Comparative evaluation of the detection rate, workflow and associated costs of a multiplex PCR panel versus conventional methods in diagnosis of infectious gastroenteritis.

Introduction. Infectious gastroenteritis is a common reason for consulting a physician. Although most cases of gastrointestinal illness are self-limiting, the identification of the etiologic pathogen by stool specimen analysis is important in cases of more severe illness and for epidemiological reasons.Due to the broad range of causative pathogens, the conventional examination of a stool specimen is labour-intensive and usually requires different diagnostic methods. Multiplex PCR tests [e.g. BioFire Gastrointestinal (GI) Panel] allow the rapid detecting of up to 22 pathogens in one test.Hypothesis. Using a multiplex PCR panel to test stool specimens for infectious gastroenteritis pathogens can improve the detection rate, reduce the time-to-result and hands-on time and lower the costs of a microbiology laboratory.Aim. This study was aimed at evaluating the detection rate, the workflow and associated costs of stool specimen management using the BioFire GI Panel versus conventional methods.Methodology. Stool specimens were evaluated prospectively during the routine operation. Pathogen detection rate, hands-on time, time-to-result and material and personnel costs were determined for the BioFire GI Panel and conventional methods-the latter based on physician request and excluding viral testing.Results. Analysing 333 specimens collected between 2019 and 2020, the detection rate of enteropathogens was significantly higher with a positivity rate of 39.9 % using the multiplex PCR panel compared with 15.0 % using the conventional methods. The BioFire GI Panel presented results in a median time of 2.2 h compared with 77.5 h for culture and 22.1 h for antigen testing, noting that no tests were performed at weekends except for toxinogenic Clostridioides difficile. Based on list prices, the BioFire GI Panel was nine times more expensive compared with conventional methods, whereas hands-on-time was significantly lower using the BioFire GI Panel.Conclusion. Multiplex PCR panels are valuable tools for laboratory identification of infectious agents causing diarrhoea. The higher costs of such a multiplex PCR panel might be outweighed by the higher detection rate, ease of handling, rapid results and most likely improved patient management. However, these panels do not provide information on antimicrobial susceptibility testing. Therefore, if this is necessary for targeted therapy or if outbreak monitoring and control is required, specimens must still be cultured.

Antimicrobial Activity of Ceftolozane-Tazobactam, Ceftazidime-Avibactam, and Cefiderocol against Multidrug-Resistant Pseudomonas aeruginosa Recovered at a German University Hospital.

Multidrug-resistant (MDR) Pseudomonas aeruginosa increasingly causes health care-associated infections. In this study, we determined the activity of ceftolozane-tazobactam, ceftazidime-avibactam, and cefiderocol against 223 MDR P. aeruginosa clinical isolates recovered from 2013 to 2017 at the University Hospital Frankfurt by using MIC test strips. Furthermore, we evaluated the presence of genes encoding major ß-lactamases, such as VIM, IMP, NDM, GIM, SPM, and KPC; the extended spectrum ß-lactamase (ESBL)-carbapenemase GES; and the virulence-associated traits ExoS and ExoU, as in particular ExoU is thought to be associated with poor clinical outcome. For MDR P. aeruginosa isolates, the MIC50/MIC90 values of ceftolozane-tazobactam, ceftazidime-avibactam, and cefiderocol were 8/>256 mg/L, 16/>256 mg/L, and 0.25/1 mg/L, respectively. Cefiderocol showed the highest susceptibility rate (97.3%) followed by ceftazidime-avibactam (48.4%) and ceftolozane-tazobactam (46.6%). In 81 (36.3%) isolates, carbapenemase gene blaVIM was detected, and in 5 (2.2%) isolates, blaGES was detected (with a positive association of exoU and blaVIM). More than half of the isolates belong to the so-called international P. aeruginosa "high-risk" clones, with sequence type 235 (ST235) (24.7%) being the most prevalent. This study underlines that ceftolozane-tazobactam, ceftazidime-avibactam, and cefiderocol are important options for the treatment of infections due to MDR P. aeruginosa, with cefiderocol currently being the most active available antipseudomonal ß-lactam agent. According to our clinical experience, the outcome of cefiderocol therapy (8 patients) was favorable especially in cases of MDR P. aeruginosa-associated complicated urinary tract infections. IMPORTANCE After testing ceftolozane-tazobactam, ceftazidime-avibactam, and cefiderocol against a collection of 233 multidrug-resistant (MDR) Pseudomonas aeruginosa, we showed that cefiderocol is the most active antipseudomonal ß-lactam agent (susceptibility rates were 46.6%, 48.4%, and 97.4%, respectively). The most prevalent one was sequence type 235 (ST235) (24.7%), followed by ST244, ST175, and ST233, with all belonging to the top 10 P. aeruginosa high-risk clones with worldwide distribution. Our data indicate that during surveillance studies special attention should be paid to the MDR and highly virulent VIM- and ExoU-producing variant of ST235. Furthermore, in the case of infections caused by carbapenemase-producing MDR P. aeruginosa, cefiderocol is the preferred treatment option, while outcomes of complicated urinary tract infections and hospital-acquired pneumonia with cefiderocol were favorable.

[Systemic and local antibiotic therapy of conservative and operative treatment of spondylodiscitis]. / Systemische und lokale Antibiotikatherapie bei konservativ und operativ behandelten Spondylodiszitiden.

An evidence-based recommendation for a standardized antibiotic therapy of spondylodiscitis has not yet been published. Crucial for conservative therapy is the verification of the causative organism and an appropriate antibiotic therapy. Intravenous antibiotic therapy should be administered for 2-3 weeks and a switched to oral administration for 6-12 weeks is then possible. If an empirical antimicrobial therapy is required a combination of ciprofloxacin and clindamycin, alternatively a combination of cefotaxim and flucloxacillin is recommended. Surgical removal of the infection by extensive debridement with stabilization and filling the resulting bone defect is desirable. Under the perception of a high local dose of antibiotic the defect filling with a mixture of cancellous bone and antibiotic-loaded hydroxyapatite and calcium sulfate is advisable.

Antibacterial resistance of community-acquired respiratory tract pathogens recovered from patients in Germany and activity of the Ketolide Telithromycin: results from the PROTEKT surveillance study (1999-2000).

BACKGROUND: The Prospective Resistant Organism Tracking and Epidemiology for the Ketolide Telithromycin (PROTEKT) longitudinal global surveillance study examines the antibacterial susceptibility of community-acquired respiratory pathogens. METHODS AND RESULTS: Data from isolates collected in Germany in 1999-2000 in the PROTEKT study show that 8.3% of pneumococcal isolates (n = 325) had reduced susceptibility to penicillin and 2.2% were fully resistant. Erythromycin resistance was 15.7% overall and particularly high in Leipzig (31.6%). All penicillin- and erythromycin-resistant strains were inhibited by telithromycin (MIC < or =0.5 mg/l) and linezolid (MIC < or =2 mg/l). Beta-lactamase was produced by 3.2% of Haemophilus influenzae (9/284) and 89.5% of Moraxella catarrhalis strains (111/124). All Streptococcus pyogenes isolates (n = 87) were susceptible to penicillin, although 9.2% were resistant to macrolides. CONCLUSIONS: Penicillin resistance in Germany remains low; however, the prevalence of antimicrobial resistance among common respiratory pathogens is rising, particularly against macrolides. Continued surveillance is necessary to guide optimal empirical therapy, and new antimicrobials, like telithromycin, need to be developed with improved potency against target pathogens and low propensity for the development of resistance.

[Evidence-based antibiotic prophylaxis in aseptic orthopedic surgery]. / Perioperative Antibiotikaprophylaxe bei aseptischen Eingriffen in der Orthopädie.

Disagreement exists on the topic of antibiotic prophylaxis in aseptic orthopedic surgery. No evidence on the usefulness of prophylactic antibiotic administration exists with regard to non-complex aseptic surgeries without placement of osteosynthetic material. Likewise, no undisputed evidence exists on the usefulness of antibiotic prophylaxis with regard to aseptic orthopedic surgeries involving placement of osteosynthetic material. However, the majority of experts agree on antibiotic prophylaxis in the latter cases. In contrast clear evidence does exist regarding the usefulness of antibiotic prophylaxis with first- or second-generation cephalosporins for surgeries of the hip involving fracture treatment or prosthetic replacement. The prophylactic use of glycopeptides should be confined to cases of high MRSA or MRSE risk. Administration of prophylactic antibiotics should precede incision time by around 30 min and tourniquet inflation by at least 10 min. Antibiotic administration may be repeated in the OR when surgery lasts longer than 3 h. The use of local antibiotics in bone cement has not proven useful as a prophylactic measure.

Antimicrobial effectiveness of povidone-iodine and consequences for new application areas.

The microbicidal action spectrum of povidone-iodine (PVP-I) is broad - even after short onset times. Unlike local antibiotics and other antiseptic substances, no resistance develops. The high degree of bactericidal efficiency in respect of highly resistant gram-positive pathogenic micro-organisms, such as methicillin-resistant Staphylococcus aureus (MRSA) and enterococcus strains, is particularly significant for hospital hygiene. An in vitro study with 10 genotypically different MRSA isolates showed an optimum bactericidal effect (logarithmic reduction factor value >5) without protein load after just 30 s exposure and even in a dilution of Betaisodona solution (Mundipharma GmbH) of 1%. With protein load (0.2% albumin), the optimum in microbicidal effectiveness shifts to concentrations > or = 10% Betaisodona solution referring to an exposure time of 30 s. Since recent results are now also available on the toxicological safety of PVP-I preparations for the ciliated epithelium of the nasal mucosa and the good tolerability on skin and other mucous membranes is a known factor, a controlled clinical study is currently being carried out to eliminate colonizations of MRSA. Evidence has also recently been produced of the antiviral activity of PVP-I against herpes simplex, adeno- and enteroviruses, as well as its high degree of efficiency against Chlamydia. Hence alongside the classical fields of application, such as the disinfection of the skin and hands, mucosa antisepsis and wound treatment, there are also useful indications for the substance, i.e. rinsing of body cavities and joints and application to the eye.

Removal of PCR inhibitors by silica membranes: evaluating the Amplicor Mycobacterium tuberculosis kit.

The effectiveness of PCR inhibitor removal by silica membranes in combination with the Amplicor Mycobacterium tuberculosis kit was analyzed for 655 respiratory and nonrespiratory specimens. The overall inhibition rate was reduced from 12.5%, when applying the Amplicor kit alone, to 1.1% with the addition of silica membrane DNA purification.

In vitro activities of fluoroquinolones against the spirochete Borrelia burgdorferi.

Little is known to date about the in vitro activity of fluoroquinolones against Borrelia species. Our study aimed at determining the in vitro activities of 15 quinolones against nine isolates of the Borrelia burgdorferi sensu lato complex in addition to one Borrelia valaisiana and one Borrelia bissettii tick isolate. For the determination of MICs, a standardized colorimetric microdilution method was applied. Determination of minimal borreliacidal concentrations providing 100% killing of the final inoculum (MBCs) after 72 h and time-kill experiments were performed by conventional culture in Barbour-Stoenner-Kelly medium in combination with dark-field microscopy. The rank order of potency on a microgram-per-milliliter basis for the substances with in vitro activity against B. burgdorferi was gemifloxacin (MIC at which 90% of the isolates tested are inhibited [MIC(90)], 0.12 microg/ml) > sitafloxacin (MIC(90), 0.5 microg/ml), grepafloxacin (MIC(90), 0.5 microg/ml) > gatifloxacin (MIC(90), 1 microg/ml), sparfloxacin (MIC(90), 1 microg/ml), trovafloxacin (MIC(90), 1 microg/ml) > moxifloxacin (MIC(90), 2 microg/ml), ciprofloxacin (MIC(90), 2 microg/ml) > levofloxacin (MIC(90), 4 microg/ml) > ofloxacin (MIC(90), 8 microg/ml), norfloxacin (MIC(90), 8 microg/ml) > fleroxacin (MIC(90), >16 microg/ml), and pefloxacin (MIC(90), 32 microg/ml) > nalidixic acid (MIC(90), 256 microg/ml). After 72 h of exposure, gemifloxacin was borreliacidal (100% killing) against the isolates investigated at a median MBC of 4 microg/ml. In the other compounds tested, median MBCs were higher (> or =8 microg/ml). Results of electron microscopy and time-kill studies clearly support an in vitro activity of some fluoroquinolones against borreliae. Our study demonstrates for the first time the enhanced in vitro effectiveness of some of the recently introduced 4-quinolones against B. burgdorferi.

Rapid molecular typing of methicillin-resistant Staphylococcus aureus by PCR-RFLP.

OBJECTIVE: To establish a new, rapid, and reliable genotypic fingerprinting technique for methicillin-resistant Staphylococcus aureus (MRSA) typing in routine epidemiological surveillance. DESIGN: The method is based on polymerase chain reaction (PCR) restriction fragment-length polymorphism (RFLP) following HaeII digestion of simultaneously amplified parts of the protein A gene, the coagulase gene, and the hypervariable region adjacent to mecA. A total of 46 MRSA initial isolates were analyzed, including 14 isolates from five countries; the six German epidemic strains; 16 isolates from the Frankfurt metropolitan area, which were known to be heterogeneous by pulsed-field gel electrophoresis (PFGE); and 10 isolates obtained during three epidemics, all of which displayed an identical genotype. RESULTS: Restriction analysis by PCR-RFLP permitted discrimination of 10 of 14 international isolates, all six German epidemic strains, and 15 of 16 national isolates. It also confirmed the homogeneous character of the 10 outbreak isolates. CONCLUSIONS: This new and rapid PCR-RFLP typing method is an attractive tool in routine epidemiological surveillance. Its impressive characteristics are ease of performance and interpretation, while at the same time guaranteeing good discriminatory power, reproducibility, and typeability.

Elution characteristics of vancomycin, teicoplanin, gentamicin and clindamycin from calcium sulphate beads.

The in vitro release of vancomycin, teicoplanin, gentamicin and clindamycin from biodegradable calcium sulphate (CaSO(4)) carrier beads is described. All antibiotics showed prolonged release from the carrier beads, which was elevated during the first 24 h, with peak levels exceeding 2500 microg/bead. Doubling the antibiotic load of the beads revealed a more prolonged elution and a two-fold increase in antibiotic release. Local carrier-associated antibiotic treatment with CaSO(4) beads may prove to be effective in the management of chronic bone infections.

In vitro activity of mezlocillin, meropenem, aztreonam, vancomycin, teicoplanin, ribostamycin and fusidic acid against Borrelia burgdorferi.

The in vitro susceptibility profile of Borrelia burgdorferi is not yet well defined for several antibiotics. Our study explored the in vitro susceptibility of B. burgdorferi to mezlocillin, meropenem, aztreonam, vancomycin, teicoplanin, ribostamycin and fusidic acid. Minimal inhibitory concentrations (MICs) and minimal borreliacidal concentrations (MBCs) were measured using a standardised colorimetric microdilution method and conventional subculture experiments. MIC values were lowest for mezlocillin (MIC(90), < or =0.06 mg/l) and meropenem (MIC(90), 0.33 mg/l). Vancomycin (MIC(90), 0.83 mg/l) was less effective in vitro. Borreliae proved to be resistant to aztreonam (MIC(90), >32 mg/l), teicoplanin (MIC(90), 6.6 mg/l), ribostamycin (MIC(90), 32 mg/l), and fusidic acid (MIC(90), >4 mg/l). The mean MBCs resulting in 100% killing of the final inoculum after 72 h of incubation were lowest for mezlocillin (MBC, 0.83 mg/l). This study gathered further data on the in vitro susceptibility patterns of the B. burgdorferi complex. The excellent in vitro effectiveness of acylamino-penicillin derivatives and their suitability for the therapy of Lyme disease is emphasised.

[Significance of methicillin resistant S. aureus (MRSA) in geriatrics--epidemiology, therapy and management]. / Bedeutung des Methicillin-resistenten S. aureus (MRSA) in der Geriatrie--Epidemiologie, Therapie und Management.

Methicillin-resistant S. aureus (MRSA) has become an important cause of severe infection in hospitalized patients all over the world. In Germany a significant increase of nosocomial infections due to MRSA has occurred during the last 10 years. Especially elderly patients with chronic illnesses are at increased risk of becoming colonized or infected with MRSA. This report focuses on epidemiology and therapy of MRSA, and on the recommendations concerning management and prevention of spread of MRSA in hospitals and nursing homes.

Colorimetric in vitro susceptibility testing of penicillins, cephalosporins, macrolides, streptogramins, tetracyclines, and aminoglycosides against Borrelia burgdorferi isolates.

The spectrum of antibiotic susceptibility of Borrelia burgdorferi has been only partially defined. In the present study the effectiveness of 12 antimicrobials, belonging to six different antibiotic classes have been tested against Borrelia burgdorferi s.s. (N=3), Borrelia garinii (N=3), Borrelia afzelii (N=3), Borrelia valaisiana (N=1), and Borrelia bissettii (N=1) isolates. These isolates were analysed by a new standardised colorimetric minimal inhibitory concentration (MIC) method based upon colour changes that result from actively metabolizing spirochaetes after 72 h of incubation. Piperacillin (MIC90: 0.08 mg/l), ceftriaxone (MIC90: 0. 04 mg/l), cefotaxime (MIC90: 0.15 mg/l), azithromycin (MIC90: 0.015 mg/l), roxithromycin (MIC90: 0.05 mg/l) and quinupristin/dalfopristin (MIC90: 0.12 mg/l) gave the lowest MIC values. Minimal inhibibitory activity of amoxycillin (MIC90: 1.04 mg/l), cefixime (MIC90: 1.33 mg/l), cefoperazone (MIC90: 0.83 mg/l) tetracycline (MIC90: 0.29 mg/l) and minocycline (MIC90: 0.30 mg/l) was slightly lower, whereas borrelia were resistant to amikacin (MIC90: >128 mg/l). Mean minimal borreliacidal concentrations (MBCs) were representatively determined for piperacillin (MBC: 1.8 mg/l), ceftriaxone (MBC: 2.0 mg/l), azithromycin (MBC: 0.82 mg/ml), roxithromycin (MBC: 1.8 mg/l), quinupristin/dalfopristin (MBC: 5.0 mg/l), minocycline (MBC: 5.8 mg/l), and amikacin (MBC: >128 mg/l) by using conventional subculture for three weeks in combination with dark-field microscopy. B. garinii proved to be the most susceptible of the genospecies tested. Our study showed excellent in vitro antimicrobial activity of all classes of antibiotics tested, except the aminoglycosides and hence their suitability for therapy of Lyme disease.

Risk of Pseudomonas aeruginosa cross-colonisation in patients with cystic fibrosis within a holiday camp--a molecular-epidemiological study.

OBJECTIVE: A study on the molecular epidemiology of Pseudomonas aeruginosa in patients with cystic fibrosis (CF) from Germany (N = 18) and Israel (N = 12) is presented. The aim is to provide an answer to the question as to whether or not social contact outside the hospital environment involves a potential risk for person-to-person spread of this pathogen. METHODS: Sputa from German and Israeli patients were obtained while these were attending a holiday camp in Israel. The sputum samples were analysed with regard to Pseudomonas aeruginosa. Strains dissimilar in macroscopic appearance and/or antibiotic resistance patterns were genotyped using pulsed-field gel electrophoresis after digestion of genomic DNA with restriction endonuclease Spel. The genetic polymorphism of DNA fragment patterns of all strains (N = 146) was studied for their overall relatedness using a fingerprint software system. RESULTS: Most of the German patients (77.7%) were colonised persistently by a unique clonal type during the four-week screening period. Isolates obtained from Israeli patients displayed a very close clonal relationship and a higher antibiotic resistance as a result of preceding epidemic spread of certain clones before the camp. Additionally, isolates showing identical PFGE patterns were demonstrated once in a single male Israeli patient and in one female German patient, suggesting previous cross-colonisation. CONCLUSION: The occurrence of person-to-person spread through social contact in patients with CF is supported by our findings, but remains a rare event outside the hospital environment, provided appropriate hygienic measures are applied.

Colonization with superantigen-producing Staphylococcus aureus is associated with increased severity of atopic dermatitis.

BACKGROUND: Atopic dermatitis (AD) is a chronic inflammatory skin disease associated with colonization of the skin with Staphylococcus aureus known to produce toxins with superantigen (SAg) activity. Besides T-cell activation these toxins induce T-cell skin homing in vitro. This may contribute to the observed induction or enhancement of skin inflammation. OBJECTIVE: The aim of this study was to determine whether colonization with SAg-producing S. aureus isolates modulates the intensity of AD. If so, it was of interest whether this may be primarily due to the toxins' effects as SAgs or as allergens. METHODS: In AD patients, healthy controls, and atopic controls SAg production by S. aureus isolated from skin or mucous membranes was investigated and correlated to the severity of the disease. Total IgE, SAg-specific IgE, and T-cell activation and recirculation markers were analysed and correlated with SAg production. RESULTS: Fifty-seven percent of S. aureus strains isolated from AD patients produced SAgs. This frequency was higher compared to healthy controls (33%). SAg production by S. aureus was correlated with a significantly higher scoring of AD (SCORAD index, 58 +/- 19 in SAg-producing vs 41 +/- 7 in non-SAg-producing germs; P < 0.05). However, the severity of the disease was not associated with sensitization against the SAgs staphylococcal enterotoxin A (SEA) and staphylococcal enterotoxin B (SEB). Furthermore, SAg production by S. aureus was inversely correlated with total IgE concentration (P < 0.05) and positively correlated with T-cell activation (as measured by HLA-DR and CD69 expression) and the expression of the T-cell skin homing phenotype cutaneous lymphocyte-associated antigen. CONCLUSION: SAg production by S. aureus is suggested to be associated with an increased severity of atopic dermatitis. Since SAg production was found neither exclusively in AD patients nor in all patients, other pathogenic factors may be additionally effective.