Acquired blaVIM and blaGES Carbapenemase-Encoding Genes in Pseudomonas aeruginosa: A Seven-Year Survey Highlighting an Increasing Epidemiological Threat.

(1) Background: Pseudomonas aeruginosa is a Gram-negative bacterium with several intrinsic and acquired antimicrobial resistance mechanisms. The spread of carbapenemase-encoding genes, an acquired mechanism, enables carbapenem resistance in clinical settings. Detection of the carbapenemase-producer strains is urgent. Therefore, we aimed to characterize carbapenemase production in the clinical strains of P. aeruginosa at a tertiary-care center. (2) Methods: We included clinical strains of P. aeruginosa (from August 2011 to December 2018) with resistance towards at least one carbapenem. Strains were isolated in a tertiary-care center in Mexico City. Antimicrobial susceptibility profiles were determined by broth microdilution. Screening for carbapenemase-encoding genes was performed in all strains. Phenotypic assays (CarbaNP and mCIM) were conducted. Additional modifications to mCIM were also tested. (3) Results: One-hundred seventy-one P. aeruginosa strains out of 192 included in this study were resistant towards at least one of the carbapenems tested. Forty-seven of these strains harbored a carbapenemase-encoding gene. VIM (59.6%) and GES (23.4%) were the most frequently found carbapenemases in our study, followed by IMP (14.9%). (4) Among the most frequent carbapenemase genes identified, metallo-ß-lactamases were the most prevalent, which impair new treatment options. Searching for carbapenemase genes should be performed in resistant isolates to stop transmission and guide antimicrobial treatment.

Next-generation Sequencing Results Require Higher Inoculum for Cutibacterium acnes Detection Than Conventional Anaerobic Culture.

BACKGROUND: Cutibacterium acnes has been described as the most common causative microorganism in prosthetic shoulder infections. Conventional anaerobic culture or molecular-based technologies are usually used for this purpose, but little to no concordance between these methodologies (k = 0.333 or less) has been observed. QUESTIONS/PURPOSES: (1) Is the minimum C. acnes load for detection higher for next-generation sequencing (NGS) than for anaerobic conventional culture? (2) What duration of incubation is necessary for anaerobic culture to detect all C. acnes loads? METHODS: Five C. acnes strains were tested for this study: Four strains were causing infection and were isolated from surgical samples. Meanwhile, the other was a reference strain commonly used as a positive and quality control in microbiology and bioinformatics. To create inoculums with varying degrees of bacterial load, we began with a standard bacterial suspension at 1.5 x 10 8 colony-forming units (CFU)/mL and created six more diluted suspensions (from 1.5 x 10 6 CFU/mL to 1.5 x 10 1 CFU/mL). Briefly, to do so, we transferred 200 µL from the tube with the highest inoculum (for example, 1.5 x 10 6 CFU/mL) to the following dilution tube (1.5 x 10 5 CFU/mL; 1800 µL of diluent + 200 µL of 1.5 x 10 6 CFU/mL). We serially continued the transfers to create all diluted suspensions. Six tubes were prepared per strain. Thirty bacterial suspensions were tested per assay. Then, 100 µL of each diluted suspension was inoculated into brain heart infusion agar with horse blood and taurocholate agar plates. Two plates were used per bacterial suspension in each assay. All plates were incubated at 37°C in an anaerobic chamber and assessed for growth after 3 days of incubation and daily thereafter until positive or Day 14. The remaining volume of each bacterial suspension was sent for NGS analysis to identify bacterial DNA copies. We performed the experimental assays in duplicate. We calculated mean DNA copies and CFUs for each strain, bacterial load, and incubation timepoint assessed. We reported detection by NGS and culture as a qualitative variable based on the identification or absence of DNA copies and CFUs, respectively. In this way, we identified the minimum bacterial load detected by NGS and culture, regardless of incubation time. We performed a qualitative comparison of detection rates between methodologies. Simultaneously, we tracked C. acnes growth on agar plates and determined the minimum incubation time in days required for CFU detection in all strains and loads examined in this study. Growth detection and bacterial CFU counting were performed by three laboratory personnel, with a high intraobserver and interobserver agreement (κ > 0.80). A two-tailed p value below 0.05 was considered statistically significant. RESULTS: Conventional cultures can detect C. acnes at a load of 1.5 x 10 1 CFU/mL, whereas NGS can detect bacteria when the concentration was higher, at 1.5 x 10 2 CFU/mL. This is represented by a lower positive detection proportion (73% [22 of 30]) for NGS than for cultures (100% [30 of 30]); p = 0.004). By 7 days, anaerobic cultures were able to detect all C. acnes loads, even at the lowest concentrations. CONCLUSION: When NGS is negative and culture is positive for C. acnes , there is likely a low bacterial load. Holding cultures beyond 7 days is likely unnecessary. CLINICAL RELEVANCE: This is important for treating physicians to decide whether low bacterial loads necessitate aggressive antibiotic treatment or whether they are more likely contaminants. Cultures that are positive beyond 7 days likely represent contamination or bacterial loads even below the dilution used in this study. Physicians may benefit from studies designed to clarify the clinical importance of the low bacteria loads used in this study at which both methodologies' detection differed. Moreover, researchers might explore whether even lower C. acnes loads have a role in true periprosthetic joint infection.

Plasma D-Dimer Is a Promising Marker to Guide Timing of Reimplantation: A Prospective Cohort Study.

BACKGROUND: Two-stage exchange arthroplasty remains the preferred surgical treatment for chronic periprosthetic joint infection. Currently, there is no single reliable marker to determine the optimal timing for reimplantation. The purpose of this prospective study was to assess the diagnostic utility of plasma D-dimer and other serological markers in predicting successful control of infection following reimplantation. METHODS: This study enrolled 136 patients undergoing reimplantation arthroplasty between November 2016 and December 2020. Strict inclusion criteria were applied including the need for a two-week "antibiotic holiday" prior to reimplantation. A total of 114 patients were included in the final analysis. Plasma D-dimer, erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), and fibrinogen were measured preoperatively. Treatment success was defined using the Musculoskeletal Infection Society Outcome-Reporting Tool. Receiver operating characteristic curves were used to assess the prognostic accuracy of each biomarker in predicting failure following reimplantation at a minimum 1-year follow-up. RESULTS: Treatment failure occurred in 33 patients (28.9%) at a mean follow-up of 3.2 years (range, 1.0 to 5.7). Median plasma D-dimer was significantly higher in the treatment failure group (1,604 versus 631 ng/mL, P < .001), whereas median CRP, ESR, and fibrinogen were not significantly different between the success and failure groups. Plasma D-dimer demonstrated the best diagnostic utility (area under the curve [AUC] 0.724, sensitivity 51.5%, specificity 92.6%), outperforming ESR (AUC 0.565, sensitivity 93.3%, specificity 22.5%), CRP (AUC 0.541, sensitivity 87.5%, specificity 26.3%), and fibrinogen (AUC 0.485, sensitivity 30.4%, specificity 80.0%). Plasma D-dimer level of ≥1,604 ng/mL was identified as the optimal cutoff that predicted failure following reimplantation. CONCLUSION: Plasma D-dimer was superior to serum ESR, CRP, and fibrinogen in predicting failure after the second stage of a two-stage exchange arthroplasty for periprosthetic joint infection. Based on the findings of this prospective study, plasma D-dimer may be a promising marker in assessing the control of infection in patients undergoing reimplantation surgery. LEVEL OF EVIDENCE: Level II.

Mark Coventry Award: Human Knee Has a Distinct Microbiome: Implications for Periprosthetic Joint Infection.

BACKGROUND: Pathogens causing prosthetic joint infection (PJI) are thought to gain access to the knee during surgery or from a remote site in the body. Recent studies have shown that there is a distinct microbiome in various sites of the body. This prospective study, and first of its kind, was set up to investigate the presence of possible microbiome in human knee and compare the profile in different knee conditions. METHODS: We obtained synovial fluid from 65 knees (55 patients) with various conditions that included normal knee, osteoarthritis (OA), aseptic revision, and those undergoing revision for PJI. The contralateral knee of patients who had a PJI were also aspirated for comparison. A minimum of 3 milliliters of synovial fluid was collected per joint. All samples were aliquoted for culture and next-generation sequencing analysis. RESULTS: The highest number of species was found in native osteoarthritic knees (P ≤ .035). Cutibacterium, Staphylococcus, and Paracoccus species were dominant in native nonosteoarthritic knees, and meanwhile a markedly high abundance of Proteobacteria was observed in the osteoarthritic joints. Moreover, the contralateral and aseptic revision knees showed a similar trend in bacterial composition (P = .75). The sequencing analysis of patients who had PJI diagnosis, confirmed the culture results. CONCLUSION: Distinct knee microbiome profiles can be detected in patients who have OA and other knee conditions. The distinct microbiome in the knee joint and the close host-microbe relationships within the knee joint may play a decisive role in the development of OA and PJI.

Sterile Setup Table in the Operating Room Is Not So Sterile.

BACKGROUND: One of the important factors for surgical site infection prevention is the implementation of an ultraclean operating room. This study was designed to evaluate back-table sterility during total joint arthroplasty. METHODS: This prospective study includes 52 patients undergoing primary total joint arthroplasty between November 2021 and January 2022. A total of 4 swabs (2 air swabs and 2 table swabs) were obtained for each case, at the conclusion of surgery and prior to the takedown of drapes. One swab from each set was sent for culture, and the other was sent for next-generation sequencing (NGS) analysis. RESULTS: Among 104 back-table swabs, a total of 13 (12.5%) organisms were isolated. Of these, 7 organisms were isolated by culture and 6 by NGS. No microorganisms were isolated by both culture and NGS from back-table swabs. Among 104 air swabs, a total of 11 (10.6%) organisms were isolated. Of these, 6 microorganisms were isolated by culture and 5 by NGS. In 4 of the 104 swabs, both culture- and NGS-isolated organisms were from air swabs. Of the 104 (12.5%) back-table and air swabs, 13 were culture positive. While more than 1 pathogen was identified in 2 air swabs, all back-table swabs were monomicrobial by culture. Pathogens were identified from 11 of 104 (10.6%) swabs by NGS, while more than 1 pathogen was identified in 4 swabs (2 air and 2 back table). CONCLUSION: The findings of this study raise an important issue that the surgical field including the sterile table setup for instruments is not "sterile" and can harbor pathogens.

Increment Antimicrobial Resistance During the COVID-19 Pandemic: Results from the Invifar Network.

Aim: This study aims to assess the changes in antimicrobial resistance among some critical and high-priority microorganisms collected previously and during the coronavirus disease 2019 (COVID-19) pandemic in Mexico. Methods: We collected antimicrobial susceptibility data for critical and high-priority microorganisms from blood, urine, respiratory samples, and from all specimens, in which the pathogen may be considered a causative agent. Data were stratified and compared for two periods: 2019 versus 2020 and second semester 2019 (prepandemic) versus the second semester 2020 (pandemic). Results: In the analysis of second semester 2019 versus the second semester 2020, in blood samples, increased resistance to oxacillin (15.2% vs. 36.9%), erythromycin (25.7% vs. 42.8%), and clindamycin (24.8% vs. 43.3%) (p ≤ 0.01) was detected for Staphylococcus aureus, to imipenem (13% vs. 23.4%) and meropenem (11.2% vs. 21.4) (p ≤ 0.01), for Klebsiella pneumoniae. In all specimens, increased ampicillin and tetracycline resistance was detected for Enterococcus faecium (p ≤ 0.01). In cefepime, meropenem, levofloxacin, and gentamicin (p ≤ 0.01), resistance was detected for Escherichia coli; and in piperacillin-tazobactam, cefepime, imipenem, meropenem, ciprofloxacin, levofloxacin, and gentamicin (p ≤ 0.01), resistance was detected for Pseudomonas aeruginosa. Conclusion: Antimicrobial resistance increased in Mexico during the COVID-19 pandemic. The increase in oxacillin resistance for S. aureus and carbapenem resistance for K. pneumoniae recovered from blood specimens deserves special attention. In addition, an increase in erythromycin resistance in S. aureus was detected, which may be associated with high azithromycin use. In general, for Acinetobacter baumannii and P. aeruginosa, increasing resistance rates were detected.

Staphylococcus epidermidis small colony variants, clinically significant quiescent threats for patients with prosthetic joint infection.

OBJECTIVES: Staphylococcus epidermidis is a leading cause of prosthetic joint infection. Its relevance is based on its high ability to develop biofilm and small colony variants. However, the clinical associations between this bacterial subpopulation and prosthetic joint infections remain highly uncertain. We aimed to define the clinical characteristics and outcome of patients affected by S. epidermidis small colony variants, as well as their antimicrobial susceptibility. METHODS: We conducted a retrospective cohort study of patients with monomicrobial prosthetic joint infection. Clinical data and time to remission after prosthesis removal was compared between groups. Antimicrobial susceptibility of small colony variants and wild type strains were analyzed. RESULTS: S. epidermidis small colony variants were identified in 16 (37.20%) cultures from eligible subjects. These patients were less likely to achieve remission throughout the follow-up period (Hazard ratio, 0.45 [95% CI, 0.22-0.89], p=0.02), compared to those with wild type strains. In vitro experiments showed higher resistant rates to trimethoprim/sulfamethoxazole (62.5%), and lower resistant rates to levofloxacin (37.5%), among small colony variants. CONCLUSIONS: The isolation of small colony variants was negatively associated with remission achievement throughout management. The search for this bacterial subpopulation and its antimicrobial susceptibility may be appropriate to adjust treatment and clinical expectations.