Multicenter Evaluation of a PCR-Based Digital Microfluidics and Electrochemical Detection System for the Rapid Identification of 15 Fungal Pathogens Directly from Positive Blood Cultures.

Routine identification of fungal pathogens from positive blood cultures by culture-based methods can be time-consuming, delaying treatment with appropriate antifungal agents. The GenMark Dx ePlex investigational use only blood culture identification fungal pathogen panel (BCID-FP) rapidly detects 15 fungal targets simultaneously in blood culture samples positive for fungi by Gram staining. We aimed to determine the performance of the BCID-FP in a multicenter clinical study. Blood culture samples collected at 10 United States sites and tested with BCID-FP at 4 sites were compared to the standard-of-care microbiological and biochemical techniques, fluorescence in situ hybridization using peptide nucleic acid probes (PNA-FISH) and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). Discrepant results were analyzed by bi-directional PCR/sequencing of residual blood culture samples. A total of 866 clinical samples, 120 retrospectively and 21 prospectively collected, along with 725 contrived samples were evaluated. Sensitivity and specificity of detection of Candida species (C. albicans, C. auris, C. dubliniensis, C. famata, C. glabrata, C. guilliermondii, C. kefyr, C. krusei, C. lusitaniae, C. parapsilosis, and C. tropicalis) ranged from 97.1 to 100% and 99.8 to 100%, respectively. For the other organism targets, sensitivity and specificity were as follows: 100% each for Cryptococcus neoformans and C. gattii, 98.6% and 100% for Fusarium spp., and 96.2% and 99.9% for Rhodotorula spp., respectively. In 4 of the 141 clinical samples, the BCID-FP panel correctly identified an additional Candida species, undetected by standard-of-care methods. The BCID-FP panel offers a faster turnaround time for identification of fungal pathogens in positive blood cultures that may allow for earlier antifungal interventions and includes C. auris, a highly multidrug-resistant fungus.

Emergence of Ceftolozane-Tazobactam-Resistant Pseudomonas aeruginosa during Treatment Is Mediated by a Single AmpC Structural Mutation.

Ceftolozane-tazobactam is a cephalosporin-ß-lactamase inhibitor combination that exhibits potent in vitro activity against Pseudomonas aeruginosa, including strains that are resistant to other ß-lactams. The emergence of ceftolozane-tazobactam resistance among clinical isolates of P. aeruginosa has rarely been described. Here we characterized ceftolozane-tazobactam-resistant P. aeruginosa strains recovered from a patient who was treated with this agent for 6 weeks for a recurrent wound infection. The results showed that the resistance was mediated by a single AmpC structural mutation.

Unanticipated Multiplex PCR-Based Identification of Polymicrobial Blood Culture Resulting in Earlier Isolation, Determination of Susceptibilities, and Optimization of Clinical Care.

We describe a case of polymicrobial bloodstream infection with six organisms identified by multiplex PCR that was initially thought to be a monomicrobial infection. Early recognition of specific Gram-positive, Gram-negative, and fungal organisms and resistance elements allowed significantly more rapid optimization of therapy.

Endoscope storage time: assessment of microbial colonization up to 21 days after reprocessing.

BACKGROUND: Insufficient data exist for how long endoscopes can be stored after reprocessing. Concern about possible microbial colonization has led to various recommendations for reprocessing intervals among institutions, with many as short as 5 days. A significant cost savings could be realized if it can be demonstrated that endoscopes may be stored for as long as 21 days without risk of clinically significant contamination. OBJECTIVE: To demonstrate whether flexible endoscopes may be stored for as long as 21 days after reprocessing without colonization by pathogenic microbes. DESIGN: Prospective, observational study. SETTING: Tertiary care center. ENDOSCOPES: Four duodenoscopes, 4 colonoscopes, and 2 gastroscopes. INTERVENTION: Microbial testing of endoscope channels. MAIN OUTCOME MEASUREMENTS: Culture results at days 0, 7, 14, and 21. RESULTS: There were 33 positive cultures from 28 of the 96 sites tested (29.2% overall contamination rate). Twenty-nine of 33 isolates were typical skin or environmental contaminants, thus clinically insignificant. Four potential pathogens were cultured, including Enterococcus, Candida parapsilosis, α-hemolytic Streptococcus, and Aureobasidium pullulans; all were likely clinically insignificant as each was only recovered at 1 time point at 1 site, and all grew in low concentrations. There were no definite pathogenic isolates. LIMITATIONS: Single center. CONCLUSION: Endoscopes can be stored for as long as 21 days after standard reprocessing with a low risk of pathogenic microbial colonization. Extension of reprocessing protocols to 21 days could effect significant cost savings.

Successful Eradication of a Highly Resistant Elizabethkingia anophelis Species in a Premature Neonate With Bacteremia and Meningitis.

Elizabethkingia anophelis is a Gram-negative bacillus that can exhibit highly resistant phenotypes against most antibiotics with evidence of efficacy and safety in the neonatal population. Given the limited antimicrobial options, clinicians may be forced into challenging treatment scenarios when faced with central nervous system infections in premature neonates caused by E. anophelis . We report a case of successful treatment of hospital-acquired meningitis and bacteremia caused by E. anophelis at 11 days of life in a male infant born at 29 weeks, 1 day gestation and birth weight of 1.41 kg. Therapy consisted of vancomycin, dose adjusted to maintain goal troughs of 15-20 mg/L, and rifampin 10 mg/kg/dose every 12 hours, with ciprofloxacin 15 mg/kg/dose every 12 hours and trimethoprim/sulfamethoxazole 5 mg/kg/dose every 12 hours added due to antimicrobial susceptibilities and unsatisfactory response, for a total of 21 days. Following initiation of this multidrug regimen, repeat cultures were negative, laboratory parameters improved [with exception of elevated cerebrospinal fluid (CSF) white blood cell count], the patient remained otherwise stable, and there were no adverse effects noted from therapy. Complications after treatment included the requirement of bilateral hearing aids and the development of hydrocephalus necessitating ventriculoperitoneal shunt placement. To our knowledge, we report the first case of meningitis in a premature neonate initially identified as E. anophelis in the United States treated with this regimen which led to successful microbiologic eradication with no antimicrobial safety concerns.

The hidden microbiome of hospital infection surveillance testing: biomarkers of health outcomes in MRSA and VRE colonization.

Background: Hospital-acquired infections present a major concern for healthcare systems in the U.S. and worldwide. Drug-resistant infections result in increased costs and prolonged hospital stays. Methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococcus (VRE) are responsible for many drug-resistant infections in the U.S. We undertook two parallel studies aimed to investigate the differences in the microbial communities of individuals colonized with MRSA (or VRE) as compared to their respective non-colonized counterparts matched for age, sex, race, ethnicity, unit of admission, and diagnostic-related group, when available. Results: The VRE study showed considerably more Enterococcus genus communities in the VRE colonized samples. Our findings for both MRSA and VRE studies suggest a strong association between 16S rRNA gene alpha diversity, beta diversity, and colonization status. When we assessed the colonized microbial communities in isolation, the differences disappeared, suggesting that the colonized microbial communities drove the change. Isolating Staphylococcus, we saw significant differences expressed across colonization in specific sequence variants. Conclusions: The differences seen in the microbial communities from MRSA (or VRE) colonized samples as compared to non-colonized match-pairs are driven by the isolated communities of the Staphylococcus (or Enterococcus) genus, the removal of which results in the disappearance of any differences in the diversity observed across the match-pairs.

Inhaled nitric oxide for adults with pulmonary non-tuberculous mycobacterial infection.

QUESTION: There is an increasing prevalence of nontuberculous mycobacteria pulmonary disease (NTM-PD) in the US. Treatment of NTM-PD typically requires multiple medications, which can be associated with unpleasant morbidity and eradication of infection is difficult. Therefore, there is a critical need for novel effective and well-tolerated therapies. Recent in vitro data and case reports have suggested that nitric oxide, inhaled as a gas (gNO), has antimicrobial activity against NTM. We sought to investigate the effect of gNO in patients with NTM-PD in an open-label proof of concept trial. METHODS: Eligible participants had NTM-PD with persistently positive respiratory cultures for NTM even if on antibiotic treatment. Participants were treated with gNO for 50 min three times daily, five days per week, for three weeks (total of 15 treatment days). RESULTS: Ten participants, of whom nine were on long-term NTM antibiotic therapy, were enrolled. All participants completed the regimen without interruption or discontinuation. Small increases in methemoglobin were noted during treatment, and all resolved to baseline within 2 h. Four participants (40%) met the primary outcome measure of negative sputum cultures after three weeks of therapy. Following treatment discontinuation, three of these participants were again culture positive during the 3-month post-treatment monitoring period, although with measures suggesting low bacterial burden. ANSWER: Patients tolerated a 3-week regimen of gNO without safety concerns, and despite highly refractory disease four individuals completed the study with negative cultures, although three were again positive in subsequent months. These data support further investigation of gNO as a potential therapy for NTM-PD.

Legionnaires' disease caused by Legionella londiniensis.

Legionella londiniensis has been isolated from aqueous environments. However, to our knowledge, this organism has never been isolated from clinical specimens. A case of Legionnaires' disease in a hematopoietic stem cell transplant recipient caused by this organism is described, which confirms that L. londiniensis can be an opportunistic pathogen.

Sustained reduction of microbial burden on common hospital surfaces through introduction of copper.

The contribution of environmental surface contamination with pathogenic organisms to the development of health care-associated infections (HAI) has not been well defined. The microbial burden (MB) associated with commonly touched surfaces in intensive care units (ICUs) was determined by sampling six objects in 16 rooms in ICUs in three hospitals over 43 months. At month 23, copper-alloy surfaces, with inherent antimicrobial properties, were installed onto six monitored objects in 8 of 16 rooms, and the effect that this application had on the intrinsic MB present on the six objects was assessed. Census continued in rooms with and without copper for an additional 21 months. In concert with routine infection control practices, the average MB found for the six objects assessed in the clinical environment during the preintervention phase was 28 times higher (6,985 CFU/100 cm(2); n = 3,977 objects sampled) than levels proposed as benign immediately after terminal cleaning (<250 CFU/100 cm(2)). During the intervention phase, the MB was found to be significantly lower for both the control and copper-surfaced objects. Copper was found to cause a significant (83%) reduction in the average MB found on the objects (465 CFU/100 cm(2); n = 2714 objects) compared to the controls (2,674 CFU/100 cm(2); n = 2,831 objects [P < 0.0001]). The introduction of copper surfaces to objects formerly covered with plastic, wood, stainless steel, and other materials found in the patient care environment significantly reduced the overall MB on a continuous basis, thereby providing a potentially safer environment for hospital patients, health care workers (HCWs), and visitors.

Characterization and control of the microbial community affiliated with copper or aluminum heat exchangers of HVAC systems.

Microbial growth in heating ventilation and air-conditioning (HVAC) systems with the subsequent contamination of indoor air is of increasing concern. Microbes and the subsequent biofilms grow easily within heat exchangers. A comparative study where heat exchangers fabricated from antimicrobial copper were evaluated for their ability to limit microbial growth was conducted using a full-scale HVAC system under conditions of normal flow rates using single-pass outside air. Resident bacterial and fungal populations were quantitatively assessed by removing triplicate sets of coupons from each exchanger commencing the fourth week after their installation for the next 30 weeks. The intrinsic biofilm associated with each coupon was extracted and characterized using selective and differential media. The predominant organisms isolated from aluminum exchangers were species of Methylobacterium of which at least three colony morphologies and 11 distinct PFGE patterns we found; of the few bacteria isolated from the copper exchangers, the majority were species of Bacillus. The concentrations and type of bacteria recovered from the control, aluminum, exchangers were found to be dependent on the type of plating media used and were 11,411-47,257 CFU cm(-2) per coupon surface. The concentration of fungi was found to average 378 CFU cm(-2). Significantly lower concentrations of bacteria, 3 CFU cm(-2), and fungi, 1 CFU cm(-2), were recovered from copper exchangers regardless of the plating media used. Commonly used aluminum heat exchangers developed stable, mixed, bacterial/fungal biofilms in excess of 47,000 organisms per cm(2) within 4 weeks of operation, whereas the antimicrobial properties of metallic copper were able to limit the microbial load affiliated with the copper heat exchangers to levels 99.97 % lower during the same time period.

Identification of the main contributors to blood culture contamination at a tertiary care academic medical center.

Background: Blood culture contamination poses an issue to all hospital systems worldwide because of the associated costs of extended length of stays, unnecessary antibiotic therapy, and additional laboratory testing that are preventable with proper handling and collection techniques. Methods: In our study, multiple units, staff, and collection methods were compared to determine the primary culprits of contamination from a tertiary care academic medical center, which includes a pediatric hospital and both adult and pediatric emergency departments. Results: Over 33 months, 2,083 out of 88,322 total blood cultures collected were contaminated, with an overall contamination rate of 2.4%. A moderate positive correlation was found between the monthly total number of cultures and monthly contamination rate (r = 0.411 P < .01). The most notable factors associated with contamination were found to be phlebotomy teams (2.7%) (P < .01), peripheral draws (2.3%) (P <.01), adult emergency departments (2.6%) (P < .01), and pediatric intensive care units (2.7%) (P < .01). A positive correlation was present between the number of hospital beds per unit and unit contamination rates (r = 0.429 P < .01). Conclusion: Our results were used to make recommendations for decreasing the rate of blood culture contamination in this institution, which includes acknowledgement of an overwhelmed staff and mandatory periodic training on acceptable aseptic technique and contamination awareness. Understanding the factors contributing to blood culture contamination can aid efforts to reduce contamination rates.

Effects of COVID-19 on Blood Culture Contamination at a Tertiary Care Academic Medical Center.

The COVID-19 pandemic has changed health care, from increased needs of personal protective equipment (PPE) to overloaded staff and influxes of patients. Blood cultures are frequently used to detect bloodstream infections in critically ill patients, but it is unknown whether the COVID-19 pandemic has had an impact on blood culture contamination rates. A total of 88,332 blood cultures taken over a 33-month period were analyzed to compare blood culture contamination rates before the COVID-19 pandemic to rates during the pandemic. A significant increase in the average number of monthly nurse-drawn and peripherally collected cultures occurred after the start of the pandemic, but there was a decrease in the average number of phlebotomy cultures. A significant increase in contamination rates (P < 0.001) was found in all nonemergency hospital departments during the COVID-19 pandemic, increasing from 2.1% to 2.5%. Increased rates during the COVID-19 pandemic were also found for nursing staff (2.0% to 2.4%) and both peripheral (2.1% to 2.5%) and indwelling line draws (1.1% to 1.7). The number of cultures drawn monthly increased in acute adult departments and both adult and pediatric emergency departments. Blood culture contamination rates in adult acute, adult emergency, and pediatric intensive care units increased after the start of the pandemic by 23%, 75%, and 59%, respectively. A positive correlation was found between blood culture contamination rates and COVID-19 incidence rates. Additional periodic staff training on proper blood collection technique and awareness of the workload of health care workers are recommended to decrease contamination rates during the COVID-19 pandemic. IMPORTANCE Understanding factors that contribute to blood culture contamination is important in order to take steps to limit contamination events. Here, we examine the effect the COVID-19 pandemic has had on blood culture contamination rates and specifically detail the effects based on the staff, draw types, and unit types. The conclusions provided here can be used as hospitals and laboratories navigate the COVID-19 pandemic or other times of high patient volume.

Increasing Recognition of Community-Acquired, Non-Tuberculous Mycobacterial Infections of the Hand and Wrist.

Background In this study, we present our experience with community-acquired, culture-positive, non-tuberculous mycobacterial (NTM) infections of the hand and wrist and compare the clinical features, risk factors, diagnostic delays, and treatment outcomes among patients referred for surgical consultation at our institution over a five-year period. Methodology We retrospectively identified patients on chart review who were diagnosed with culture-positive, extrapulmonary, cutaneous NTM infections between January 1, 2014, and December 31, 2018. Only patients with community-acquired NTM infections of the hand and wrist were included. Patient demographics, risk factors, location, diagnostic delays, NTM species isolated, treatment modalities, and treatment outcomes were collected and analyzed. These variables were further compared between patients who participated in fishing-related activities and those who did not. Results A total of 10 patients were identified with community-acquired NTM infections of the hand or wrist. Of these patients, eight (80%) were male, and six (60%) had participated in fishing-related activities prior to the initial presentation. The majority of patients had Mycobacterium marinum isolates (n = 6, 60%) and involved the hand (n = 8, 80%). M. marinum isolates were associated with a significantly shorter time to diagnosis (p = 0.02). All patients underwent surgical management with a prolonged course of postoperative antibiotics and were cured of their infection at the end of their treatment course. Conclusions Proper risk factor documentation and heightened clinical awareness are essential to reduce delays in the diagnosis of NTM skin and soft tissue infections and provide the best chance for curative therapy.

Intracavitary voriconazole for the treatment of hemoptysis complicating Pseudallescheria angusta pulmonary mycetomas in fibrocystic sarcoidosis.

Pulmonary mycetomas often occur in fibrocystic sarcoidosis. When this condition is complicated by hemoptysis, definitive surgery is usually precluded because of poor lung function. Intracavitary antifungal therapy has been described for the treatment of symptomatic pulmonary mycetomas. We report the first use of intracavitary voriconazole in the management of a Pseudallescheria angusta pulmonary mycetoma complicated by hemoptysis in a patient with fibrocystic sarcoidosis and renal transplant.

Heterogeneous vancomycin-intermediate susceptibility phenotype in bloodstream methicillin-resistant Staphylococcus aureus isolates from an international cohort of patients with infective endocarditis: prevalence, genotype, and clinical significance.

BACKGROUND: The significance of heterogeneous vancomycin-intermediate Staphylococcus aureus (hVISA) is unknown. Using a multinational collection of isolates from methicillin-resistant S. aureus (MRSA) infective endocarditis (IE), we characterized patients with IE with and without hVISA, and we genotyped the infecting strains. METHODS: MRSA bloodstream isolates from 65 patients with definite IE from 8 countries underwent polymerase chain reaction (PCR) for 31 virulence genes, pulsed-field gel electrophoresis, and multilocus sequence typing. hVISA was defined using population analysis profiling. RESULTS: Nineteen (29.2%) of 65 MRSA IE isolates exhibited the hVISA phenotype by population analysis profiling. Isolates from Oceania and Europe were more likely to exhibit the hVISA phenotype than isolates from the United States (77.8% and 35.0% vs 13.9%; P < .001). The prevalence of hVISA was higher among isolates with a vancomycin minimum inhibitory concentration of 2 mg/L (P = .026). hVISA-infected patients were more likely to have persistent bacteremia (68.4% vs 37.0%; P = .029) and heart failure (47.4% vs 19.6%; P = .033). Mortality did not differ between hVISA- and non-hVISA-infected patients (42.1% vs 34.8%, P = .586). hVISA and non-hVISA isolates were genotypically similar. CONCLUSIONS: In these analyses, the hVISA phenotype occurred in more than one-quarter of MRSA IE isolates, was associated with certain IE complications, and varied in frequency by geographic region.

First reported case of disseminated Microascus gracilis infection in a lung transplant patient.

Microascus gracilis is a specie of the genus Microascus in the family of Microascaceae and has been isolated from lung. It has never been reported as the cause of disseminated infection in humans. Herein, we report a fatal case of disseminated Microascus gracilis infection in a 65-year-old man with a history of primary idiopathic pulmonary fibrosis, status-post bilateral lung transplant. His course was complicated by donor lung cultures positive for multiple organisms and persistent pleural effusions. Multiple lung biopsy and bronchial lavage specimens were negative for mold. Later, pleural fluid cultures grew M. gracilis confirmed by DNA sequencing. Despite aggressive antifungal treatment, the patient continued to deteriorate with altered mental status. Imaging showed scattered hemorrhagic and hypodense lesions in the brain. The patient eventually succumbed to his infections and a restricted autopsy was performed. Autopsy findings included multiple hemorrhagic foci and abscesses involving the whole brain. Numerous punctuate, tan-white circular lesions were on the endocardium and diffuse tan exudates covered the pericardium and lungs. Histologically, similar fungal organisms with septate branching hyphae and short chains of conidia were identified, along with hemorrhage, neutrophilic inflammation, and necrosis in the brain, pleura, peripheral parenchyma of lungs and heart. This is the first reported case of disseminated M. gracilis infection in an immunosuppressed human, indicating it can cause localized infections and disseminated infections. This case increases our awareness of such fatal opportunistic infections, particularly in lung transplant patients, and urges earlier aggressive prophylaxis, diagnosis, and treatment.

Cutibacterium acnes Infection in Orthopedics: Microbiology, Clinical Findings, Diagnostic Strategies, and Management.

Cutibacterium (formerly called Propionibacterium) acnes is a human skin flora often implicated in orthopedic infections. The unique characteristics of this microorganism make the diagnosis of infection difficult. The diagnosis often is made based on clinical evidence, radiographic signs, and laboratory and/or surgical findings combined. Treatment often involves both pharmacologic and surgical methods. In addition, formation of biofilms and increased resistance to drugs exhibited by the microorganism can require combined antimicrobial therapy. Prophylactic measures are particularly important, but no single method has been shown to fully eliminate the risk of C acnes infections. Previous reports have focused on C acnes infections involving surgical implants or after certain orthopedic procedures, particularly in the shoulder and spine. This article reviews current clinical, diagnostic, and treatment principles for C acnes in orthopedics in general. [Orthopedics. 2020; 43(1):52-61.].

Risk Factors for Fungal Prosthetic Joint Infection.

Background: Fungal prosthetic joint infections (PJIs) are rare and often associated with poor outcome; however, risk factors are not well described. Methods: This was a retrospective case control study among all patients with PJIs from 2006-2016 at two major academic centers. Each fungal PJI case was matched 1:1 with a bacterial PJI control by joint (hip, knee, shoulder) and year of diagnosis. We compared demographics, comorbidities, and clinical characteristics between cases and controls using chi square/Fisher's exact or Wilcoxon rank sum test. Independent risk factors were identified with multivariable logistic regression. Results: Forty-one fungal PJIs occurred over the study and 61% were due to Candida albicans. The hip was involved in 51.2% of cases, followed by the knee (46.3%). Compared to bacterial PJI, fungal PJI cases were more likely to have received antibiotics within the previous 3 months (70.7% vs 34%, P=.001), wound drainage lasting >5 days (48% vs 9%, P=.0002), had a lower median CRP (2.95 mg/dl vs 5.99, P=.013) and synovial fluid white blood cell count (13,953 cells/mm3 vs 33,198, P=.007), and a higher proportion of prior two-stage exchanges (82.9% vs 53.6%, P=.008). After controlling for center, prolonged wound drainage (OR, 7.3; 95% CI, 2.02-26.95) and recent antibiotics (OR, 3.4; 95% CI, 1.2-9.3) were significantly associated with fungal PJI. Conclusion: In our study, Candida albicans was the most common species in fungal PJIs and prolonged wound drainage and recent antibiotics were independent risk factors. These clinical characteristics may help providers anticipate fungal PJI and adjust management strategies.