Xld agar

RIVET reporters were used for the quantification of Salmonella gene expression in tomatoes. Activation of a promoter of interest cloned upstream of the promoterless tnpR recombinase gene was determined by scoring the frequency with which TnpR excised an antibiotic resistance cassette cloned in between the ‘res’ sites that are recognized and acted upon by TnpR (Angelichio and Camilli, 2002 (link)). For the RIVET assays in tomatoes, Salmonella cultures were grown at 37°C overnight in LB supplemented with the appropriate antibiotic(s) (Noel et al., 2010a (link)). Bacterial cultures were then pelleted, washed three times in an equal volume of sterile PBS. Approximately 10² cfu (in 3 μl of water) were inoculated onto superficial 1 mm deep wounds on surfaces of unwaxed fruits. At least two technical (individual infections) and three biological (tomatoes from different plants) replications were carried out for each experiment. Unless otherwise stated, infected tomatoes were incubated at 22°C in vented chambers. All RIVET assays were carried out for a week. To harvest samples, 15 × 0.5 mm cores were removed from fruits, homogenized in PBS, and aliquots were then plated onto XLD agar (Oxoid) with appropriate antibiotics. Individual colonies were then patched on LB agar with tetracycline (10 μl ml⁻¹) to detect constructs in which TnpR recombinase was active.

This study has conveniently targeted 52 broiler chicken flocks (Avian 48, Abdelsalam Hegazy Company), of which 26 were one-week-old chick flocks and 26 were six-week-old birds. These farms were investigated for Salmonella infections. The broiler chicken flocks were surveyed in Kafrelsheikh, Gharbia, and Menofeya provinces in Egypt during 2020–2022. The birds showed different clinical signs, including reluctance to move, pasty diarrhea, huddling near the source of the heat, ruffling feathers, dehydration, decreased body weight gain, droopy wings, lameness, and high mortalities of 9.64% ± 1.72 in one-week-old chicks from each broiler chicken farm. Four living, diseased birds were selected randomly and sacrificed. At postmortem, sections from the liver, gallbladder, spleen, and intestinal contents were collected under aseptic conditions for Salmonella isolation. Within five hours of collection, samples were delivered to the lab and stored on ice until then. Selenite-F broth (SFB) (Oxoid, UK) was combined with one gramme of tissue from each organ and incubated statically at 37°C for an overnight period. The enrichments were applied to XLD agar (Oxoid, UK) using a swap, and they were then incubated at 37°C overnight. One colony from each plate that appeared to be Salmonella spp. was chosen for additional examinations based on appearance [29 (link)].

In accordance with the testing method of the Korean Food Code laid out by the MFDS, a 25 g sample and 225 ml of Buffered Peptone Water (Oxoid, UK) were mixed thoroughly and enriched in the incubator at 37°C for around 24 h. The enriched culture solution was added to two enrichment media, 1 ml to 10 ml of teterathionate medium (Biomeriux Inc., Spain) and 0.1 ml to 10 ml of Rappaport-Vassiliadis medium (Oxid, UK), which underwent secondary enrichment at 37°C (tetrathionate) and 42°C (RV medium) for 20-24 h. The secondary enrichment culture solution was smeared on the selective media of XLD agar (Oxoid) and Brilliant green sulfa agar (Remel, UK), and cultured at 37°C for 18-24 h, and then a typical colony was selected and subcultured in the nutrient medium, which was identified by Vitek MS (Biomeriux Inc., France).

After 24 hours incubation sub-culturing was performed from the Typtic Soya broth on XLD agar (OXOID, England). After overnight incubation positive cultures were proceed further while Negative broth cultures were incubated for seven days and sub cultured before reported negative. Suspected colonies obtained on the above media were screened by biochemical tests using Triple Sugar Iron agar (TSI) (BBL™), citrate utilization test, motility (Difco™), urease test (Himedia ltd. India) and lysine decarboxylation (LDC) [Difco™] test.

Different clinical specimens were inoculated onto appropriate culture media (sheep blood agar, XLD agar, and MacConkey agar plates (Oxoid Ltd, UK)) and incubated overnight under the aerobic condition at 37°C for 18-24 hours. Identification of Enterobacteriaceae was done using colony characteristics, Gram staining reaction, and ability to ferment lactose. In addition, the Phoenix system (BD Diagnostic Systems, Oxford, UK) was used for the identification of the bacteria to species level. The combination panel includes identification (ID) side with dried substrates for bacterial identification, and the instrument tests panels every 20 minutes. After three hours, the identified bacteria were displayed on the screen of the Phoenix system [13 ].