Evaluation of two different semi-automated homogenization techniques in microbiological diagnosis of periprosthetic joint infection: disperser vs. bead milling method.

BACKGROUND: In microbiological diagnosis of periprosthetic joint infection (PJI) there is no consensus regarding the most suitable and optimal number of specimens to be cultured or the most effective technique of tissue processing. This comparative study analysed the accuracy of two semi-automated homogenization methods with special focus on the volume and exact origin of each sample. METHODS: We investigated a total of 722 periprosthetic tissue samples. PJI was defined according to the new scoring system for preoperative and intraoperative criteria. We compared the performance of our routinely used single tissue processing by disposable high-frequency disperser with the bead milling method. RESULTS: Eighty patients were included. Among forty classified PJIs, 34 patients yielded positive culture results. In 23 cases (68%) exact concordant results were generated with both techniques. However, in seven cases (20%) processing by the disperser and in four cases (12%) by bead milling provided additional positive samples, but without significant difference since the major definition criteria were met in all cases. The percentage of positive results was influenced by the volume and origin of the tissue samples. Results for small tissue samples tended to be better using the bead milling method. This might lead to improved preoperative arthroscopic diagnosis, as the volume of biopsies is generally limited. Six patients had negative results due to previous antimicrobial therapy. Forty other patients were classified as aseptic failures. Neither procedure resulted in any contamination. CONCLUSION: Both methods enable reliable processing of tissue samples for diagnosis of PJI and are suitable for routine use.

Microbiological diagnosis of polymicrobial periprosthetic joint infection revealed superiority of investigated tissue samples compared to sonicate fluid generated from the implant surface.

OBJECTIVES: In the microbiological diagnosis of periprosthetic joint infection (PJI), there is much discussion about the methodology of obtaining proper specimens, the processing technique, and suitable culture media. This retrospective study was conducted to analyse the accuracy of our culture techniques. METHODS: Tissue samples and components from 258 patients after revision arthroplasty of the hip, knee, and shoulder were investigated, and the results of tissue cultures (TC) were compared to those of sonicate fluid cultures (SFC). Furthermore, an evaluation was performed of the influence of different culture media on the detection rate. RESULTS: PJI was confirmed in 186 patients. The overall sensitivity of TC was no different to that of SFC (91.3% vs 90.8%, P = 1). In 153 cases (82.3%), TC and SFC showed concordant positive results. Results were discordant in 33 cases (17.7%). When differentiated according to the type of infection, TC showed significantly better results than SFC in detecting polymicrobial infections (97.0% vs 67.0%, P = 0.004). There were also significant differences between the culture media regarding the yield of microorganisms. CONCLUSIONS: TC was more effective in detecting co-infections. The best results were obtained using both TC and SFC. The choice of culture media has a significant influence on the quality of results.

Sonicate fluid inoculated into blood culture bottles does not improve diagnosis of periprosthetic joint infection caused by anaerobes. A retrospective analysis.

BACKGROUND: In microbiological diagnosis of periprosthetic joint infection (PJI) there is much controversial discussion about culture media and incubation time, especially if anaerobic bacteria are the causative agents. This retrospective analysis was conducted to compare the results obtained by inoculation of sonicate fluid from prosthetic components into BD Bactec blood culture bottles with those obtained by our culture method using sensitive supplemented growth media. METHODS: Twenty-eight cases were included in this study. For definition of PJI, the criteria of the Musculoskeletal Infection Society (MSIS) were considered. The quantity and time to positivity of anaerobes detected in sonicate fluid were monitored both from inoculated supplemented liver thioglycollate broth and anaerobic blood culture bottles. Furthermore, phenotypic testing was performed on the antimicrobial activity within the sonicate fluid. RESULTS: The most frequently isolated microbes were Cutibacterium species, followed by Finegoldia magna, Parvimonas micra, Robinsoniella peoriensis, Clostridium species, Peptoniphilus harei and Slackia exigua. In 24 cases, the microorganisms became detectable within five days (median time 3.2 days) when sonicate fluid was incubated in supplemented liver thioglycollate broth, regardless of whether the patients had taken antimicrobial agents prior to surgery. However, when sonicate fluid was inoculated into anaerobic Bactec bottles, the median time to positivity was 7.4 days and only 12 cases (43%) were correctly identified. Sixteen cases remained negative after 14 days of incubation. CONCLUSION: Depending on the pathogen, incubation of sonicate fluid using blood culture bottles can support diagnosis of PJI but compared with our culture medium it is less efficient if anaerobes are the suspected cause of infection. Microbiological expertise is therefore indispensable to ensure reliable detection of these microorganisms in PJI until a gold standard for laboratory handling of anaerobes has been established.

Periprosthetic joint infection associated with Mycoplasma hominis after transurethral instrumentation in an immunocompetent patient. Unusual or underestimated? A case report and review of the literature.

Judging by the small number of published cases, periprosthetic joint infections (PJI) caused by Mycoplasma species are regarded as unusual. This is not surprising as special growth conditions are necessary for diagnosis and therefore the laboratory must be informed of any clinical suspicion. However, surgeons are generally not aware of the risk factors associated with certain microorganisms causing an infection. Our laboratory therefore decided to adopt a new strategy: first, to address specific questions concerning the medical history of the patient and second, to make diagnosis of rare and fastidious microorganisms part of routine investigation, even if detailed information is not available.

Slackia exigua, an anaerobic Gram-positive rod and part of human oral microbiota associated with periprosthetic joint infection of the hip. First case and review of the literature.

We report on the first case of a periprosthetic joint infection with the anaerobic Gram-positive rod Slackia exigua as the causative agent. The bacterium is part of human oral microbiota and has so far mainly been associated with periodontal diseases.

Whole genome analyses of CMY-2-producing Escherichia coli isolates from humans, animals and food in Germany.

BACKGROUND: Resistance to 3rd-generation cephalosporins in Escherichia coli is mostly mediated by extended-spectrum beta-lactamases (ESBLs) or AmpC beta-lactamases. Besides overexpression of the species-specific chromosomal ampC gene, acquisition of plasmid-encoded ampC genes, e.g. blaCMY-2, has been described worldwide in E. coli from humans and animals. To investigate a possible transmission of blaCMY-2 along the food production chain, we conducted a next-generation sequencing (NGS)-based analysis of 164 CMY-2-producing E. coli isolates from humans, livestock animals and foodstuff from Germany. RESULTS: The data of the 164 sequenced isolates revealed 59 different sequence types (STs); the most prevalent ones were ST38 (n = 19), ST131 (n = 16) and ST117 (n = 13). Two STs were present in all reservoirs: ST131 (human n = 8; food n = 2; animal n = 6) and ST38 (human n = 3; animal n = 9; food n = 7). All but one CMY-2-producing ST131 isolates belonged to the clade B (fimH22) that differed substantially from the worldwide dominant CTX-M-15-producing clonal lineage ST131-O25b clade C (fimH30). Plasmid replicon types IncI1 (n = 61) and IncK (n = 72) were identified for the majority of blaCMY-2-carrying plasmids. Plasmid sequence comparisons showed a remarkable sequence identity, especially for IncK plasmids. Associations of replicon types and distinct STs were shown for IncK and ST57, ST429 and ST38 as well as for IncI1 and ST58. Additional ß-lactamase genes (blaTEM, blaCTX-M, blaOXA, blaSHV) were detected in 50% of the isolates, and twelve E. coli from chicken and retail chicken meat carried the colistin resistance gene mcr-1. CONCLUSION: We found isolates of distinct E. coli clonal lineages (ST131 and ST38) in all three reservoirs. However, a direct clonal relationship of isolates from food animals and humans was only noticeable for a few cases. The CMY-2-producing E. coli-ST131 represents a clonal lineage different from the CTX-M-15-producing ST131-O25b cluster. Apart from the ST-driven spread, plasmid-mediated spread, especially via IncI1 and IncK plasmids, likely plays an important role for emergence and transmission of blaCMY-2 between animals and humans.

Periprosthetic joint infection caused by anaerobes. Retrospective analysis reveals no need for prolonged cultivation time if sensitive supplemented growth media are used.

BACKGROUND: In microbiological diagnosis of periprosthetic joint infection (PJI) culture media and incubation time are controversially discussed, especially if anaerobic bacteria are the causative agent. This study was conducted to demonstrate the influence of sensitive supplemented growth media on the duration of culturing anaerobes. METHODS: Twenty-five consecutive cases were included in this retrospective study. For definition of PJI, the criteria of the Musculoskeletal Infection Society (MSIS) were considered. Histopathological analysis was interpreted according to the classification by Krenn et al. The quantity and time to positivity of detected anaerobes were monitored. Furthermore, antimicrobial activity within the tissue and sonicate fluid was phenotypically tested. RESULTS: In all cases, even if the patients had received antibiotics before recovery, culture of anaerobes (Propionibacterium species, Finegoldia magna, Parvimonas micra and Robinsoniella peoriensis), both from tissue samples and prosthetic components, first became detectable in supplemented liver thioglycollate broth within six days (median: four days). CONCLUSION: Recommendations for prolonged cultivation for up to 14 days mostly aim at detection of anaerobes. Here we present a laboratory procedure that can shorten cultivation time considerably.

Protracted Regional Dissemination of GIM-1-Producing Serratia marcescens in Western Germany.

The metallo-beta-lactamase GIM-1 has been found in various bacterial host species nearly exclusively in western Germany. However, not much is known about the epidemiology of GIM-1-positive Serratia marcescens Here we report on a surprisingly protracted regional dissemination. In-hospital transmission was investigated by using conventional epidemiological tools to identify spatiotemporal links. Strain typing was performed using pulsed-field gel electrophoresis (PFGE) and whole-genome sequencing (WGS). Bayesian phylogeny was used to infer the time axis of the observed occurrence. Thirteen S. marcescens strains from 10 patients from 6 different German hospitals were investigated. Suspected in-hospital transmissions were confirmed by molecular typing at a higher resolution by WGS than by PFGE. A detailed sequence analysis demonstrated the spread of one predominant strain variant but also provided evidence for transfer of the blaGIM-1 gene cassette between different strains. A Bayesian phylogenetic analysis showed that the most recent common ancestor of the identified clonal cluster could be dated back to April 1993 (95% highest posterior density interval, January 1973 to March 2003) and that this strain might have already harbored the blaGIM-1 at that time and, therewith, years before the first detection of this resistance gene in clinical specimens. This study shows a long-standing clonal and plasmid-mediated expansion of GIM-1-producing S. marcescens that might have gone unnoticed in the absence of a standardized and effective molecular screening for carbapenemases. The systematic and early detection of resistance is thus highly advisable, especially for the prevention of potentially long-term dissemination that may progress beyond control.

Molecular Investigation of Carbapenem-Resistant Acinetobacter spp. from Hospitals in North Rhine-Westphalia, Germany.

Emergence of carbapenem-resistant Acinetobacter spp., especially Acinetobacter baumannii, in hospitals has been increasingly detected worldwide. In the present study, we analyzed carbapenem-resistant isolates (70 A. baumannii and one Acinetobacter pittii) collected in a period of 4 years (February 2008 to January 2012) in one diagnostic laboratory in Germany. All isolates were carbapenemase positive with OXA-23 as by far the most common enzyme (n = 66, 93%). Carbapenemases OXA-24-like and OXA-58 were not present in the isolates, but genes blaGIM-1 and ISAba1+blaOXA-80/82 were found to be the cause of carbapenem resistance in one and four isolates, respectively. Polymerase chain reaction typing revealed that the majority of A. baumannii isolates could be assigned to the very successful international clone 2. ApaI-macrorestriction and pulsed-field gel electrophoresis (PFGE) indicated clonal transmission of resistant strains (eight different PFGE types) within several hospitals. By multilocus sequence typing, the isolates were to be assigned to ST195 (n = 44), ST236 (n = 12), ST208 (n = 4), ST437 (n = 3), ST231 (n = 3), ST448 (n = 2), ST556 (n = 1), and ST945 (n = 1). The wide spread of carbapenem-resistant clones of A. baumannii is facilitated by international travelling and needs continuous surveillance in hospitals and diagnostic laboratories.

Robinsoniella peoriensis, originally isolated from swine manure, and early periprosthetic hip infection: Case report and review of the literature.

We report on the first case of a periprosthetic joint infection with the anaerobic spore-forming Gram-positive rod Robinsoniella peoriensis as the causative agent. The bacterium was first isolated from a swine manure storage pit and has so far rarely been associated with human infections.

Emergence of metallo-ß-lactamase GIM-1 in a clinical isolate of Serratia marcescens.

The metallo-ß-lactamase GIM-1 (German imipenemase) has been found so far only in clinical isolates of Pseudomonas aeruginosa from Germany. Here we report the detection of bla(GIM-1) in a clinical strain of Serratia marcescens that was isolated from urine, blood, and wound samples over a period of 20 months. The strain was repeatedly isolated from one patient in two German hospitals and an outpatient department located in the region in which all previously described GIM-1-producing P. aeruginosa strains were identified.

Actinomyces neuii subsp. neuii Associated with periprosthetic infection in total hip arthroplasty as causative agent.

Actinomyces neuii has until now not been described as a pathogen associated with periprosthetic infection in total joint replacement. The case presented here suggests that A. neuii subsp. neuii is a causative pathogen. The discussion and review of the literature indicate the impact that detection of Actinomyces species could have.