Aim. A significant number of patients undergoing shoulder arthroplasty surgery have C acnes contamination at the end of the primary surgery. The objective of this study is to determine whether patients with C acnes contamination at the end of their primary shoulder surgery have a worse prognosis than those who end up without C. acnes contamination. Method. Prospective study including all patients who underwent a reverse shoulder prosthesis from January 2015 to December 2018. In all of them, 5 to 12 cultures were performed during primary surgery. The patients underwent surgery for shoulder arthritis secondary to rotator cuff tears, acute fracture of the proximal humerus, and sequelae of fracture of the proximal humerus. Exclusion criteria included the existence of previous surgeries on the affected shoulder, the presence of signs of infection, having received infiltrations and / or complementary invasive examinations (Arthro-MRI and Arthro-CT). Follow-up from 2 to 5 years. Functional assessment according to the Constant Functional Scale. All complications were also recorded. Results. 162 patients were included. Of these, 25 had positive cultures for C. acnes at the end of primary shoulder surgery. Average age of 74.8 years. 136 women and 26 men. 75.9% Shoulder arthritis secondary to rotator cuff tears, 13.6% acute fractures and 10.5% sequelae of fractures. There were no differences between patients with C. acnes and those without C. acnes regarding age and indication for surgery. Predominance of men in the group with positive C. acnes (p <0.001). No differences at 2 and 5 years in the Constant functional scale between the two groups (2 years, 59.6 vs 59.2 p 0.870) (5 years, 62.4 vs 59.5 p 0.360). Significant differences regarding the number of complications (p 0.001). Patients without C. acnes had 1 aseptic loosening of the metaglene and patients with C. acnes had 2 infections, 1 dislocation, and 1 revision surgery. Patients with contamination by C. acnes had more comorbidities (p 0.035) than patients without contamination. Conclusions. Patients with C acnes contamination at the end of primary surgery do not have functional differences when compared with patients without contamination at 2 and 5 years, but they have a higher number of complications in the medium term

Background. Although described as a commensal bacterium with low pathogenicity, Cutibacterium acnes involvement has been reported in many clinical entities: infections associated with devices, such as shoulder prosthetic joint infections, osteosynthesis, breast implants or cerebrospinal fluid shunts. Various studies show that C. acnes grows as a biofilm, contributing to its persistence by allowing its escape from the action of the immune system and antibiotics. Purpose. Our aim was to assess the activity of different active substances (erythromycin, clindamycin, doxycycline and Myrtacine. ®. ) on eight different well-characterized C. acnes strains after growth in biofilm mode. Methods. Eight susceptible strains of C. acnes were selected for this study, including two reference strains (ATCC6919 and ATCC11827) and six clinical strains. All C. acnes strains were studied using two different methods to study the biofilm production at different time points: the BioFilm Ring Test. ®. technique (early stages of adhesion) and the Crystal Violet (CV) method (mature biofilm). In a second step, the impact of different active substances (erythromycin, clindamycin, doxycycline and Myrtacine. ®. ) was studied. For the CV technique, two types of tests were performed: preventive tests (addition of active substances and bacteria at the same time) and curative challenge tests (addition of active substances on a biofilm already formed after 48h). Transmission electron microscopy was performed to investigate the morphology modifications. Results. C. acnes isolates from phylotypes IA. 1. and IA. 2. , seem to produce more mature biofilm in the first stages of adhesion than other phylotypes. Curative assays were performed to evaluate the efficacy of antibiotics and Myrtacine. ®. on mature biofilm. Significant efficacy of Myrtacine. ®. at 0.03% was observed for C. acnes strains. Moreover, the combination of Myrtacine. ®. and doxycycline appears to decrease the total biofilm biomass. The effect of doxycycline as a preventive measure was minimal. On the contrary, a similar use of Myrtacine. ®. as early as 0.001% showed significant efficacy with a significant decrease in total biofilm biomass for all C. acnes strains. Transmission electron microscopy revealed a significantly decreased biofilm growth in treated bacteria with Myrtacine. ®. compared to untreated bacteria. Moreover, the total number of bacteria decreased as the concentration of Myrtacine. ®. increased suggesting also an antimicrobial effect. Conclusion. These results confirm the difference in biofilm producing ability depending on C. acnes phylotypes. These results suggest that Myrtacine. ®. may be a promising alternative antibacterial and anti-biofilm agent like peroxide de benzoyle to prevent shoulder prosthetic joint infection involving planktonic and biofilm C. acnes

Aim. Cutibacterium acnes is a major skin commensal that may also act as an opportunistic pathogen. Findings of C. acnes in tissue cultures obtained during arthroplasty revision surgery are difficult to interpret, since they may represent true infection or contamination. This study investigated whether C. acnes isolates obtained from prosthetic joint infections (PJIs) were related and shared common genomic traits that might correlate with clinical courses and patient outcomes. Method. C. acnes isolates from revision surgery of patients with PJIs of the hip, shoulder, and knee were characterized using molecular methods to determine sequence type (ST) and the presence of virulence determinants (CAMP factors, dermatan sulfate-binding adhesion 1, hyaluronidase lyase, and linear plasmid). A standardized review of the patients’ medical charts was performed. Results. The study included 37 patients with C. acnes culture-positive tissue samples where multiple isolates of C. acnes belonged to the same ST. Most of the isolates belonged to phylotype IA. 1. Phylogenetic analysis of virulence determinants revealed no shared pattern among PJI isolates. Seven patients had a polymicrobial infection. Exchange revision was performed in 70% of the patients, and >50% of all patients received antibiotic treatment for ≥3 months. Failure was noted in seven patients, all of whom had shoulder PJIs. Conclusions. No specific ST or any identifiable unique feature among virulence determinants were found among C. acnes isolated from PJIs of hips and shoulders. The majority of all included patients had low inflammatory markers and were treated successfully, even when the infection consisted of a polymicrobial infection

Aim. Cutibacterium acnes (C. acnes) is the most cultured organism implicated in periprosthetic shoulder infections. Nevertheless, the clinical significance of its persistence on the skin surface and in the deep layers during shoulder arthroplasty surgery remains still unknown. The purpose of this study was to know if the C. acnes isolate present in deep tissues at the end of a primary shoulder arthroplasty could be responsible for shoulder arthroplasty infection. Method. Prospective study including 156 patients undergoing primary shoulder arthroplasty. In all the patients included 5 to 12 tissue samples were obtained and were specifically cultured to detect C. acnes presence. DNA was extracted from the C. acnes colonies selected with the QIAsymphony DSP Virus/Pathogen Midi Kit (Qiagen, Hilden, Germany). Libraries were prepared using Nextera XT kit (Illumina) and sequenced in an Illumina MiSeq sequencer. Sequencing files were pre-processed using The Microbial Genome Atlas pipeline. Samples that failed on QC analysis were discarded for further analysis. Isolate nucleotide distances were calculated using Genome-based distance matrix calculator from the enveomics collection. Comparative genomic analysis was performed between intra- and inter-patients’ isolates. Data analysis was performed using R 3.6.3. Results. For twenty-seven out of 156 patients (17.31%), C. acnes was present at the end of the primary surgery. Two of these patients (both male) developed a C. acnes periprosthetic shoulder infection after 6 and 4 months from the primary surgery. DNA from the C. acnes responsible for the periprosthetic infection was further analysed by whole genome sequencing (WGS). Average Nucleotide Identity (ANI) value was assessed, measuring the nucleotide-level genomic similarity between genome pairs. We found a clear ANI clustering in two major groups which corresponded, mainly, to the associated phylotype (97%–98% ANI). Moreover, when analysing both isolates that developed a periprosthetic shoulder infection, we found that all the revision-surgery isolates clustered nearer to their corresponding primary-surgery isolates (99.4% of similarity) than to the other independent bacterial isolates, supporting the causal relationship between the initial and the delayed infection. Conclusions. C. acnes present at the end of the primary surgery can be the cause of early- or delayed-periprosthetic joint infections in shoulder arthroplasty, revealing the potential route of infection. Therefore, efforts must be made in terms of antibiotic prophylaxis and skin preparation to limit infections of total shoulder arthroplasties

Aim. The aim of our study was to analyze putative genes for virulence factors of Cutibacterium isolates obtained from implant-associated infections. Methods. We analyzed 64 isolates of Cutibacterium spp. (C. acnes (53/64), C. avidum (6/64), C. granulosum (4/64), C. namnetense (1/64)) using NextSeq 550 (Illumina, San Diego, CA, USA) and performed genomic analysis of 24 genes associated with virulence factors (VFs) of C. acnes previously reported in the literature. Most isolates were obtained from implant-associated infections (IAI) between 2012–2021 at the Institute of Microbiology and Immunology, Faculty of Medicine, Ljubljana. Additionally, we included the first C. namnetense isolated in our laboratory from surgical site infection. Results. C. acnes and C. namnetense have the highest number of VFs among those examined. The VFs gntK (shikimate kinase) and HYL-IB / II (hyaluronate lyase) are absent in phylotype IA. 1. (sequence types (ST) A, C, D according to the SLST scheme). Repressor gene of porphyrin synthesis, deoR is present in all Cutibacterium spp. isolates. The phylotypes II and IB show a similar distribution of VFs, with the presence of the VFs rcsB (compound for biofilm formation) and HYL-IA (hyaluronate lyase), which are absent in other C. acnes phylotypes and other Cutibacterium spp. In phylotypes IA. 1. and IB, the sequence of genes encoding VFs dsA1 and dsA2 does not have 100% genomic coverage, possibly indicating homologs between species. The isolates of C. acnes and C. namnetense possess all three CAMP (1,2,4) factors, which are not detected in other Cutibacterium spp. However, further analysis revealed species-specific CAMP factors in C. avidum and C. granulosum. Both species also have similar other genes for VFs, mainly encoding heat shock proteins and lipases, while VFs related to biofilm production are mostly absent (rcsB, ytpA). Conclusion. We found several differences in the distribution of VFs among Cutibacterium spp. isolated from IAI

Aim. Cutibacterium acnes is a significant cause of late-onset spinal implant infection (SII). In addition, usual preoperative prophylactic measures may be insufficient to prevent C. acnes operating site colonisation and infection, as demonstrated for prosthetic shoulder surgery. However, little information is available regarding risk factors for SII due to this microorganism. The aims of this study were to determine the characteristics of and risk factors for C. acnes SII. Method. we conducted a retrospective unmatched case-control study including all adult patients treated for mono and polymicrobial C. acnes SII during 2010–2015. Controls were randomly selected among patients diagnosed with SII due to other microorganisms during the same period. Results. Fifty-nine patients with C. acnes SII were compared with 59 controls. There was no difference in sex distribution (39% vs 53% men). Patients with C. acnes SII were younger (median age 42 vs. 65, p< 0.001), thinner (median body mass index (BMI) 21 vs. 25 kg/m. 2. , p< 0.001), and presented a better health status (ASA score≤ 2, 83% vs. 65%, p= 0.015; and presence of immunosuppression, 3% vs. 27%, p= 0.002). Patients with C. acnes SII were more likely to experience delayed/late infections (i.e. diagnosed >3 months post-instrumentation, 66% vs. 22%, p< 0.001) and to be instrumented for scoliosis (83% vs. 27%, p< 0.001) with an extended osteosynthesis (median number of fused vertebrae 12 vs. 5, p< 0.001). However, 20 C. acnes SII (34%) developed early (≤3 months) after instrumentation. The clinical presentation was significantly more indolent in the C. acnes group (presence of fever, 27% vs. 61%, p= 0.001; wound inflammation 39% vs. 61%, p< 0.001 and median C-reactive protein level 38 vs. 146 mg/L). Mixed C. acnes SII were diagnosed on 24 occasions (41%), 22 of which involving both C. acnes and staphylococcal strains. In the multivariate logistic regression model, factors independently associated with the development of SII involving C. acnes were age less than 65 (adjusted odds ratio [aOR] 7.13, 95% CI [2.44–24.4], p= 0.001), BMI< 22kg/m. 2. (aOR 3.71 [1.34–10.7], p= 0.012) and a number of fused vertebrae >10 (aOR 3.90 IC 95% [1.51–10.4], p= 0.005). Conclusions. There were significant differences between SII involving C. acnes and those involving other microorganisms. We identified a specific profile of patients at increased risk of developing C. acnes SII. These findings could contribute to improve both the prevention and treatment of such infections

Aim. Cutibacterium acnes is involved in chronic/low-grade pathologies such as prosthetic joint infection (PJI) or sarcoidosis. During these diseases, granulomatous structures are frequently observed. In this study, we induced a physiological granulomatous reaction in response to different well-characterized clinical C. acnes isolates in order to investigate the cellular process during the granulomatous formation. Method. An acne C. acnes ATCC6919 isolate (clonal complex (CC) 18, phylotype IA1), a PJI C. acnes BL clinical isolate (CC36, phylotype IB) and a sarcoidosis C. acnes S8 strain (CC28, phylotype IA. 2. ) were included and co-culture with PBMC. Cellular aggregation was followed daily using light microscopy. At various time points of incubation (day 3 and day 7), granuloma structures were processed for microscopic observation, colony forming unit enumeration after Triton ×100 lysis to release the internalized bacteria (bacterial load within the granulomas), as well as for flow cytometric analysis (detection of CD4, CD8 and NK lymphocytes). Results. All C. acnes isolates generated granulomatous structures in our experimental conditions. The bacterial burden was better controlled by granulomas induced by sarcoidosis C. acnes isolate. PJI C. acnes isolate, belonging to CC36, promoted the recruitment of CD8. +. lymphocytes inside the granuloma. At the opposite, acne and sarcoidosis C. acnes isolates, belonging, respectively, to phylotype IA. 1. /CC18 and phylotype IA. 2. /CC28 generated a higher number of granuloma and promoted the recruitment of CD4. +. lymphocytes inside the granuloma. Conclusions. Our results provide new arguments supporting the role of C. acnes in the development of infections and new explanations concerning the mechanisms underlying PJI due to C. acnes. This model appears to be a possible alternative assay to animal models for studying the immune response to C. acnes infection

Aim. The aim of this study was to confirm that Mirra's criterion (≥ 5 Polymorphonuclears (PMNs) per field in 5 high power fields (HPFs)) is not adequate for diagnosis of chronic bone and joint infections (BJIs) due to Cutibacterium acnes (C. acnes). The second objective was to determine if plasma-cell infiltration, that is a classical marker of chronic inflammation, could be useful for the diagnosis of chronic BJIs due to C. acnes. Methods. We retrospectively selected 25 patients from 2009 to 2013 with chronic BJIs due to C. acnes. In addition of Mirra's criterion, the number of plasma-cells (≥5 plasma-cells/5 HPFs, defined as “CRIOAc Lyon's criterion”) was implemented in the histopathological analysis. Patients were defined as infected, if at least one of the two criteria were present. Results. According to Mirra's and CRIOAc Lyon's histopathological criteria, positive histology was observed in respectively 12 (48%) and 16 (64%) cases. In 1 case the samples were not analyzable. Considering the 12 cases with negative Mirra's criterion, high plasma-cell infiltration (≥5 plasma-cells/5 HPFs; Figure 1) was observed in 6 cases (50%), and low plasma-cells infiltration (2–5 plasma-cells/5 HPFs) was observed in 5 other cases (42%). Conclusions. Mirra's criterion is not an adequate criterion to defining chronic BJIs [1, 2]. In our study, more cases of chronic BJIs due to C. acnes have been diagnosed using CRIOAc Lyon's criterion than Mirra's criterion. Adding CRIOAc Lyon's criterion might restore some histopathological diagnosis of chronic BJIs due to C. acnes, when a clinical chronic BJI is suspected. For any tables or figures, please contact the authors directly

Aim. Cutibacterium acnes, a skin commensal, is responsible for 5–10% of prosthetic joint infections (PJI). All current microbiological definitions of PJI require two or more identical commensal isolates to be recovered from the same procedure to diagnose PJI and rule out contamination. Unlike coagulase negative staphylococci, C.acnes shows a highly stereotypical susceptibility profile making impossible to phenotypically assess the clonal relationship of isolates. In order to determine the clonal relationship of multiple C.acnes isolates recovered from arthroplasty revisions, we analyzed by multi-locus sequence typing (MLST) C.acnes isolates grown from orthopedic device-related infections (ODRI) in a reference center for bone and joint infection. Methods. Laboratory records from January 2009 to January 2014 were searched for monomicrobial C.acnes ODRI with growth of C. acnes in at least 2 intraoperative and/or preoperative samples. Clinical, biological and demographic information was collected from hospital charts. All corresponding isolates biobanked in cryovials (−80°C) were subcultured on anaerobic blood agar, and identification confirmed by MALDI-TOF-MS. C.acnes isolates were typed using the MLST scheme described by Lomholt et al. Plasmatic pre-operative C-reactive protein (CRP) levels were determined using DimensionEXL (Siemens). A threshold of 10 mg/L was used to determine serologically positive ODRIs from negatives. Results. Over a 5-year period, 37 cases of monomicrobial C.acnes ODRI were diagnosed in our center. Among these 37 cases, 113/153 C.acnes isolates were cryopreserved. 110/113, corresponding to 36/37 cases, were typed by MLST: 14/36 (39%) ODRI cases were found to feature isolates belonging to two or more different STs and were qualified to be heteroclonal whereas 22/36 (61%) of ODRI cases were found to feature isolates belonging to the same ST and were qualified to be homoclonal. Homoclonal infections were significantly more likely to have elevated CRP levels compared to heteroclonal cases (p=0.0011, Fisher test). Patients with only two positive intraoperative samples had significantly lower CRP values than patients with three or more positive intraoperative samples (12,7mg/L vs 67mg/L; p=0,01, homoscedastic two-tailed Student's t test). Conclusions. This study suggests that what is classified microbiologically as C.acnes ODRIs comprises: i) true homoclonal infections eliciting an inflammatory response, ii) heteroclonal infections lacking inflammatory response where C.acnes could be an innocent bystander and iii) false positives where no strain achieves true microbiological significance. Our study shows that a stricter threshold of 3 intraoperative positive samples could be more adequate than 2. These results reinforces the need for a more specific definition of C.acnes ODRI

Aim. The aim of this study was to develop an in-house multiplex PCR real-time assay on the LightCycler 480 system (Roche, Basel, Switzerland) with the aim of rapid detection of common pathogens in prosthetic joint infections (PJI), followed by validation on clinical samples (sonication fluid and tissue biopsies) routinely collected for PJI diagnosis. Methods. Using the PrimerQuest and CLC WorkBench tool, we designed six primer sets with specific fluorescently labelled TaqMan probes for the nuc gene in different Staphylococcus species (S. aureus, S. epidermidis, S. capitis, S. lugdunensis, S. hominis, S. haemolyticus). In addition, primers previously developed by Renz et al. (2022) for C. acnes were integrated into our assay with internal control of isolation, leading to the development of specific mPCR assay with seven included targets. Analytical sensitivity and specificity were evaluated using reference bacterial strains. To determine the assay's limit of detection (LOD), we conducted serial dilutions of eluates containing known concentrations of bacterial DNA copies/µl. The overall LOD in spiked clinical samples, including sample preparation and DNA isolation on MagnaPure24, was measured through 10-fold serial dilutions (from 10. 9. to 10. -1. CFU/ml) including additional dilutions of 5000, 500, 50 and 5 CFU/ml. Results. The results with LOD in serial dilutions of eluates and spiked clinical samples, together with analytical sensitivity and specificity, are shown in Table 1. Conclusion. The mPCR assay showed excellent analytical sensitivity and specificity, but with considerably lower LOD after sample preparation and further DNA isolation in spiked clinical samples. Although still promising in diagnostics of acute infections, the use of mPCR could be challenging in chronic, low-grade infections with lower microbial burden. Nevertheless, PCR offers significant advantages in terms of speed and can shorten the time to result, especially for C. acnes infections. Additionally, it represents a promising complementary approach in patients with suspected PJI on antibiotic therapy with negative culture results. For any tables or figures, please contact the authors directly

Aim. The aim of our study was to identify pathogens involved in septic knee arthritis after ACLR and to describe clinical features, treatment and outcome of infected patients. Methods. We conducted a retrospective observational study including all patients with ACLR infection in 3 orthopedic centers sharing the same infectious disease specialists. Results. During a seven-year period (2011–2017) we identified 74 infected patients among 9858 patients who had ACLR (incidence rate = 0.0075). Fourteen patients had polymicrobial infection. We identified 89 pathogens. Twenty four patients (34.4 %) were infected with S. aureus (27% of all isolates)(only one oxacillin-resistant strain). C. acnes was the second most frequent pathogen, identified in 14 patients (18.9%) (15.7% of all isolates). S. lugdunensis was identified in 9 patients (12.2%) (10.1% of all isolates). S. caprae was as frequent as S. epidermidis identified in 8 patients each (10.8%) (9 % of all isolates for each). No strain of S. lugdunensis and S. caprae was resistant to oxacillin, levofloxacin or rifampicin. Ten patients infected by C. acnes, 8 infected by S. lugdunensis, and 7 infected by S. caprae had an early acute infection. In all cases but one an arthroscopic lavage was performed, in 14 cases two lavages were required and in 4, 3 lavages. All patients infected by a strain susceptible to levofloxacin and rifampicin, including those with C. acnes, S. caprae and S. lugdunensis infection, were treated with an oral combination of levofloxacin and rifampicin, after a couple of days of IV empirical treatment with vancomycin and a broad spectrum beta-lactam. The median duration of treatment was 6 weeks. Seventy one patients were considered cured. Conclusions. To our knowledge this is the largest reported series of infection after ACLR. S. aureus is the main pathogen (27% of all strains). C. acnes, S. lugdunensis and S. caprae accounted for almost 35% of pathogens and 38% of infections. A conservative strategy consisting in arthroscopic lavage(s) and a 6-week treatment with levofloxacin and rifampicin was effective

Aim. To evaluate whether sonication of implant material and subsequent culturing add clinical relevance to culturing of tissue biopsies for improved antibiotic treatment in treatment of bone and joint infection. Method. A retrospective examination of patients’ charts and microbiological analyses in patients who had explanted material (plates, screws, k-wires and prostheses) send for sonication between December 2020 and April 2022. Results. 77/143 (54 %) patients had complete agreement between the cultures from tissue biopsies and sonication fluid. 66/143 (46 %) patients had partial or no agreement between the cultures from tissue biopsies and sonication fluid. Of the 66 patients, 31 (47 %) had a culture positive sonication fluid and tissue biopsies that were positive with one or more bacterial isolates. 26/66 (39 %) patients had a culture positive sonication fluid and tissue biopsies that were negative. 9/66 (14 %) patients had negative sonication fluid and positive tissue biopsies. Of the 26 patients with culture positive sonication fluid and culture negative tissue biopsies, virulent bacteria were found in 5 (19 %) patients, making the diagnosis and treatment of infection straight forward. The remaining 21 (81 %) patients had C. acnes, S. epidermidis and CoNS in the sonication fluid, which made the diagnosis less evident but none the less gave the clinician a relevant treatment option. Conclusion. In this study a high concordance was found between cultures from tissue biopsies and sonication fluid. Additionally, in a small group of patients with culture negative tissue biopsy, the culture of sonication fluid was essential to the identification infections agent. This indicates that culture of sonication fluid is an important diagnostic tool in bone and joint infection, especially in the absence of positive tissue cultures

Aim. Periprosthetic joint infection (PJI) is one of the most devastating complications after joint replacement. It is associated with high morbidity and economic burden when misdiagnosed as an aseptic failure. Among all cases of PJI, up to 25% could yield negative cultures. Conversely, among cases of aseptic failures, up to 30% may actually be undiagnosed PJIs. In PJIs microbiological diagnosis is a key step for successful treatment. Sonication of the removed prosthesis is more sensitive than conventional periprosthetic-tissue culture, especially in patients who received antimicrobial therapy before surgery. This study aimed to compare the diagnostic value of classic sonication fluid cultures (SF-C) and sonication fluid incubation in blood culture bottle (SF-BCB). Method. Between 2016 and 2018 we analysed 160 revision procedures of joint arthroplasties. For each procedure, at least 5 microbiological and multiple histopathological samples were harvested, and explant sonication was performed which was further analysed by SF-C and SF-BCB. For SF-C classical cultivation of sonication fluid was performed. While for SF-BCB, 10 mL of sonication fluid was inoculated into aerobic and anaerobic lytic blood culture bottles. The definite diagnosis of PJI was based on the EBJIS definition. Results. Among 160 revisions, 59 PJIs were identified, 15 patients were treated with the debridement and implant retention, 7 patients with the one-stage and 35 with the two-stage exchange, remaining 2 were partial revisions. The sensitivity of SF-C and SF-BCB were 81.5% and 94.9%, respectively. The mismatch of microbe identification was observed in 5 cases. We observed positive SF-C while negative SF-BCB in 4 cases, among them having 2 positive histology. While 12 patients have negative SF-C and positive SF-BCB, among them 3 have positive and 6 negative histology. Among these 12 patients, typical low-grade microbes were identified in 9 cases (5 cases of C. acnes, 3 cases of S. epidermidis, and 1 case of S. capitis). Conclusions. The weakest point in all PJI diagnostic criteria is their sensitivity. SF-BCB demonstrates higher sensitivity in diagnosing PJI compared to SF-C. Therefore, it appears prudent to incorporate SF-BCB into the diagnostic protocol for all patients exhibiting either low-grade PJI symptoms or experiencing undiagnosed, presumably aseptic failures, where the likelihood of misdiagnosing infection is greatest

Aim. Diagnosing or excluding a chronic prosthetic joint infection (PJI) prior to revision surgery can be a clinical challenge. To enhance accuracy of diagnosis, several biomarkers were introduced in recent years, but most are either expensive or not available as a rapid test. We compared the diagnostic accuracy of leucocyte esterase (€0.20 per sample), calprotectin (€20 per sample) and alpha defensin (€200 per sample). Method. We prospectively evaluated PJI patients with chronic pain with or without prosthetic loosening between 2017 and 2018. Synovial fluid was collected prior to revision surgery. Leucocyte esterase was measured using a reagent strip (2+ considered as positive), and calprotectin and alpha defensin were measured using a lateral flow immunoassay. Intraoperative cultures (5 periprosthetic tissue samples, synovial fluid and sonication fluid) incubated for 9 days, were used as gold standard. At least two positive cultures of low-grade microorganisms with the same antibiogram were required to diagnose PJI. Results. A total of 19 patients were included (knee =11, hip =8). None of the patients were treated with antibiotics prior to revision surgery. A PJI was diagnosed in 8 patients (42.1%). The diagnostic accuracy of leucocyte esterase vs. calprotectin vs. alpha defensin was as follows; sensitivity 50.0% vs. 87.5% vs. 87.5%, specificity 81.8% vs. 90.9% vs. 100%, positive predictive value 60.0% vs. 87.5% vs. 100% and negative predictive value 75.0% vs. 90.9% vs. 91.6%, respectively. Both calprotectin and alpha defensin were false negative in one PJI caused by Cutibacterium acnes. The other two C. acnes PJIs were correctly diagnosed with both tests. Conclusions. Calprotectin is as accurate as alpha defensin in excluding a chronic PJI at 10% of the costs. Future studies with a large number of patients are necessary to analyze its diagnostic accuracy in very low-grade infections, in particularly caused by C. acnes

Aim. Arthroscopic interventions have revolutionized the treatment of joint pathologies. The appropriate diagnostics and treatment are required for infections after ligament reconstructions using non-resorbable material such as tendon grafts, anchors, and sutures, prone to biofilm formation. The infection rate is around 1% for knee and shoulder, while up to 4% for Achilles tendon reconstructions. Despite high number of these procedures worldwide, there is limited evidence about the best treatment protocol. Our study aimed to provide a general protocol for the treatment of small implants for soft tissue reconstruction. Method. Between 2019 and 2023, we treated 48 infections of ligament, meniscus, and tendon reconstructions out of 7291 related procedures performed in the same time period. Early infection (<30 days) were treated with an arthroscopic debridement and implant retention (DAIR), except Achilles tendons had open DAIR, while those with delayed or chronic infection (>30 days) were treated with extensive debridement and lavage combined with one-stage exchange (OSE) or implant removal. During surgery, at least 5 microbiological s and samples for histopathology were obtained. The removed material was sonicated. After surgery, all patients were one week on iv. antibiotics, followed by oral antibiofilm antibiotics for 6 weeks including rifampicin and/or a quinolone. All patients were followed for at least 1 year. Failure was defined as the need for additional revision surgery after finished iv. antibiotic treatment. Results. Among 48 patients, 38 were early and 10 were late acute or chronic infections. The incidence of infection for our cohort was 0.7%. We observed 27 infections after ligament reconstruction of the knee, 15 of the shoulder, 5 of the ankle, and 1 infection of the elbow joint. 40 patients were treated with DAIR, 5 with OSE, and 3 with implant removal. We had 11 C. acnes, 10 S. aureus, 6 S. epidermidis, 2 P. aeruginosa, 2 S. lugdunensis, 10 mixed flora, and 3 culture-negative infections. 12 patients received antibiotics before surgery, and all culture-negative infections were related to this subgroup. We observed 2 failures, both in a combination of proximal tibial osteotomy and ligament reconstruction of the knee joint. The success rate of our protocol was 96%. Conclusions. Prompt surgical treatment followed by 6 weeks of antibiotic treatment cured 96% of infections of small implants after reconstruction procedures of knee, shoulder, and ankle joints. Our study is the first to provide a treatment protocol for infections of small implants after ligament reconstruction procedures

Aim. Treatment algorithms for fracture-related nonunion depend on the presence or absence of bacterial infection. However, the manifestation of septic nonunion varies. Low-grade infections, unlike manifest infections, lack clinical signs of infection and present similarly to aseptic nonunion. The clinical importance of low-grade infection in nonunion is not entirely clear. Therefore, the aim of this study was to evaluate the clinical relevance of low-grade infection in the development and management of femoral or tibial nonunion. Method. A prospective, multicenter clinical study enrolled patients with nonunion and regular healed fractures. Preoperatively, complete blood count without differential, C-reactive protein (CRP), and procalcitonin were obtained, clinical signs of infection were recorded, and a suspected septic or aseptic diagnosis was made based on history and clinical examination. During surgical nonunion revision or routine implant removal, tissue samples were collected for microbiology and histopathology, and osteosynthesis material for sonication. Nonunion patients were followed for 12 months. Definitive diagnosis of “septic” or “aseptic” nonunion was made according to diagnostic criteria for fracture-related infection, considering the results of any further revision surgery during follow-up. Results. 34 patients with regular healed fractures were included. 62 nonunion patients were diagnosed as aseptic, 22 with manifest, and 23 with low-grade infection. The positive predictive value was 88% and the negative predictive value 72% for the suspected diagnosis. The nonunion groups had significantly higher CRP levels than the regular healer group. Differentiation between septic and aseptic nonunion based on blood values was not possible. Low-grade infection demonstrated less frequently histopathologic signs of infection than manifest infection (22% vs. 50%, p=0.048), with 15% of regular healers having histopathologic signs of infection. Cutibacterium acnes was less present in manifest compared to low-grade infection (p=0.042). Healing rates for septic nonunion involving C. acnes were significantly lower for manifest infection (20%) than for low-grade infection (100%, p=0.002). Patients with low-grade infection were treated with systemic antibiotics less frequently than patients with manifest infection (p=0.026), with no significant difference in healing rate (83% vs. 64%), which was slightly lower for low-grade infection than for aseptic nonunion (90%). Conclusions. Low-grade infections play a significant role in nonunion development and are difficult to diagnose preoperatively due to the lack of clinical signs of infection and unremarkable blood counts. However, our results imply that for low-grade infections, antibiotic therapy may not always be mandatory to heal the nonunion. This study was supported by the German Social Accident Insurance (FF-FR0276)

Aim. We prospectively evaluated four different microbiological tools for diagnostics of prosthetic joint infections (PJI), and assessed their impact on the categorization of infection according to EBJIS guidelines. We compared culture, in-house real-time mPCR for S. aureus, S. lugdunensis, S. hominis, S. epidermidis, S. capitis, S. haemolyticus, C. acnes (mPCR), broad-spectrum PCR (Molzym) with 16S rRNA V3-V4 amplicon Sanger sequencing (16S PCR), and 16S rRNA V3-V4 amplicon next-generation sequencing (16S NGS) on MiSeq (Ilumina). Methods. A total of 341 samples (sonication fluid, tissue biopsy, synovial fluid) were collected from 32 patients with suspected PJI who underwent 56 revision surgeries at the Orthopaedic Centre University Hospital Ljubljana, between 2022 and 2024. Samples were processed using standard protocols for routine culture, followed by DNA isolation using the MagnaPure24 (Roche). All samples were tested with mPCR, and an additional ≥4 samples from each revision (244 in total) were subjected to further metagenomic analysis. Culture results were considered positive if the same microorganism was detected in ≥2 samples, ≥50 CFU/ml were present in the sonication fluid, or ≥1 sample was positive for a more virulent microorganism or if the patient had received antibiotic treatment. Results. Each tool demonstrated high sensitivity for correct EBJIS categorization (100% culture and 16S NGS, 96.88% mPCR and 16S PCR). The highest specificity was observed with mPCR and 16S PCR (87.5%), while culture (79.17%) and NGS (37.5%) showed lower specificity. In 27% (15/56) of revisions, all microbiological tests were negative, although infection was confirmed with histology in one case, and four cases were classified as infection-likely based on clinical signs. In 20% (11/56) of cases, all microbiological tests were positive; in three cases a combination of other EBJIS criteria (without microbiology) categorized the episodes as infection-likely and one as infection-unlikely, emphasizing the importance of microbiological tests in diagnostic criteria. In 43% (24/56) of revisions categorized as infection-unlikely using a combination of other EBJIS criteria, five had positive culture, and three had positive mPCR and 16S PCR. Fifteen (62%) had positive 16S NGS, 12 due to a low number of reads, which may indicate low-grade infection or possible contamination. Conclusion. To date, no test can be established as the ultimate gold standard. The lack of interpretation criteria can result in low specificity of some methods, as the threshold is difficult to determine. A multidisciplinary approach with combination of microbiological tools is still considered the most efficient

Aim. Prosthetic joint infection (PJI) represents the second most frequent complication of total joint arthroplasty (TJA) with up to 20% of low-grade PJI treated as aseptic failure. Sensitive diagnostic criteria have been provided by EBJIS. However, to date there is no single test to reliably diagnose all PJIs. Studies of Mazzucco et al. and Fu et al. suggest that synovial fluid (SF) viscosity could be considered as an important marker for PJI. The primary aim of our study was to determine if SF viscosity is a more reliable diagnostic criterion of PJI than the SF cell count with differential (CCD), and the combined diagnostic value of SF viscosity and CCD. Method. We prospectively analysed the viscosity of SF samples obtained during TJA of hip and knee revisions. We sampled 2.5–5mL of SF for viscosity and CCD. Intraoperatively, 1mL of the sample was analysed for the CCD. The remaining SF was centrifuged for 4min at 7000rpm. The viscosity of the supernatant was determined on Ostwald viscometer as the time required to pass the viscometer at 20°C. During each surgery at least 5 microbiological and multiple histopathological samples were harvested, and explant sonication was performed. The diagnosis was based on EBJIS definition. The viscosity threshold for detecting PJI was set at 65 seconds. Results. Between December 2020 and January 2023, we analysed 65 knee and 47 hip TJA revision procedures. There were 55 septic and 57 aseptic diagnoses. As a diagnostic marker of PJI, SF viscosity achieved 100% sensitivity and 82.5% specificity, with area under the receiver operating characteristic curve (AUC) of 0.832 (95% CI 0.739, 0.925). The specificity and sensitivity of SF CCD were 98.2% and 78.2%, respectively, with AUC of 0.921 (95% CI 0.869, 0.974). Of the 10 cases incorrectly diagnosed as aseptic based on SF viscosity, 2 were acute traumas and 8 metalloses. The SF CCD in all these cases was <0.5. Of the 12 cases incorrectly diagnosed as aseptic based on SF CCD, 6 cases were culture negative, 4 C. acnes and 2 S. epidemidis isolates in microbiology. Taken together, SF viscosity and CCD achieved a combined AUC of 0.953 (95% CI 0.919, 0.987). Conclusions. Our study is the first to report that SF viscosity is more sensitive but slightly less specific for PJI than SF CCD. The study demonstrates diagnostic value of combining SF viscosity with CCD in decision making in TJA revision surgery

Aim. To assess the clinical characteristics, diagnostic tests and treatment strategies in orthopedic implant-associated infections (OIAI) caused by Cutibacterium spp. Method. We retrospectively included consecutive patients with OIAI caused by Cutibacterium spp. treated at our institution from January 2012 to January 2017. OIAI was diagnosed when: (i) macroscopic purulence, sinus tract or exposed implant was present; (ii) acute inflammation in peri-implant-tissue was documented; (iii) Cutibacterium spp. grew in joint aspirate, ≥2 intraoperative peri-implant tissue samples or in sonication fluid of the removed implant (>50 CFU/ml). Results. Of 67 patients with Cutibacterium OIAI, 42 (63%) had an infected joint prosthesis (21 hip, 12 shoulder, 9 knee) and 25 (37%) an infected fixation device (10 spinal hardware, 11 osteosynthesis, 2 anchorages after rotator cuff reparation, 2 cruciate ligament grafts). 53 (84%) presented with a delayed (3–24 months) or late (>24months) infection. 62 infections were caused by C. acnes and 5 by C. avidum, all being susceptible to levofloxacin and rifampin. Among non-culture-based diagnostic tests, tissue histology had the highest sensitivity (68%), followed by increased synovial fluid leukocyte count/differential (59%). Of culture-based tests, sonication fluid culture showed the highest sensitivity (83%), followed by tissue culture (71%) and synovial fluid culture (61%). Culture positivity rates of synovial fluid, peri-implant tissue and sonication fluid were 20%, 41% and 40%, respectively, after 7 days of incubation and 58%, 74% and 78%, respectively, after 14 days. Most patients were treated with one-stage (24%) or two-stage (55%) implant exchange of the implant. The majority of patients received oral levofloxacin and rifampin for 6–12 weeks. Conclusions. Cutibacterium spp. affected various types of orthopaedic implants in different anatomic locations (lower and upper limbs and spine). Conventional diagnostic tests showed limited sensitivity of Cutibacterium OIAI and can be easily missed when cultures are incubated less than 10–14 days. All Cutibacterium isolates were susceptible to levofloxacin and rifampin

Aims

This study aimed to assess the performance of an automated multiplex polymerase chain reaction (mPCR) technique for rapid diagnosis of native joint septic arthritis