), Typhimurium monofazni (1%), Mbandaka (6%), Senftenberg (7%), Jerusalem (2%), Thompson (1%), Amsterdam (1%), Colindale (1%), Dahra (1%), Livingstone (1%). In four isolates serovars were not determined (Salmonella enterica subspecies enterica (1,3,19:i:-)). According to the results of Congo red agar test, when grown at 20C, 98% of the isolates expressed one or both components of the Salmonella biofilm matrix and formed colonies of various morphotypes (RDAR, PDAR and BDAR)...