Combining capture and lysis of the bacteria with partial purification of the plasmid DNA is beneficial for the design of efficient plasmid production processes at larger scale. Such an approach is possible when the bacteria are captured by filtration. Taking industrial requirements into account, however, such a capture requires complex filtration mixtures containing retentive additives such as bentonite and polycations. This makes the straightforward transfer of established lysis protocols to in situ lysis difficult. In this contribution, the different steps of such a protocol are designed for complex filter cakes, including fragilization (by lysozyme), lysis (alkaline pH/acidic pH, 70/37 degrees C, urea/NaCl/Triton), and specific elution (pH, NaCl, CaCl2, guanidinium hydrochloride). Results are compared in regard to plasmid quality (topoisomeric form) and quantity (compared to the yield obtained by a commercial miniprep of a small aliquot of the bacteria suspension from the bioreactor). Best results in these terms were obtained by the Triton lysis protocol performed at 37 degrees C (30 min of contact with a lysis buffer composed of 50 mM Tris pH 8, 1% Triton, 1 g/L lysozyme, and 6 M guanidinium hydrochloride) followed by the specific elution of the plasmid DNA in 50 mM Tris buffer pH 8.
EUROCKOT is now a firmly established and operational launch service provider for small and medium sized satellites into low earth orbit and beyond using the Rockot launch vehicle. Eurockot, which is backed by strong parent companies, namely Europe's EADS SPACE Transportation (51%) and Russia's Khrunichev (49%) owns world class payload preparation facilities at its launch site, Plesetsk in northern Russia. ROCKOT is a three stage liquid fuelled launch vehicle with the Russian SS-19 Intercontinental Ballistic Missile providing its first two stages.
Background
Antimicrobial stewardship (AMS) strategies worldwide focus on optimising the use of antibiotics. Selective susceptibility reporting is recommended as an effective AMS tool although there is a lack of representative studies investigating the impact of selective susceptibility reporting on antibiotic use. The aim of this study was to investigate the impact of selective susceptibility reporting of Staphylococcus aureus (S. aureus) on antibiotic consumption. Enhancing the use of narrow-spectrum beta-lactam antibiotics such as flucloxacillin/cefazolin/cefalexin is one of the main goals in optimising antibiotic therapy of S. aureus infections.
Methods
This interventional study with control group was conducted at a tertiary care hospital in Germany. During the one-year interventional period susceptibility reports for all methicillin-sensitive S. aureus (MSSA) were restricted to flucloxacillin/cefazolin/cefalexin, trimethoprim-sulfamethoxazole, clindamycin, gentamicin and rifampin/fosfomycin, instead of reporting all tested antibiotics. The impact of implementing selective reporting was analysed by monitoring total monthly antibiotic consumption in our hospital and in a reference hospital (recommended daily dose/100 occupied bed days: RDD/100 BD), as well as on an individual patient level by analysing days of therapy adjusted for bed days (DOT/ 100 BD) for patients with S. aureus bacteremia (SAB) and respectively skin and soft tissue infections (SSTI).
Results
MSSA-antibiograms were acquired for 2836 patients. The total use of narrow-spectrum beta-lactams more than doubled after implementing selective reporting (from 1.2 to 2.8 RDD/100 BD, P < 0.001). The use of intravenous flucloxacillin/cefazolin for SAB rose significantly from 52 to 75 DOT/100 BD (plus 42%), just as the use of oral cefalexin for SSTI (from 1.4 to 9.4 DOT/100 BD, from 3 to 17 of 85/88 patients). Considering the overall consumption, there was no decrease in antibiotics omitted from the antibiogram. This was probably due to their wide use for other infections.
Conclusions
As narrow-spectrum beta-lactams are not widely used for other infections, their increase in the overall consumption of the entire hospital was a strong indicator that selective reporting guided clinicians to an optimised antibiotic therapy of S. aureus infections. On a patient level, this assumption was verified by a significant improved treatment of S. aureus infections in the subgroups of SAB and SSTI. As useful AMS tool, we recommend implementing selective reporting rules into the national/international standards for susceptibility reporting.
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