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Mycobacterium abscessus pulmonary disease (M abscessus-PD) is challenging to treat because of its resistance to antibiotics.What are the outcomes of treatment-naive patients with M abscessus-PD treated with inhaled amikacin-containing multidrug regimens?We identified 82 treatment-naive patients with M abscessus-PD from a prospective observational cohort treated with regimens containing inhaled amikacin with or without clofazimine between March 2015 and June 2018 (ClinicalTrials.gov identifier: NCT00970801). During the initial phase, all patients received IV amikacin, imipenem (or cefoxitin), and oral azithromycin. Oral clofazimine was added in cases of (1) M abscessus subspecies abscessus (here M abscessus) or (2) M abscessus subspecies massiliense (here M massiliense) with cavitary lesions. During the continuation phase, amikacin was changed from an injectional to inhalational form.Of 82 patients, 46 (56%) had M massiliense-PD and 36 (44%) had M abscessus-PD. Among 59 patients with nodular bronchiectatic disease (72%), 23 of 59 had a concurrent cavitary lesion. The remaining 23 patients (28%) had fibrocavitary disease. Twelve months after treatment initiation, cure was achieved in 53 patients (65%): 42 of 46 patients (91%) with M massiliense-PD and 11 of 36 patients (31%) with M abscessus-PD (P .001). Symptomatic and radiologic improvements were observed in 72 patients (88%) and 64 patients (78%), respectively, with significantly greater improvement in patients with M massiliense-PD (symptom improvement, 96% vs 78% [P = .047]; improvement on CT scanning, 93% vs 61% [P = .002]).Inhaled amikacin with or without clofazimine in the regimen provides favorable treatment outcomes in M massiliense-PD. However, more effective treatments are needed for M abscessus-PD.

Objective:To evaluate the impact of a vancomycin-resistant Enterococcus (VRE) screening policy change on the incidence of healthcare-associated (HA)-VRE bacteremia in an endemic hospital setting.Design:A quasi-experimental before-and-after study.Setting:A 1,989-bed tertiary-care referral center in Seoul, Republic of Korea.Methods:Since May 2010, our hospital has diminished VRE screening for admitted patients transferred from other healthcare facilities. We assessed the impact of this policy change on the incidence of HA-VRE bacteremia using segmented autoregression analysis of interrupted time series from January 2006 to December 2014 at the hospital and unit levels. In addition, we compared the molecular characteristics of VRE blood isolates collected before and after the screening policy change using multilocus sequence typing and pulsed-field gel electrophoresis.Results:After the VRE screening policy change, the incidence of hospital-wide HA-VRE bacteremia increased, although no significant changes of level or slope were observed. In addition, a significant slope change in the incidence of HA-VRE bacteremia (change in slope, 0.007; 95% CI, 0.001–0.013; P = .02) was observed in the hemato-oncology department. Molecular analysis revealed that various VRE sequence types appeared after the policy change and that clonally related strains became more predominant (increasing from 26.1% to 59.3%).Conclusions:The incidence of HA-VRE bacteremia increased significantly after VRE screening policy change, and this increase was mainly driven by high-risk patient populations. When planning VRE control programs in hospitals, different approaches that consider risk for severe VRE infection in patients may be required.

Background Cancer patients can be at a higher risk of infection due to drug-resistant bacteria than the general population for various reasons. We performed a retrospective study to evaluate possible risk factors and outcomes of extended-spectrum beta-lactamase-producing Klebsiella pneumoniae (ESBL-KP) bacteremia in cancer patients. Materials and Methods Cases were divided into two groups based on whether or not the isolated strain produced ESBL and multivariable regressions were done to identify possible risk factors of ESBL-KP bacteremia and mortality. For ESBL-producing strain, additional molecular analysis was done. Results 278 cases with KP bacteremia were identified between 2010 and 2012, of which ESBL-producers were 50 (18%). The presence of percutaneous drainage catheter [odds ratio (OR) 4.99, P

An accurate quantitative estimation of the radioactive tracer distribution in positron emission tomography (PET) requires correction of the observed emission data for the attenuation factors. For this purpose, transmission data, provided by X-ray computerized tomography (CT) or magnetic resonance imaging (MRI), are often used to measure the attenuation rate in observed PET data. In this paper, we propose an iterative simultaneous attenuation correction and activity estimation method without transmission data. In each iteration, the proposed method disperses the attenuation effect in the time-of-flight (TOF) PET sinogram in each line-of-response to the current iterates for the activity estimate, then uses that iterate to update the attenuation rate in the TOF PET sinogram, and finally use that updated attenuation rate in computing the next iterates. The simulation result shows that the proposed method corrects attenuation artifact accurately and produces activity estimates comparable to reconstructed ones from attenuation-free data.

The recent increase in severe fever with thrombocytopenia syndrome (SFTS) cases has led to the development of the SFTS-QS kit (MiCoBioMed, Seongnam, Korea) for detecting the SFTS virus (SFTSV, now renamed Huaiyangshan banyangvirus). SFTS-QS is a qualitative real-time reverse transcription PCR assay based on lab-on-a-chip technology. We evaluated the performance of the SFTS-QS kit and compared it with that of the PowerChek SFTSV Real-time PCR kit (PowerChek; Kogene Biotech, Seoul, Korea). A total of 117 serum samples were simultaneously assayed using the SFTS-QS and PowerChek kits. Sanger sequencing targeting the S and M segments of SFTSV was performed as the reference method. The total turnaround time of the two kits was compared. The SFTS-QS results agreed with those of PowerChek with a kappa value of 0.92. The diagnostic sensitivity and specificity of the SFTS-QS kit were both 100% (14/14 and 103/103, respectively), whereas those of the PowerChek kit were 100% (14/14) and 98.1% (101/103), respectively. The results of SFTS-QS and PowerChek were comparable; however, the SFTS-QS kit required a shorter total turnaround time. The SFTS-QS kit produced accurate and fast results and thus could serve as a useful tool for detecting SFTSV.

Objectives The performance of the investigational-use-only version of the BioFire FilmArray Pneumonia Panel (FA-Pneumo), a high-order nested multiplex PCR, was evaluated for the detection of typical respiratory bacterial pathogens and antibiotic resistance genes in sputa and endotracheal aspirate (ETA) specimens. Methods Thirty-one sputa and 69 ETA specimens were analyzed. The diagnostic performance of FA-Pneumo was assessed using routine microbiological methods as the reference standard. Results Overall sensitivity and specificity for organism detection using FA-Pneumo were 98.5% and 76.5%, respectively. The sensitivity for each pathogen was 100%, except for Klebsiella aerogenes, and the range of specificity was 83.3–99.0%. FA-Pneumo detected antimicrobial resistance genes in 17 out of 18 specimens (94.4%) that were resistant by antimicrobial susceptibility testing. FA-Pneumo additionally detected 25 resistance genes in 22 specimens, and sequencing for the presence of resistance genes confirmed the majority of these results (20/25, 80%). Semi-quantitative analysis of bacterial nucleic acid amounts by FA-Pneumo revealed that 88.2% of the identified bacteria (67/76) with ≥106 copies/ml also gave culture-positive results with significant amounts of bacteria. Conclusions FA-Pneumo is a rapid test with high sensitivity for the detection of bacteria and antimicrobial resistance genes in sputum and ETA specimens and could aid in determining antibiotic therapy.

The etiologic diagnostic yield of community-onset pneumonia (COP) using conventional methods is low. Bacterial multiplex polymerase chain reaction (mPCR) has been shown to be more sensitive than conventional methods. This study assessed the clinical factors influencing bacterial mPCR results in patients with COP. Patients with COP admitted to a tertiary care hospital between November 2015 and April 2016 were retrospectively assessed. Conventional methods included culture-based methods and serology for Mycoplasma pneumoniae. Bacterial mPCR that could identify Streptococcus pneumoniae, Haemophilus influenzae, Mycoplasma pneumoniae, and Legionella pneumophilia was performed. Bacterial mPCR was performed in a total of 342 patients with COP in the study. Bacterial mPCR alone provided etiology in 99 patients. The total etiologic diagnosis rates improved from 22.2 to 51.1% when bacterial mPCR was added to conventional methods. Additional diagnostic benefits of bacterial mPCR were more prominent in the prior antibiotic non-exposure group (77.8% vs 63.5%, P = 0.015) and in the low-risk group with low CURB 65 score (62.6% vs 44.9%, P = 0.005). Patients who required ICU care, those with healthcare-associated pneumonia (HCAP), and patients with any underlying diseases were not associated with the additional pathogen detection rates using bacterial mPCR. By supplementing conventional diagnostic methods with bacterial mPCR-based methods, the overall pathogen detection rates improved in patients with COP. Moreover, the additional diagnostic usefulness of bacterial mPCR was significantly higher in patients without prior antibiotic exposure and in the mild-to-moderate-risk group with lower CURB 65 score.

Primary ciliary dyskinesia (PCD) is a genetically heterogeneous disorder characterized by defects in the function or structure of motitle cilia. In most cases, causative variants result in axonemal dynein arm anomalies, however, PCD due to radial spoke (RS) and central pair (CP) of microtubules has been rarely reported. To identify the molecular basis of PCD characterized by RS/CP defects, we performed whole exome sequencing in PCD patients with RS/CP defects. We identified a homozygous nonsense variant (c.572G>A; p.Trp191*) in NME5, which encodes a protein component of the RS neck, in one PCD patient with situs solitus. Morpholino knockdown of nme5 in zebrafish embryos resulted in motile cilia defects with phenotypes compatible with ciliopathy. This is the first study to show NME5 as a PCD-causative gene in humans. Our findings indicate that NME5 screening should be considered for PCD patients with RS/CP defects.

The GENEDIA MTB/NTM Detection Kit (GENEDIA MTB/NTM; Green Cross Medical Science Corp., Chungbuk, Korea) is a multiplex real-time PCR assay used for differential identification of Mycobacterium tuberculosis complex (MTBC) and nontuberculous mycobacteria (NTM). While the importance of differential identification of MTB/NTM is recognized, there is limited data on the performance of GENEDIA MTB/NTM assay to date. A total of 687 consecutive sputum specimens were cultured and analyzed with the GENEDIA MTB/NTM and GENEDIA MTB assays. Nineteen specimens (2.8%) were MTBC-positive, and 69 (10.0%) were NTM-positive based on mycobacterial culture. All specimens showed concordant results for MTBC using both assays, with a kappa value of 1.00, overall sensitivity of 63.2% (12/19), and specificity of 100% (668/668). The overall NTM sensitivity and specificity were 23.2% (16/69) and 99.7% (616/618) for GENEDIA MTB/NTM. The association between NTM-positivity using GENEDIA MTB/NTM and the diagnosis of NTM pulmonary disease was not statistically significant. In conclusion, the two real-time PCR assays showed similar diagnostic performance for MTBC detection. However, the sensitivity for NTM detection was lower than that for MTBC detection.

Background: A major complication of peritoneal dialysis (PD) is peritonitis, and bacterial culture of PD effluent in a blood culture bottle is the preferred technique for diagnosis of peritonitis. In this study, we compared dialysate inoculation and culture using the BacT/AlerTⓇ Fastidious Antimicrobial Neutralization Plus blood culture bottles (FAN Plus; bioMérieux, France) to the conventional centrifugation culture method.Methods: A total of 170 PD effluents were simultaneously processed by the conventional centrifugation culture method and by culture using FAN Plus media with two different inoculation procedures: inoculation after centrifugation and direct bedside inoculation.Results: Of the 52 cultures that were positive on at least one of the culture methods, 27 samples were positive on conventional centrifugation. However, 46 samples showed growth following inoculation into the FAN Plus media after centrifugation, and 47 samples were positive on the direct FAN Plus inoculation method. Using the case definition for PD peritonitis to classify samples, sensitivity of the conventional method was 50.0% (95% CI, 33.7-66.3%), whereas the sensitivity of the FAN Plus media was 78.9% (95% CI, 62.2-89.9%) by inoculation after centrifugation and 86.8% (95% CI, 71.1-95.1%) by direct inoculation. Use of both inoculation methods with FAN Plus media resulted in 92.1% sensitivity (95% CI, 89.2-99.9%).Conclusion: Culture using FAN Plus media demonstrated a superior bacterial recovery rate to the conventional centrifugation culture method. A combination of the two inoculation methods with FAN Plus media is recommended for the best diagnostic yield, while direct inoculation alone can be useful due to its simplicity and cost-effectiveness. (Ann Clin Microbiol 2019;22:90-95)

Shiga toxin-encoding genes (stx) of enterohemorrhagic Escherichia coli (EHEC) can be lost during infection or in vitro cultivation, and in clinical practice, it is difficult to distinguish EHEC that have lost stx (EHEC-LST) from enteropathogenic E. coli (EPEC), as both are stx-negative and eae-positive. In this study, we performed whole-genome sequencing (WGS) of a stx-negative, eae-positive E. coli O63:H6 isolate from a child with hemolytic uremic syndrome and compared its genome with those of nine E. coli O63:H6 strains in public databases. Virulence gene profiles were analyzed and core-genome multilocus sequence typing (cgMLST) was conducted. The virulence gene profile of our isolate was consistent with EHEC, except for the absence of stx, and the isolate clustered with seven EHEC strains but was distant from two EPEC strains in cgMLST. In genome alignment, our isolate exhibited a high nucleotide identity with EHEC strain 377323_2f but displayed a gap corresponding to the stx-harboring prophage sequence. Overall, our isolate was genetically closely related to EHEC strains, consistent with this being an EHEC-LST strain. As EHEC-LST may be misdiagnosed as EPEC in routine laboratories, comparative genomic analysis using WGS can be useful to determine whether stx-negative and eae-positive isolates are EHEC-LST or EPEC.

Limited data are available regarding the in vitro activity of clofazimine against nontuberculous mycobacteria (NTM) or on outcomes of clofazimine-containing regimens in NTM-pulmonary disease (PD). Therefore, we evaluated the in vitro activity of clofazimine and the clinical outcomes of clofazimine-containing regimens. We evaluated clofazimine in vitro activity for 303 NTM isolates from NTM-PD patients. Fifty-seven clarithromycin-resistant and 35 amikacin-resistant isolates were also analyzed. Culture conversion after a 12-month treatment regimen containing clofazimine was evaluated in 58 NTM-PD patients, including 20 patients with drug-resistant isolates. Most of the 303 isolates (238/303) had minimum inhibitory concentrations (MICs) ≤ 0.25 µg/mL for clofazimine (57/63 Mycobacterium avium, 53/57 M. intracellulare, 49/52 M. kansasii, 22/64 M. abscessus, and 57/67 M. massiliense). For the 57 clarithromycin-resistant and 35 amikacin-resistant isolates, most had MICs ≤ 0.25 µg/mL (47/57 and 32/35, respectively). Among the 38 NTM-PD patients without resistance to clarithromycin or amikacin, 47% achieved culture conversion (8/27 M. abscessus, 9/9 M. massiliense, 0/1 M. avium, and 1/1 M. intracellulare). The conversion rate was higher in the MIC ≤ 0.25 µg/mL group than in the MIC = 0.5 µg/mL group (13/18 vs. 5/20, p = 0.004), and an MIC ≤ 0.25 µg/mL remained a significant factor in multivariable analysis. Culture conversion was achieved in 20% of 20 patients with clarithromycin- or amikacin-resistant isolates. However, a clofazimine MIC ≤ 0.25 µg/mL was not significant for culture conversion in the 58 NTM-PD patients, regardless of the drug resistance pattern. Clofazimine was effective in vitro against NTM species. Some patients on clofazimine-containing regimens achieved culture conversion.

Background The difference in clinical outcomes between Klebsiella aerogenes (formerly Enterobacter aerogenes) bacteremia (KAB) and Enterobacter cloacae complex bacteremia (ECB) is controversial. Methods We compared the clinical outcomes of patients with KAB and ECB and examined the risk factors associated with mortality. We conducted a retrospective case-control study of hospitalized patients with monobacterial KAB and ECB between January 2011 and June 2020. The primary outcome measure was 30-day all-cause mortality. Multiple logistic regression and propensity-score (PS) matching were used to identify independent risk factors for mortality. The models included demographic characteristics, comorbidities, recent healthcare contact, patient status at the onset of bacteremia, and severity of infection as covariates. Results A total of 282 patients with KAB or ECB were included, among whom 194 patients were selected after PS matching. The 30-day all-cause mortality rate was higher in the ECB group than in the KAB group (24.1% vs 10.6%, P = .003). In a multivariable model, ECB was an independent risk factor for 30-day mortality in both overall and PS-matched cohorts (adjusted odds ratio, 3.528; 95% confidence interval, 1.614–7.714; P = .002). Stay in the intensive care unit at the onset of bacteremia and higher Pitt bacteremia score were found to be independent risk factors for 30-day mortality. Conclusions In our study, mortality was significantly higher in patients with ECB than in those with KAB. Further studies are warranted to clarify the virulence mechanisms of E cloacae complex.

As various linezolid resistance mechanisms have been identified in methicillin-resistant Staphylococcus aureus (MRSA), we investigated the molecular characteristics of MRSA with elevated linezolid minimum inhibitory concentrations (MICs), using the VITEK 2 system (bioMerieux, Marcy-l'Etoile, France). Twenty-seven MRSA isolates from 14 patients exhibiting linezolid MICs ≥8 μg/mL were examined by broth microdilution (BMD) test as well as by sequencing for mutations in the 23S rRNA gene or ribosomal proteins (L3, L4, and L22) and the presence of the optrA, cfr, and cfr(B) genes. Of the 27 isolates, four (14.8%) from one patient were confirmed as linezolid resistant by BMD and harbored a 23S rRNA T2500A mutation. The remaining 23 were confirmed as linezolid susceptible, indicating that the linezolid-resistant results were major errors generated by VITEK 2. The most commonly detected mutation (19/27, 70.4%), L3 Gly152Asp, was detected in only linezolid-susceptible isolates. No isolates contained optrA, cfr, or cfr(B) or any L4 or L22 protein alterations. Our results show that the 23S rRNA T2500A mutation was mainly associated with linezolid resistance, while the L3 Gly152Asp mutation was not related to linezolid resistance. A confirmatory test is recommended for VITEK 2 linezolid-resistant results owing to the high probability of false resistant results.

Background Information about the natural history of nontuberculous mycobacterial pulmonary disease (NTM-PD) is limited. The purpose of this study was to evaluate the long-term natural history of non-cavitary nodular bronchiectatic NTM-PD and the factors associated with treatment initiation and the frequency of spontaneous sputum culture conversion after diagnosis of NTM-PD. Methods We evaluated 1,021 patients with newly diagnosed non-cavitary nodular bronchiectatic NTM-PD caused by Mycobacterium avium complex or M. abscessus between 2003 and 2013. Results Of 1,021 patients, 562 (55%) initiated antibiotic treatment and 459 (45%) did not. Young age (adjusted hazard ratio [aHR] = 0.99; 95% confidence interval [CI] = 0.98–0.99), low body mass index (aHR = 0.96; 95% CI = 0.93–0.99), previous history of tuberculosis (aHR = 1.23; 95% CI = 1.01–1.50), respiratory complaints such as cough (aHR = 1.36; 95% CI = 1.05–1.75) and sputum production (aHR = 1.47; 95% CI = 1.13–1.91), and high number of involved lobes on high-resolution computed tomography (aHR = 1.22; 95% CI = 1.14–1.31) were associated with treatment initiation. Of 459 patients who did not initiate treatment, 157 (34%) showed spontaneous sputum culture conversion. None of the clinical factors was associated with spontaneous conversion. After spontaneous culture conversion, 26 of 157 (17%) showed redeveloped NTM-PD caused by a species different from the original species. Conclusions The natural history of non-cavitary nodular bronchiectatic NTM-PD is variable. After diagnosis, the decision to initiate antibiotic therapy should be individualized based on consideration of the risk factors for disease progression. However, for patients who do not start antibiotic therapy, continuous and lifetime follow-up is recommended to manage underlying bronchiectasis and the possibility of late progression of NTM-PD.

BACKGROUND Real-time PCR is more sensitive than microscopic examination for detecting Pneumocystis jirovecii. We compared the performance of two assays for detecting P. jirovecii DNA: the RealStar Pneumocystis jirovecii PCR Kit 1.0 CE (Altona Diagnostics, Hamburg, Germany) and the AmpliSens Pneumocystis jirovecii (carinii)-FRT PCR kit (InterLabService Ltd., Moscow, Russia). METHODS We used 159 samples from the lower respiratory tract (112 bronchoalveolar lavage [BAL] fluid, 37 sputum, and 10 endotracheal aspirate [ETA] samples) of non-HIV immunocompromised patients. Nested PCR and sequencing were used to resolve discordant results. The performance of the two assays was evaluated according to clinical categories (clinical Pneumocystis pneumonia [PCP], possible PCP, or unlikely PCP) based on clinical and radiological observations. RESULTS The positive and negative percent agreement values were 100% (95% confidence interval [CI], 85.4-100%) and 96.6% (95% CI, 90.9-98.9%), respectively, and kappa was 0.92 (95% CI, 0.84-0.99). P. jirovecii DNA load was significantly higher in the clinical PCP group than in the other groups (P

Background Mycobacterium kansasii is a major pathogen associated with nontuberculous mycobacterial pulmonary disease. For treatment of M. kansasii pulmonary disease, daily therapy with isoniazid, rifampin, and ethambutol is traditionally recommended. Although a regimen containing a macrolide, instead of isoniazid, has been recently recommended, supporting data are limited. We compared the treatment outcomes of a macrolide-containing regimen (macrolide group) and an isoniazid-containing regimen (isoniazid group) on patients with M. kansasii pulmonary disease. Methods A total of 49 patients were identified between January 2002 and December 2016. Treatment outcomes for the isoniazid group (n = 24) and the macrolide group (n = 25) were compared. Results Baseline characteristics of the isoniazid and macrolide groups were similar. Favorable outcomes did not differ between the isoniazid group (79%, n = 19) and macrolide group (88%, n = 22, P = 0.463). Total treatment duration (median 17.9 months vs. 15.4 months; P = 0.712) and time to culture conversion (median 2.0 months vs. 1.2 months; P = 0.838) were also similar between the isoniazid and macrolide groups. Five patients who completed three-times-weekly intermittent treatment containing a macrolide for non-cavitary M. kansasii pulmonary disease achieved negative sputum culture conversion within 12 months of treatment. Only one patient experienced recurrence of M. kansasii pulmonary disease in the isoniazid group. Conclusions A macrolide-containing regimen appears to be as effective as an isoniazid-containing regimen for treatment of M. kansasii pulmonary disease. Additionally, intermittent therapy containing a macrolide could be an alternative treatment option for non-cavitary M. kansasii pulmonary disease.

In this study, we aimed to retrospectively investigate and compare the drug susceptibility patterns of two major Mycobacterium abscessus complex (MABC) species; M. abscessus and M. massiliense. A total of 546 MABC respiratory isolates (277 M. abscessus and 269 M. massiliense) from 2011 to 2016 were analyzed in this study. We estimated minimum inhibitory concentrations (MICs) using the broth microdilution method, and we calculated MIC50 and MIC90 values from the MIC distribution. Both M. abscessus and M. massiliense were highly susceptible to amikacin and linezolid. For M. abscessus, the proportions of inducible and acquired resistance to clarithromycin were 68.6% and 12.3%, respectively. Only 15.2% of M. abscessus remained susceptible at day 14. On the other hand, none of the M. massiliense showed inducible resistance and 6.3% showed acquired resistance to clarithromycin. A total of 92.6% of the M. massiliense remained susceptible at day 14. The resistance rate of M. abscessus to moxifloxacin (90.3%) was significantly higher than that of M. massiliense (83.3%; p = 0.016). These susceptibility differences may explain the divergent treatment outcomes between patients with pulmonary disease caused by these two species.

Background The AdvanSure MDR-TB GenoBlot Assay detects isoniazid- and rifampin-resistant tuberculosis using mutation-specific probes, including probes to disputed rpoB mutations. The aim of this study was to evaluate the clinical usefulness of molecular drug susceptibility testing (DST) using the AdvanSure assay with weekly batch testing in routine clinical laboratory settings in a country with an intermediate tuberculosis burden. Methods The AdvanSure assay was evaluated against an absolute concentration (AC) method and the Mycobacterial Growth Indicator Tube (MGIT) 960 System, which are phenotypic DST methods, using 496 Mycobacterium tuberculosis (MTB) isolates. We retrospectively reviewed and compared DST results and turnaround times (TATs), the time intervals from MTB culture identification to final reporting, for these methods. Results For rifampin, the AdvanSure assay showed 99.2% (492/496) concordance with both the AC and MGIT methods. AdvanSure also detected an rpoB mutation (D516Y) conferring low-level resistance in three isolates categorized as rifampin-susceptible by both phenotypic DST methods. For isoniazid, AdvanSure concordance rates with the AC method and MGIT DST were 96.6% (479/496) and 95.4% (473/496), respectively. The median TAT of AdvanSure in weekly batch testing was 5.8 days, shorter than the times for the phenotypic DST methods, which were 35.1 days for the AC method and 8.9 days for MGIT DST. Conclusions AdvanSure shows promising clinical usefulness for rapid detection of rifampin- and/or isoniazid-resistant tuberculosis when used as a complementary method to phenotypic DST assays in weekly batch testing. Furthermore, MTB isolates with disputed mutations for rifampin resistance were detectable by the AdvanSure assay.

We evaluated the in vitro activity of rifamycin derivatives, including rifampin, rifapentine, rifaximin, and rifabutin, against clinical nontuberculous mycobacteria (NTM) isolates. Of the rifamycin derivatives, rifabutin showed the lowest MICs against all NTM species, including Mycobacterium avium complex, M. abscessus , and M. kansasii .

We evaluated the association between 16S rRNA gene (rrs) mutations and susceptibility in clinical isolates of amikacin-resistant nontuberculous mycobacteria (NTM) in NTM-pulmonary disease (PD) patients. Susceptibility was retested for 134 amikacin-resistant isolates (minimum inhibitory concentration [MIC] ≥ 64 µg/ml) from 86 patients. Amikacin resistance was reconfirmed in 102 NTM isolates from 62 patients with either Mycobacterium avium complex-PD (MAC-PD) (n = 54) or M. abscessus-PD (n = 8). MICs and rrs mutations were evaluated for 318 single colonies from these isolates. For the 54 MAC-PD patients, rrs mutations were present in 34 isolates (63%), comprising all 31 isolates with amikacin MICs ≥ 128 µg/ml, but only three of 23 isolates with an MIC = 64 µg/ml. For the eight M. abscessus-PD patients, all amikacin-resistant (MIC ≥ 64 µg/ml) isolates had rrs mutations. In amikacin-resistant isolates, the A1408G mutation (n = 29) was most common. Two novel mutations, C1496T and T1498A, were also identified. The culture conversion rate did not differ by amikacin MIC. Overall, all high-level and 13% (3/23) of low-level amikacin-resistant MAC isolates had rrs mutations whereas mutations were present in all amikacin-resistant M. abscessus isolates. These findings are valuable for managing MAC- and M. abscessus-PD and suggest the importance of phenotypic and genotypic susceptibility testing.

Limited data are available regarding optimal treatment for refractory Mycobacterium avium complex-pulmonary disease (MAC-PD). We evaluated outcomes of inhaled amikacin (AMK) with clofazimine (CFZ) regimens as an add-on salvage therapy for refractory MAC-PD. We retrospectively analyzed 52 patients with refractory MAC-PD, characterized by persistently positive sputum cultures despite &gt, 6 months of treatment. Thirty-five (67%) patients had M. intracellulare-PD, and 17 (33%) patients had M. avium-PD. Twenty-seven (52%) patients received the salvage therapy for &ge, 12 months, whereas 25 (48%) patients were treated for &lt, 12 months due to adverse effects or other reasons. Seventeen (33%) patients had culture conversion: 10 (10/27) in the &ge, 12-month treatment group and seven (7/25) in the &lt, 12-month treatment group (p = 0.488). Microbiological cure, defined as maintenance of culture negativity, was achieved in 12 (23%) patients, six (6/12) with accompanying symptomatic improvement were considered to have reached cure. Clinical cure, defined as symptomatic improvement with &lt, 3 consecutive negative cultures, was achieved in three (6%) patients. Overall, 15 (29%) patients achieved favorable outcomes, including microbiological cure, cure, and clinical cure. Inhaled AMK with CFZ may provide favorable outcomes in some patients with refractory MAC-PD. However, given the adverse effects, more effective strategies are needed to maintain these therapeutic regimens.

Background The Automated Fluorescent Immunoassay System ROTA (AFIAS‐Rota) and NORO (AFIAS‐Noro) assays (Boditech Med Inc.) are newly developed diagnostic tests for rotavirus and norovirus infections. Methods Performance of AFIAS‐Rota/Noro assays was evaluated in comparison with RIDASCREEN® Rotavirus and Norovirus ELISA kits (R‐Biopharm) using clinical stool samples submitted from November 2018 to January 2019. Multiplex real‐time reverse transcription‐polymerase chain reaction was used as reference method. Results A total of 256 clinical specimens were analyzed. AFIAS‐Rota and RIDASCREEN Rotavirus had almost perfect agreement (Kappa value = 0.95), and substantial agreement was observed between AFIAS‐Noro and RIDASCREEN Norovirus (Kappa value = 0.80). For detection of rotavirus, AFIAS and RIDASCREEN assays showed satisfactory diagnostic sensitivity (100% and 97.8%, respectively) and specificity (99.5% and 99.1%). For detection of norovirus, the RIDASCREEN assay showed significantly higher sensitivity than the AFIAS‐Noro (86.0% and 66.0%, respectively; P = .002). Analytic specificity of AFIAS‐Rota/Noro assays showed no cross‐reactivity against any other bacteria (14 strains) or viruses (2 strains). Hands‐on time (6 minutes) and turnaround time (26 minutes) required to perform AFIAS assays were much shorter than those required for RIDASCREEN assays (20 and 150 minutes, respectively). Conclusion The AFIAS‐Rota/Noro assays showed overall excellent agreement with the RIDASCREEN assays. Although the AFIAS‐Noro assay exhibited lower sensitivity than the RIDASCREEN Norovirus assay for detection of norovirus, the AFIAS‐Rota/Noro assays could be useful as a rapid initial screening test in clinical laboratories due to its convenience and rapid turnaround time.

Objectives We evaluated the performance of the MicroIDSys Elite system, a newly developed matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry system for identification of mycobacteria directly from positive MGIT liquid cultures. Methods Analytical specificity was evaluated with 63 reference strains grown in mycobacteria growth indicator tube media. Prospective performance evaluation was conducted with primary liquid cultures of sputum samples for identification of mycobacteria, and results were compared to multigenerational sequencing as the reference method. Liquid media subcultures were also analyzed. Results The accuracy for the 63 reference strains was 98.4% (62/63). A total of 167 paired mycobacterial primary cultures and subcultures in liquid media, comprised of seven Mycobacterium tuberculosis isolates, 109 slowly growing nontuberculous mycobacterial isolates, and 51 rapidly growing nontuberculous mycobacterial isolates, was identified by the MicroIDSys Elite system. Using primary liquid cultures, the MicroIDSys Elite system correctly identified 143 (85.6%) isolates; 21 (12.6%) resulted in “no identification”; and three (1.8%) isolates were misidentified. Using liquid media subcultures with this system, 159 (95.2%) isolates were correctly identified; seven (4.2%) resulted in “no identification”; and one (0.6%) isolate was misidentified. Conclusion The MicroIDSys Elite system is a useful routine diagnostic tool for identification of mycobacterial species from liquid culture.

With the increasing number of fungal infections and immunocompromised patients, rapid and accurate fungal identification is required in clinical microbiology laboratories. We evaluated the applicability of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) system, MicroIDSys Elite (ASTA Corp., South Korea) for the identification of medically important filamentous fungi. A total of 505 strains comprising 37 genera and 90 species collected from 11 Korean hospitals were sent to the microbiology laboratory of International St. Mary’s Hospital. All isolates were tested using MicroIDSys Elite, and data were analyzed using the MoldDB v.1.22 database (ASTA). Correct identification rates were compared with the multigene sequencing results. MicroIDSys Elite correctly identified 86.5% (437/505) and 88.9% (449/505) of all tested isolates at the species and genus level, respectively. About 98.2% of Aspergillus isolates were identified at the species level, including cryptic and rare species of A. calidoustus, A. tamarii, A. lentulus, A. versicolor and A. aculeatus. MicroIDSys Elite identified 75.0% of basidiomycetes, including Schizophyllum commune, and 84.3% of the dermatophytes. It also distinguished Sprothrix globosa at the species level. The mean scores of total isolates corresponding to correct species identification were significantly higher than those obtained for genus-level identification (253.5 ± 50.7 vs. 168.6 ± 30.3, P

Background We investigated the treatment outcomes before and after the addition of amikacin to cefepime monotherapy as an initial empirical antibiotic treatment in pediatric cancer patients with febrile neutropenia. Methods This was a retrospective historical cohort study. The subjects were pediatric cancer patients who visited the emergency room at the Samsung Medical Center, Seoul, Korea, due to chemotherapy-induced febrile neutropenia, between January 2011 and December 2016. Since September 2014, the empirical antimicrobial treatment regimen for febrile neutropenia was changed from high-dose cefepime monotherapy to combination therapy of adding a single dose of amikacin. Results Two hundred twenty-five bacteremia episodes in 164 patients were reported during the study period. Bacteremia caused by cefepime-resistant Gram-negative bacteria was observed in 16% of patients before September 2014 and in 21% of the patients after September 2014 (P = 0.331). Use of appropriate empirical antibiotic treatments increased from 62% to 83% following addition of amikacin to cefepime treatment (P = 0.003). The duration of fever was shorter in the cefepime plus amikacin group than in the cefepime group (22 vs. 34 hours, P = 0.014); however, rates of septic shock and pediatric intensive care unit hospitalizations were not significantly different between the 2 groups (septic shock, both 7%, P = 0.436; pediatric intensive care unit 3% vs. 1%, P = 0.647). Conclusions We observed no additional benefit of amikacin addition to high-dose cefepime monotherapy. Therefore, adding amikacin to cefepime monotherapy in conditions where cefepime-resistant Gram-negative bacteremia amounts to 20% or less may not be justified.

Patients with refractory Mycobacterium avium complex lung disease (MAC-LD) undergo long-term macrolide therapy, but macrolide resistance develops infrequently.The aim of this study was to determine whether reinfection was a factor in the low incidence of macrolide resistance in patients with refractory MAC-LD.Among 481 patients with treatment-naive MAC-LD who started antibiotic treatment between January 2002 and December 2013, we identified 72 patients with refractory disease, characterized by persistently positive sputum cultures despite ≥12 months of treatment. Molecular analyses of the 23S ribosomal RNA gene responsible for macrolide resistance and serial mycobacterial genotyping were performed using stored MAC isolates.The median duration of treatment was 32 months (interquartile range, 24-41 mo) in 72 patients. After treatment for a median of 33 months (interquartile range, 21-44 mo), macrolide resistance developed in 16 (22%) patients. Molecular analysis of isolates from 15 patients revealed that 80% (12 of 15) had a point mutation at position 2,058 or 2,059 of the 23S ribosomal RNA gene. Of the 49 patients who had stored pre- and post-treatment isolates, mycobacterial genotyping revealed that reinfection by new MAC strains occurred in 36 (73%) patients. New MAC strains were found in 24 (49%) patients, and mixed infections with original and new strains occurred in 12 (24%) patients. Only 13 (27%) patients had persistent infections with their original MAC strains.Refractory MAC-LD is commonly caused by reinfection with new strains rather than persistence of the original strain, which may explain the infrequent development of macrolide resistance in refractory MAC-LD. Clinical trial registered with www.clinicaltrials.gov (NCT00970801).

To evaluate the impacts of fluconazole minimum inhibitory concentration (MIC) according to primary antifungal agents on Candida glabrata bloodstream infection (BSI), a multicenter retrospective cohort study was conducted in Korea, concerning the time period from January 2010 to February 2016. A total of 197 adult patients with C. glabrata BSI were included in the study, and neutropenia (P = 0.026), APACHE II score (P = 0.004), and fluconazole resistance (HR 3.960, 95% CI 1.395-11.246, P = 0.010) were associated with 30-day mortality in multivariate analysis. In subgroup analysis, fluconazole MIC = 32 μg/mL in the azole-treated group (HR 6.691, 95% CI 1.569-28.542, P = 0.010) and fluconazole MIC ≥ 64 μg/mL in the non-azole-treated group (HR 3.337, 95% CI 1.183-9.411, P = 0.023) showed the highest hazard ratio (HR) for 30-day mortality. Increased fluconazole MIC was associated with poor outcome both in azole- and non-azole-treated patients with C. glabrata BSI.

We evaluated the efficacy of intermittent azithromycin and ethambutol therapy for noncavitary Mycobacterium avium complex pulmonary disease (MAC-PD). Twenty-nine (76%) of 38 patients achieved sputum culture conversion after 12 months of treatment, and sputum smear positivity was an independent factor for failure to achieve culture conversion (adjusted odds ratio, 26.7; 95% confidence interval, 2.1 to 339.9; P = 0.011).

The use of fluoroquinolones to treat respiratory tract infections and pneumonia due to Streptococcus pneumoniae has affected the emergence of resistance to this class of drugs. Increasing pneumococcal resistance to levofloxacin has become a major public health concern. We investigated the prevalence and genetic characteristics of levofloxacin-nonsusceptible S. pneumoniae (LNSP) clinical isolates in Korea. A total of 43 LNSP isolates collected from a national surveillance study at 13 tertiary hospitals between 2008 and 2014 were analyzed for serotype and antimicrobial susceptibilities to 19 antimicrobial agents as well as the quinolone resistance-determining region mutation. Multilocus sequence typing was performed to investigate the genetic relatedness among LNSP isolates. All LNSP isolates (MIC, ≥4 μg/mL) exhibited multidrug-resistant or even extensively drug-resistant (XDR) phenotypes (8 isolates, 18.6%). Most LNSP isolates belonged to sequence type (ST) 8279 and its variants (16 isolates, 37.2%). ST8279 is a double-locus variant of ST156, which is identical to the pneumococcal Spain9V-3 international clone. The high prevalence of nonvaccine types in LNSP isolates could pose significant therapeutic challenges. A limited number of clones dominated the population of LNSP XDR isolates, and homogeneous antimicrobial resistance profiles support the possibility of clonal dissemination of LNSP. More information on the emergence and spread of these LNSP isolates is necessary in order to prevent its spread.

Objectives This study aimed to determine the prevalence of linezolid-resistant (LR) vancomycin-resistant enterococci and to investigate the mechanisms of linezolid resistance with clinical and microbiological characterisation. Methods All vancomycin-resistant Enterococcus faecium (VREF) isolated from blood and rectal swab cultures during 2012–2015 were tested for linezolid resistance. LR-VREF isolates were tested for antimicrobial susceptibility, glycopeptide resistance genes and virulence genes. Multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE) were performed. Isolates were tested for known mechanisms of linezolid resistance. Results Among 389 VREF isolates, 7 (1.8%) were found to be resistant to linezolid. All LR-VREF isolates carried the vanA gene. Five isolates had both hyl and esp genes. The isolates were susceptible to tigecycline, daptomycin and quinupristin/dalfopristin, except for one isolate with daptomycin resistance. Two LR-VREF isolates recovered from patients with previous linezolid exposure contained the G2576T mutation in 23S rRNA and exhibited high-level resistance to linezolid (MIC > 64 mg/L). The other five isolates recovered from linezolid-naive patients revealed no known linezolid resistance mechanism and exhibited low-level resistance to linezolid (MICs = 8–16 mg/L). Plasmid-mediated genes encoding cfr or optrA were not detected. LR-VREF isolates were represented by six different sequence types, belonging to hospital lineages, and were assigned to seven PFGE types. Conclusions The prevalence of LR-VREF in this centre was low. Both linezolid exposure and horizontal transmission appear to be responsible for acquisition of LR-VREF in hospitalised patients. Prudent use of linezolid and improved infection control strategies are needed to limit the spread of LR-VREF.

Objectives To evaluate the performance of a rapid antimicrobial susceptibility testing (AST) platform based on microfluidic chip technology, the QMAC-dRAST, which enables AST from colony isolates or positive blood culture broth (PBCB), and to compare the performance of the QMAC-dRAST for staphylococci and enterococci with that of the VITEK-2 system based on reference broth microdilution (BMD). Methods A total of 110 staphylococcal and enterococcal isolates from blood cultures were included. AST was performed directly using the QMAC-dRAST with PBCB. Thereafter, colony isolates derived from subculture of PBCB were used for the QMAC-dRAST, the VITEK-2 system and BMD. Results The overall agreement between the QMAC-dRAST with PBCB and BMD was 91.5%. There were 1.2% very major errors (VMEs), 4.3% major errors (MEs) and 5.4% minor errors (mEs). The QMAC-dRAST with colony isolates yielded 94.6% agreement and error rates of 1.0% VMEs, 1.8% MEs and 4.0% mEs. The VITEK-2 system showed 96.2% agreement and error rates of 2.3% VMEs, 0.5% MEs and 2.6% mEs. The incubation time in the QMAC-dRAST was significantly shorter than in the VITEK-2 system (median of 6 versus 10 h; P

Objectives: The aim of this study was to evaluate whether rifabutin can improve treatment outcomes in patients with rifabutin-sensitive MDR-TB. Methods: A retrospective cohort study was performed on 76 patients with rifabutin-sensitive MDR-TB who were treated with or without rifabutin between 2006 and 2011. Results: Overall, 75% (57/76) of patients achieved favorable outcomes, including cure (53/76, 70%) and treatment completion (4/76, 5%). In contrast, 25% (19/76) had unfavorable treatment outcomes, which included treatment failure (6/76, 8%), death (2/76, 3%), loss to follow-up (4/76. 5%), and no evaluation due to transfer to other institutions (7/76, 9%). Rifabutin was given to 52 (68%) of the 76 patients with rifabutin-sensitive MDR-TB. Although favorable treatment outcomes were more frequent in patients who received rifabutin [81% (42/52)] than in those who did not receive rifabutin [63% (15/24)], this difference was not statistically significant (P = 0.154). However, in multivariable regression logistic analysis, use of rifabutin was significantly associated with favorable treatment outcomes in patients with rifabutin-sensitive MDR-TB (adjusted odds ratio = 9.80, 95% confidence interval = 1.65â58.37, P = 0.012). Conclusions: These results suggest that the use of rifabutin can improve treatment outcomes in patients with rifabutin-sensitive MDR-TB. Keywords: Multidrug-resistant tuberculosis, Extensively drug-resistant tuberculosis, Rifabutin, Treatment outcome

For an accurate quantitative reconstruction of the radioactive tracer distribution in positron emission tomography (PET), we need to take into account the attenuation of the photons by the tissues. For this purpose, we propose an attenuation correction method for the case when a direct measurement of the attenuation distribution in the tissues is not available. The proposed method can determine the attenuation factor up to a constant multiple by exploiting the consistency condition that the exact deconvolution of noise-free time-of-flight (TOF) sinogram must satisfy. Simulation studies shows that the proposed method corrects attenuation artifacts quite accurately for TOF sinograms of a wide range of temporal resolutions and noise levels, and improves the image reconstruction for TOF sinograms of higher temporal resolutions by providing more accurate attenuation correction.

We describe the laboratory identification of Leptotrichia species from clinical isolates collected over a six-year period. Five isolates from blood cultures were identified as Leptotrichia species. Gram stain showed large, fusiform, gram-negative or -variable bacilli. Identification based on biochemical testing was unsuccessful; however, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry proved to be a useful tool for identifying Leptotrichia species to the genus level. Species level identification was successfully achieved by using 16S ribosomal RNA gene sequencing.

Background/aims Although multidrug resistance (MDR) among extended-spectrum β-lactamase-producing Escherichia coli (ESBL-EC) poses significant therapeutic challenges, little is known regarding the risk factors and epidemiology of community-onset MDR-ESBL-EC infections. We performed this study to investigate risk factors and the molecular epidemiology of community-onset MDR-ESBL-EC infections. Methods We conducted a case-control-control study of community-onset infections. MDR-ESBL-EC was defined as ESBL-EC that demonstrated in vitro resistance to trimethoprim-sulfamethoxazole, fluoroquinolones (FQs), and gentamicin. Patients with MDR-ESBL-EC infections were designated as case patients. A control group I (CG I) patient was defined as a person whose clinical sample yielded ESBL-EC that did not meet the criteria for MDR. A control group II (CG II) patient was defined as a patient with a non-ESBL-EC infection. Results Of 108 patients with ESBL-EC infections, 30 cases (27.8%) were due to MDR-ESBL-EC. Compared with CG I, prior use of FQs (odds ratio [OR], 3.16; 95% confidence interval [CI], 1.11 to 8.98) and immunosuppressant use (OR, 10.47; 95% CI, 1.07 to 102.57) were significantly associated with MDR-ESBL-EC. Compared with CG II, prior use of FQs (OR, 15.53; 95% CI, 2.86 to 84.27) and healthcare-associated infection (OR, 5.98; 95% CI, 2.26 to 15.86) were significantly associated with MDR-ESBL-EC. CTX-M-15 was the most common in MDR-ESBL-EC infections (59.1% [13/22]), while CTX-M-14 was the most common in non-MDR-ESBL-EC infections (41.6% [32/77]). CTX-M-15 was significantly associated with MDR-ESBL-EC (59.1% vs. 32.5%, p = 0.028). Pulsed-field gel electrophoresis showed clonal diversity of MDR-ESBL-EC isolates. Conclusions The emergence of strains of MDR-ESBL-EC in the community poses an important new public health threat. More information on the emergence and transmission of these strains will be necessary in order to prevent their spread.

Although Mycobacterium massiliense lung disease is increasing in patients with cystic fibrosis and non-cystic fibrosis bronchiectasis, optimal treatment regimens remain largely unknown. This study aimed to evaluate the efficacy of oral macrolide therapy after an initial 2-week course of combination antibiotics for the treatment of M massiliense lung disease.Seventy-one patients received oral macrolides, along with an initial 4-week (n = 28) or 2-week (n = 43) IV amikacin and cefoxitin (or imipenem) treatment. These patients were treated for 24 months (4-week IV group) or for at least 12 months after negative sputum culture conversion (2-week IV group).Total treatment duration was longer in the 4-week IV group (median, 23.9 months) than in the 2-week IV group (15.2 months; P .001). The response rates after 12 months of treatment were 89% for symptoms, 79% for CT scanning, and 100% for negative sputum culture results in the 4-week IV group. In the 2-week IV group, these values were 100% (P = .057), 91% (P = .177), and 91% (P = .147), respectively. Acquired macrolide resistance developed in two patients in the 2-week IV group. Genotyping analyses of isolates from patients who did not achieve negative sputum culture conversion during treatment and from those with positive culture results after successful treatment completion revealed that most episodes were due to reinfection with different genotypes of M massiliense.Oral macrolide therapy after an initial 2-week course of combination antibiotics might be effective in most patients with M massiliense lung disease.ClinicalTrials.gov; No.: NCT00970801; URL: www.clinicaltrials.gov.

Background Long-term nucleoside analogue (NA) treatment leads to selection for drug-resistant mutations in patients undergoing hepatitis B virus (HBV) therapy. The Abbott RealTime HBV Sequencing assay (Abbott assay; Abbott Molecular Inc., Des Plaines, IL, USA) targets the reverse transcriptase region of the polymerase gene and as such has the ability to detect NA resistance-associated mutations in HBV. Objectives We evaluated the analytical performance of the Abbott assay and compared its diagnostic performance to that of a laboratory-developed nested-PCR and sequencing method. Study design The analytical sensitivity of the Abbott assay was determined using a serially-diluted WHO International Standard. To validate the clinical performances of the Abbott assay and the laboratory-developed assay, 89 clinical plasma samples with various levels of HBV DNA were tested using both assays. Results The limit of detection of the Abbott assay, was 210 IU/ml and it successfully detected mutations when the mutant types were present at levels ≥20%. Among 89 clinical specimens, 43 and 42 were amplification positive in the Abbott and laboratory-developed assays, respectively, with 87.6% overall agreement (78/89; 95% confidence interval [CI], 78.6–93.4). The Abbott assay failed to detect the minor mutant populations in two specimens, and therefore overall concordance was 85.3% (76/89), and the kappa value was 0.79 (95% CI, 0.67–0.90). Conclusions The Abbott assay showed comparable diagnostic performance to laboratory-developed nested PCR followed by direct sequencing, and may be useful as a routine method for detecting HBV NA resistance-associated mutations in clinical laboratory settings.