Maldi tof vitek ms

Rectal swab specimens were collected as part of the Antimicrobial Resistance Surveillance Program, in which patients were screened for CRE carriage by rectal swab on admission, according to the hospital protocols. For the isolation of CRE strains from rectal swab specimens, a 10 μL aliquot from each specimen was inoculated into a CHROMID^® CARBA SMART selective chromogenic media bi-plate (bioMérieux, Marcy l’Etoile, France).
The plates were incubated at 37 °C for 18 h and then examined for growth (Escherichia coli showing dark pink or red colonies; Klebsiella spp., Enterobacter spp., and Citrobacter spp. showing metallic blue colonies).
If a swab tested positive, any suspected colony was further identified using the Vitek MS MALDI-ToF (bioMérieux, Marcy-l’Etoile, France).
Species identification was followed by a molecular test (Xpert^® Carba-R test, Cepheid, Sunnyvale, CA, USA) to detect the gene sequences from pure colonies. The following genes were identified: bla_KPC, bla_NDM, bla_VIM, bla_OXA-48, and bla_IMP.

The skin covering the mammary glands was cut and peeled off. From each dam, the mammary gland revealing most intensive signs of inflammation (redness, induration and swelling) was opened by a transverse cut with a sterile scalpel and swabbed for bacterial isolation (Transport Swabs, VWR, Radnor, PA, USA). If no signs of inflammation were present, a random gland was sampled. Swabs from the mammary gland tissue and the intestinal contents of the mink kits were transported to Section of Veterinary Clinical Microbiology, University of Copenhagen, where they were inoculated on blood agar [blood agar base (Oxoid, Thermo Scientific, Waltham, MA, US)] enriched with 5% sterile bovine blood. The agar plates were incubated aerobically at 37 °C for 1–2 days. Bacterial colonies were sub-cultured and identified with Matrix-Assisted Laser Desorption Ionization Time-Of-Flight mass spectrometry (MALDI-TOF–MS) using a VITEK MS MALDI-TOF (BioMérieux, Marcy-l’Etoile, France) as described elsewhere [23 (link)]. From the opened rectum of one random kit from each litter, a swab was taken from the intestinal contents for bacterial isolation.

Post mortem, the intestine was removed from the abdomen as previously described [24 ]. The aboral part of the colon was aseptically opened with a sterile scalpel and swabbed for bacterial cultivation (Transport Swabs, VWR, Radnor, PA, USA). The total of 17 pooled swab control samples were generated by pooling swabs from two healthy kits from the same litter, on the same day, from 17 litters age-matched to the kits affected by PWD. The age-matching of the mink kits unfortunately resulted in some of the kits being pooled into the same tube. This left only 17 pooled samples, instead of 20, from kits without PWD. From the diarrheic kits, the samples were cultured individually. The swabs were stored at 5 °C until analysis by blinded cultivation. The swabs were inoculated on blood agar plates enriched with 5% bovine blood (blood agar base, Oxoid, Thermo Scientific, Watham, MA, USA) and incubated at 37 °C aerobically for 1–2 days. The two most frequently occurring bacterial colonies were then sub-cultivated before they were identified by Matrix-Assisted Laser Desorption Ionization Time-Of-Flight mass spectrometry (MALDI–TOF–MS) using a VITEK MS MALDI-TOF (BioMérieux, Marcy-l’Etoile, France) as previously described [25 (link)]. If two bacterial species were equally frequent next to a single more frequent species both were sub-cultivated, and hence three isolates were identified.