Augmented humoral and cellular immunity against severe acute respiratory syndrome coronavirus 2 after breakthrough infection in kidney transplant recipients who received 3 doses of coronavirus disease 2019 vaccine.

Diminished immune response to coronavirus disease 2019 (COVID-19) vaccines and breakthrough infection (BI) is a major concern for solid organ transplant recipients. Humoral and cellular immune responses of kidney transplant (KT) recipients after a third COVID-19 vaccination were investigated compared to matched health care workers. Anti-severe acute respiratory syndrome coronavirus 2 spike protein antibody and severe acute respiratory syndrome coronavirus 2 specific interferon-gamma releasing assay (IGRA) were assessed. A total of 38 KT recipients, including 20 BI and 18 noninfection, were evaluated. In the KT BI group, antibody titers were significantly increased (median 5 to 724, binding antibody units/mL (P = 0.002) after the third vaccination, but IGRA responses were negligible. After BI, antibody titers increased (median 11 355 binding antibody unit/mL; P < 0.001) and there was a significant increase of IGRA responses to spike proteins (Spike1-Nil, median 0.05 to 0.41 IU/mL; P = 0.009). Antibody titers and IGRA responses were significantly higher in the BI than in the noninfection group after 6 months. Immune responses were stronger in the health care worker than in the KT cohort, but the gap became narrower after BI. In conclusion, KT recipients who experienced BI after 3 COVID-19 vaccinations acquired augmented humoral and cellular immune responses.

Triple immune modulator therapy for aberrant hyperinflammatory responses in severe COVID-19.

A dysregulated hyperinflammatory response is a key pathogenesis of severe COVID-19, but optimal immune modulator treatment has not been established. To evaluate the clinical effectiveness of double (glucocorticoids and tocilizumab) and triple (plus baricitinib) immune modulator therapy for severe COVID-19, a retrospective cohort study was conducted. For the immunologic investigation, a single-cell RNA sequencing analysis was performed in serially collected PBMCs and neutrophil specimens. Triple immune modulator therapy was a significant factor in a multivariable analysis for 30-day recovery. In the scRNA-seq analysis, type I and II IFN response-related pathways were suppressed by GC, and the IL-6-associated signature was additionally downregulated by TOC. Adding BAR to GC and TOC distinctly downregulated the ISGF3 cluster. Adding BAR also regulated the pathologically activated monocyte and neutrophil subpopulation induced by aberrant IFN signals. Triple immune modulator therapy in severe COVID-19 improved 30-day recovery through additional regulation of the aberrant hyperinflammatory immune response.

Breakthrough invasive fungal infection in patients with myeloid malignancy receiving posaconazole tablet prophylaxis: Clinical features, risk factors, and posaconazole profiles.

Posaconazole (PSC) delayed-release tablet prophylaxis is the standard of care for preventing invasive fungal infection (IFI) in patients with acute myeloid leukemia undergoing myelosuppressive chemotherapy. The clinical features, risk factors, and PSC profiles of breakthrough IFI (bIFI) in patients receiving PSC tablet prophylaxis were investigated. A single-center retrospective cohort study was conducted, including adult patients with myeloid malignancy who received prophylactic PSC tablets while undergoing chemotherapy from June 2016 to June 2021. Logistic regression analysis was used to identify risk factors for bIFI. A receiver operating characteristic curve was used to predict the relationship between PSC trough level at steady state and bIFI. A total of 434 patients with myeloid malignancy who received PSC tablets were screened. A total of 10 patients with bIFI were compared with 208 non-IFI patients. There were four proven and six probable IFI cases, nine due to Aspergillus, and one due to Fusarium species. The bIFI patients had higher in-hospital mortality (30.0%) than the non-IFI patients (1.9%; P < 0.001). History of allogeneic hematopoietic stem cell transplantation (odds ratio [OR] 6.27; 95% confidence interval [CI] 1.63-24.09), prolonged neutropenia ≥28 days (OR 4.33; 95% CI 1.20-15.70), and low plasma PSC concentration <0.7 µg/ml (OR 16.33; 95% CI 4.15-64.26) were risk factors for bIFI. The optimal cutoff value of plasma PSC concentration predicting bIFI was 0.765 µg/ml (sensitivity, 60.0%; specificity, 91.3%; area under the curve, 0.746). bIFI was not uncommon in patients with myeloid malignancy receiving PSC tablet prophylaxis and associated with poor outcomes. Therapeutic drug monitoring may still be necessary, even in patients receiving PSC tablets.

Invasive fungal infections increase mortality in acute myeloid leukemia patients. This study investigated breakthrough invasive fungal infection cases in patients receiving posaconazole tablet prophylaxis. Our results will contribute to improving the outcome of patients with myeloid malignancy.

Risk factors and clinical outcomes of cytomegalovirus diseases in hematologic malignancy patients without hematopoietic stem-cell transplantation.

PURPOSE: This study aims to evaluate the risk factors and prognosis for CMV diseases in hematologic malignancy patients without hematopoietic stem-cell transplantation (HSCT). METHODS: We performed a case-control study (1:2) between 2012 and 2022. Adults with pathologic-confirmed CMV diseases (n=60) among hematologic malignancy patients were matched and compared to whom without CMV disease. RESULTS: Lymphoma was the most common underlying malignancy, and gastrointestinal tract involvement was the most common CMV disease. In the case group, high-dose steroid administration and transfusion within one month before diagnosis were higher (p<0.001). Steroid administration (aOR=5.78; 95% confidence interval: 1.25-26.68, p=0.024), red blood cell transfusion within one month (aOR=14.63; 2.75-77.76, p=0.002), low BMI (aOR=13.46, 2.07-87.45, p=0.006), and hypoalbuminemia (aOR=26.48, 5.93-118.17, p<0.001) were independent risk factors associated with CMV disease. The 30-day mortality was higher in the case group and CMV disease was significantly associated with all-cause mortality (aOR=14.41, 3.23-64.31, p<0.001). CONCLUSION: In hematologic malignancy patients without HSCT, risk factors for CMV organ disease included high-dose steroid administration and RBC transfusion within one month, low BMI, and hypoalbuminemia. Overall mortality was significantly higher with CMV disease, and CMV disease occurrence was a significant risk factor for mortality.

Clinical Utility of Sero-Immunological Responses Against SARS-CoV-2 Nucleocapsid Protein During Subsequent Prevalence of Wild-Type, Delta Variant, and Omicron Variant.

As nucleocapsid protein of severe acute respiratory syndrome coronavirus 2 is immunogenic but not targeted in vaccines, it could be useful in distinguishing natural infection from vaccination. We aimed to investigate the clinical utility of sero-immunological responses against the nucleocapsid protein. Nucleocapsid antibody immunoassay study with 302 coronavirus disease 2019 (COVID-19) patients showed lower titers in immunocompromised patients (P < 0.001), higher titers in higher severity (P = 0.031), and different seroconversion rates and titers according to variants of concern. Longitudinal evaluation of nucleocapsid antibodies using 513 samples from 291 COVID-19 patients revealed that it could persist up to 556 days from symptom onset. Interferon gamma release assay against the nucleocapsid protein showed poor response, precluding the deduction of a cut-off for the nucleocapsid protein. In conclusion, nucleocapsid antibody provides instructive clues about the immunogenicity of nucleocapsid proteins by different seroconversion rates and titers according to the severity of infection, host immune status, and different variants of concern.

Loss of Neutralizing Activity of Tixagevimab/Cilgavimab (Evusheld™) Against Omicron BN.1, a Dominant Circulating Strain Following BA.5 During the Seventh Domestic Outbreak in Korea in Early 2023.

Tixagevimab/cilgavimab is a monoclonal antibody used to prevent coronavirus disease 2019 among immunocompromised hosts and maintained neutralizing activity against early omicron variants. Omicron BN.1 became a dominant circulating strain in Korea early 2023, but its susceptibility to tixagevimab/cilgavimab is unclear. We conducted plaque reduction neutralization test (PRNT) against BN.1 in a prospective cohort (14 patients and 30 specimens). BN.1 PRNT was conducted for one- and three-months after tixagevimab/cilgavimab administration and the average PRNT ND50 of each point was lower than the positive cut-off value of 20 (12.9 ± 4.5 and 13.2 ± 4.2, respectively, P = 0.825). In the paired analyses, tixagevimab/cilgavimab-administered sera could not actively neutralize BN.1 (PRNT ND50 11.5 ± 2.9, P = 0.001), compared with the reserved activity against BA.5 (ND50 310.5 ± 180.4). Unlike virus-like particle assay, tixagevimab/cilgavimab was not active against BN.1 in neutralizing assay, and would not be effective in the present predominance of BA.2.75 sublineages.

Pharmacokinetic/pharmacodynamic parameters of vancomycin for predicting clinical outcome of enterococcal bacteremia.

PURPOSE: To find pharmacokinetic/pharmacodynamic parameters of vancomycin associated with the optimal outcome of severe infection due to Enterococcus species. METHODS: We retrospectively reviewed enterococcal bacteremia cases treated with vancomycin from January 2015 to December 2020. The primary outcome was 30-day mortality. We calculated cutoff values of the ratio of vancomycin area under the concentration-time curve over 24 h to the minimum inhibitory concentration (AUC24/MIC) and trough concentration (Ctrough) during the initial 72 h of treatment. The optimal cutoff value was determined using the Youden index. Binary variables created based on these cutoffs were further assessed using multivariable analysis. RESULTS: A total of 65 patients were included. The majority (87.7%) had solid or hematologic malignancies. Thirty-day mortality and nephrotoxicity occurred in nine (13.4%) and 14 (21.5%) patients, respectively. Both vancomycin AUC24/MIC and Ctrough showed fair performance in predicting 30-day mortality (AUC of receiver-operator curve for AUC24/MIC, 0.712; 95% confidence interval [CI] 0.539-0.886; AUC for Ctrough, 0.760; 95% CI 0.627-0.892; pairwise AUC comparison: p = 0.570). Ctrough ≥ 13.94 µg/mL, but not AUC24/MIC ≥ 504, had a significant association with 30-day mortality after adjusting for confounders (odds ratio, 8.40; 95% CI 1.60-86.62; p = 0.010). CONCLUSION: Mean Ctrough ≥ 13.94 µg/mL during the initial 72 h was associated with higher 30-day mortality in enterococcal bacteremia. Further studies are warranted to elucidate optimal pharmacokinetic targets for enterococcal bacteremia.

Paper-Based Multiplex Surface-Enhanced Raman Scattering Detection Using Polymerase Chain Reaction Probe Codification.

We construct a multiplex surface-enhanced Raman scattering (SERS) platform based on a plasmonic paper substrate and a double-labeled probe for the detection of multiple fluorescent dyes at high sensitivity in a single-wavelength light source system. Plasmonic paper, made of silver nanodots on three-dimensional cellulose fibers, enables highly sensitive SERS biosensing based on localized surface plasmon resonance (LSPR). The proposed method enables the identification and quantification of a range of fluorescent dyes ranging from picomolar to millimolar concentrations. The use of 5' fluorescent dyes and 3' biotin-modified probes as SERS-coded probes renders possible the separation of fluorescent dyes with streptavidin-coated magnetic beads (SMBs) and the sensitive detection of multiple dyes after the reverse transcription polymerase chain reaction (RT-PCR). This experimental study reveals the multiplex detection capability of PCR-based SERS under existing PCR conditions without modifying primer and probe sequences. The combination of magnetic bead-based separation and paper SERS platform is efficient, economical, and can be used for the simultaneous detection of two or more pathogens.

Poor outcomes of early recurrent post-transplant bloodstream infection in living-donor liver transplant recipients.

Bloodstream infection (BSI) is a common complication after living-donor liver transplantation (LDLT). Some patients develop recurrent BSIs. We evaluated the impacts of early recurrent BSIs (ER-BSIs) on outcomes in LDLT recipients. LDLT cases between 2008 and 2016 were included. Early BSI (E-BSI) was defined as a BSI event that occurred within 2 months after LDLT. ER-BSIs were defined as new-onset BSIs within 2 months due to another pathogen at a ≥ 48-h interval or a relapse of BSIs by the same pathogen at a ≥ 1-week interval, with negative cultures in between. The primary objective was evaluating the all-cause mortality of each group of LDLT recipients (90 days and 1 year). The secondary objectives were analyzing associated factors of each all-cause mortality and risk factors for early single BSI and ER-BSI. Among 727 LDLT recipients, 108 patients experienced 149 events of E-BSI with 170 isolated pathogens. Twenty-eight patients (25.9%, 28/108) experienced ER-BSI. The 1-year survival rates of patients without BSI, with early single BSI event, and with ER-BSIs were 92.4%, 81.3%, and 28.6%, respectively. ER-BSI was the most significant risk factor for 1-year mortality (adjusted HR = 5.31; 95% CI = 2.27-12.40). Intra-abdominal and/or biliary complications and early allograft dysfunction were risk factors for both early single BSI and ER-BSI. Interestingly, longer cold ischemic time and recipient operative time were associated with ER-BSI. LDLT recipients with ER-BSI showed very low survival rates accompanied by intra-abdominal complications. Clinicians should prevent BSI recurrence by being aware of intra-abdominal complications.

Impact of Difficult-to-Treat Resistance in Gram-negative Bacteremia on Mortality: Retrospective Analysis of Nationwide Surveillance Data.

BACKGROUND: Clinically relevant categorization of antimicrobial resistance is critical to mitigating the threat it poses. Difficult-to-treat resistance (DTR) is a recently proposed category defined as nonsusceptibility to all first-line antibiotic agents. METHODS: A retrospective study was conducted with nonduplicate cases of gram-negative bloodstream infection (GNBSI) caused by 4 major taxa (Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Acinetobacter species) identified from a nationwide surveillance database. DTR was defined as nonsusceptibility to all the ß-lactams and fluoroquinolones tested. Patient characteristics and mortality were compared between DTR GNBSI and GNBSI caused by carbapenem-resistant but not DTR and extended-spectrum cephalosporin-resistant but not DTR isolates using Centers for Disease Control and Prevention definitions. Adjusted odds ratios (aORs) for 30-day in-hospital mortality were examined for DTR in overall and in propensity score-matched cohorts. RESULTS: A total of 1167 episodes of monomicrobial GNBSI were identified, and 147 (12.6%) of the isolates were DTR. The majority of DTR isolates were Acinetobacter species (79.6%) and P. aeruginosa (17.7%). DTR infections were associated with previous antibiotic use, healthcare contact, ventilator use, and lower respiratory tract infection. Crude mortality for GNBSI caused by DTR was 50.3%. A multivariable model showed that only DTR, but not other categories, was significantly associated with mortality (adjusted odds ratio [aOR], 3.58 [95% confidence interval {CI}, 1.27-10.19]). DTR was also a significant predictor for mortality in the analysis of propensity score-matched cohorts (aOR, 3.48 [95% CI, 1.82-6.79]). CONCLUSIONS: In patients with GNBSI, DTR was associated with higher mortality than those in other resistance categories. Our findings suggest that DTR could be useful for surveillance and prognostication.

Co-introduction of plasmids harbouring the carbapenemase genes, blaNDM-1 and blaOXA-232, increases fitness and virulence of bacterial host.

BACKGROUND: Bacterial isolates with multiple plasmids harbouring different carbapenemase genes have emerged and been identified repeatedly, despite a general notion that plasmids confer fitness cost in bacterial host. In this study, we investigated the effects of plasmids with carbapenemase genes on the fitness and virulence of bacteria. METHODS: Different plasmids harbouring the carbapenemase genes, blaNDM-1 and blaOXA-232, were isolated from a carbapenem-resistant K. pneumoniae strain. Each plasmid was conjugated into the Escherichia coli strain DH5α, and a transconjugant with both plasmids was also obtained by transformation. Their in vitro competitive ability, biofilm formation, serum resistance, survival ability within macrophage and fruit fly, and fly killing ability were evaluated. RESULTS: The transconjugants with a single plasmid showed identical phenotypes to the plasmid-free strain, except that they decreased fly survival after infection. However, significantly increased fitness, virulence and biofilm production were observed consistently for the transconjugant with both plasmids, harbouring blaNDM-1 and blaOXA-232. CONCLUSIONS: Our data indicate that bacteria carrying multiple plasmids encoding different carbapenemases may have increased fitness and virulence, emphasizing the need for diverse strategies to combat antimicrobial resistance.

Emergence of vancomycin-resistant Enterococcus faecium ST1421 lacking the pstS gene in Korea.

Although multilocus sequence typing (MLST) has been used to study molecular epidemiology and to explore the population structure of Enterococcus faecium, vancomycin-resistant E. faecium (VREF) strains lacking the pstS gene that were non-typable using conventional MLST methods were reported recently. We found nationwide emergence of VREF isolates lacking pstS in Korea and hereby report the molecular characteristics of these isolates. Forty-six VREF isolates lacking the pstS gene were identified among 300 VREF rectal isolates collected from hospitalized patients between 2014 and 2015. MLST was performed and clonal relatedness was determined by pulsed-field gel electrophoresis (PFGE). Four VREF ST1421 isolates were whole-genome sequenced. Among the VREF rectal isolates lacking pstS, 98% were classified as ST1421, which has identical allelic profiles to ST17 for all housekeeping genes except pstS. PFGE pattern analyses revealed 32 pulsotypes. All isolates harbored Tn1546 components with various transposase and insertion sequences. The whole-genome sequencing of four VREF ST1421 isolates showed that the pstS gene region was deleted at various locations with considerable inversion. The pstS gene was also depleted in 12.1% of 33 VREF clinical isolates in 2006-2007 and in 11.8% of 59 clinical isolates in 2012-2013. VREF ST1421 strains lacking the pstS gene have emerged in Korea. The emergence and spread of pstS-deleted VREF strains pose a serious challenge for epidemiological investigation. Alternative molecular typing methods to MLST will be increasingly necessary.

Clinical factors influencing the performance of bacterial multiplex polymerase chain reaction in patients with community-onset pneumonia.

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.

Risk factors for development and mortality of invasive pulmonary Aspergillosis in kidney transplantation recipients.

Invasive pulmonary aspergillosis (IPA) is a high mortality opportunistic infection among kidney transplant recipients. This study assessed the risk factors and outcomes of IPA after KT. A retrospective study was conducted at a tertiary-care referral hospital in Korea. Electronic medical records of patients diagnosed with IPA after KT between February 1995 and March 2015 were reviewed. The control patients comprised two patients who received KT before and after each IPA case. Twenty-six cases were diagnosed with IPA among 1963 recipients at a median of 58 years old. The most common cause of end-stage renal disease was diabetic nephropathy. The median time to diagnosis was 161 days. Delayed graft function was associated with the development of IPA. The overall 12-week mortality rate of IPA was 57.5%. Serum GM level ≥ 2 and BAL GM level ≥ 5 were associated with 12-week mortality in the Kaplan-Meier survival analyses. Approximately half of IPA in KT recipients developed during the late posttransplant period (> 6 months), especially after treatment for acute rejection. Careful monitoring for IPA is required in patients with delayed graft function, DM, and who received rejection therapy. Higher serum and BAL GM were associated with 12-week mortality.

Clinical effectiveness of oral antimicrobial therapy for acute pyelonephritis caused by extended-spectrum ß-lactamase-producing Enterobacteriales.

Infections caused by extended-spectrum ß-lactamase-producing Enterobacteriales (ESBL-PE) are commonly treated with intravenous antibiotics. This study investigated whether oral antimicrobial therapy (OAT) is as effective as intravenous antimicrobial therapy (IVT) for acute pyelonephritis (APN) caused by ESBL-PE. A retrospective cohort of patients with APN caused by ESBL-PE was studied at a tertiary-care hospital from January 2014 through December 2016. The OAT group comprised patients treated with an appropriate oral antimicrobial agent following 7 days or less of IVT. The primary endpoint was treatment failure defined as clinical and/or microbiological failure. The secondary endpoint was length of hospital stay and recurrences of APN within 2 months and within 1 year. Propensity score matching and multivariable Cox proportional hazard modeling were used to minimize bias. Among 238 eligible cases, Escherichia coli (83.6%) was the most common pathogen. Sixty patients received OAT after a median of four days of appropriate IVT, and 178 patients completed treatment with IVT. Fluoroquinolones (58.3%) were the most commonly prescribed OAT, followed by trimethoprim-sulfamethoxazole and amoxicillin-clavulanate. OAT was not associated with treatment failure (adjusted OR 0.66; 95% CI 0.18-2.44) and hospitalization length was shorter in the OAT group (6.2 days versus 10.7 days; P < 0.01). APN recurrence caused by ESBL-PE infection within 2 months was not associated with OAT (adjusted HR 0.56; 95% CI 0.16-2.00). OAT reduced hospital stay without adverse effects on treatment outcome. OAT could be safely applied as a carbapenem-saving option in treatment of APN.

A case of gonococcal meningitis caused by Neisseria gonorrhoeae MLST ST7363 in a healthy young adult.

A 37-year-old healthy man was admitted with fever, skin rash, migratory arthralgia, and headache without preceding urogenital symptoms. Sexual contact history and positive CSF culture for Neisseria gonorrhoeae using BacT/Alert blood culture bottles were diagnostic for gonococcal meningitis. Multilocus sequence typing of this isolate showed sequence type (ST) 7363, the most predominant ST among ceftriaxone-resistant strains. The isolate from this case remained susceptible to ceftriaxone although it was resistant to penicillin, tetracycline, and ciprofloxacin. With the high selective pressure of ceftriaxone for treatment of plasmid-mediated ß-lactamase producing N. gonorrhoeae, resistance to ceftriaxone and molecular characteristics should be monitored.

Antifungal prophylaxis with posaconazole tablet and oral suspension in patients with haematologic malignancy: Therapeutic drug monitoring, efficacy and risk factors for the suboptimal level.

BACKGROUND: Posaconazole is used to prevent invasive fungal infections (IFIs) in patients with haematologic malignancy. In this study, we compared plasma posaconazole concentrations (PPCs) and the incidence of breakthrough IFIs between patients with haematologic malignancy receiving posaconazole oral suspension vs tablet. METHODS: We retrospectively collected data on adult patients with haematologic malignancies who received posaconazole prophylaxis during chemotherapy from April 2014 through May 2018. A total of 242 cases with PPCs, 88 in the oral suspension group and 154 in the tablet group, were included in this study. RESULTS: Patients receiving tablets achieved a significantly higher mean PPC than did those on oral suspension (1.631 ± 0.878 µg/mL in the tablet group vs. 0.879 ± 0.585 µg/mL in the oral suspension group). One hundred and thirty-seven of 154 patients (89.0%) receiving tablets had PPCs of 0.7 µg/mL or more, while only 41 of 88 patients (46.6%) receiving oral suspension attained an optimal level (P < .001). The incidence of breakthrough IFIs was significantly higher in the oral suspension group compared with in the tablet group (14.8% of oral suspension vs. 4.5% of tablet; P = .005). In the analysis including patients receiving posaconazole tablets, hypoalbuminemia (< 3.5 g/dL) was found to be a risk factor associated with suboptimal levels (odds ratio: 8.872; 95% confidence interval: 3.011 - 26.141; P < .001). CONCLUSIONS: Suboptimal PPCs in the tablet group were less common than those in the oral suspension group. Therapeutic drug monitoring may be still necessary even in patients receiving posaconazole tablets, especially in those with hypoalbuminemia.

Quantitative Serodiagnosis of Scrub Typhus Using Surface-Enhanced Raman Scattering-Based Lateral Flow Assay Platforms.

A surface-enhanced Raman scattering-based lateral flow assay (SERS-LFA) technique has been developed for the rapid and accurate diagnosis of scrub typhus. Lateral flow kits for the detection of O. tsutsugamushi IgG (scrub typhus biomarker) were fabricated, and the calibration curve for various standard clinical sera concentrations were obtained by Raman measurements. The clinical sera titer values were determined by fitting the Raman data to the calibration curve. To assess the clinical feasibility of the proposed method, SERS-LFA assays were performed on 40 clinical samples. The results showed good agreement with those of the standard indirect immunofluorescence assay (IFA) method. SERS-LFA has many advantages over IFA including the less sample volume, simpler assay steps, shorter assay time, more systematic quantitative analysis, and longer assay lifetime. As SERS strips can be easily integrated with a miniaturized Raman spectrophotometer, field serodiagnosis is also more feasible.

Evolution of Klebsiella pneumoniae with mucoid and non-mucoid type colonies within a single patient.

We obtained nine Klebsiella pneumoniae isolates successively isolated from a single patient. Four pairs (M1-M4 and NM1-NM4) obtained simultaneously from the same site showed different colony types, mucoid and non-mucoid, while the final isolate (M5) was isolated alone from the blood and showed a mucoid phenotype. The whole genome of isolate M5 was sequenced de novo using the PacBio RSII system, while the others were sequenced with an Illumina Hiseq4000 and mapped to the genome sequences of M5. To identify insertions or deletions in the cps locus, we amplified and sequenced cps locus genes. We identified insertion sequence (IS) elements in several genes of the cps locus or one amino acid substitution in WcaJ in all non-mucoid isolates. Five additional amino acid alterations in RpsJ, LolE, Lon-2, PpsE, and a hypothetical protein were detected in some mucoid and non-mucoid isolates. Based on the genome data and cps locus sequences, the mucoid phenotype may have been lost or converted into the non-mucoid phenotype because of the insertion of IS elements or amino acid alterations at this locus. We inferred a within-host evolutionary scenario, in which non-mucoid variants emerged repeatedly from mucoid isolates, but may be short-lived because of their low fitness.

Plasmid analysis of Escherichia coli isolates from South Korea co-producing NDM-5 and OXA-181 carbapenemases.

Recently, Escherichia coli isolates co-producing New Delhi metallo-ß-lactamase (NDM)-5 and oxacillinase (OXA)-181 were identified in a tertiary-care hospital of South Korea. Isolate CC1702-1 was collected from urine in January 2017 and isolate CC1706-1 was recovered from a transtracheal aspirate of a hospitalized patient in May 2017. Carbapenemase genes were identified by multiplex PCR and sequencing, and whole genome sequencing was performed subsequently using the PacBio RSII system. Both E. coli isolates belonged to the same clone (ST410) and were resistant to all ß-lactams including carbapenems. We obtained whole plasmid sequences of the isolates: pCC1702-NDM-5 from CC1702-1 and pCC1706-NDM-5 and pCC1706-OXA-181 from CC1706-1. The two E. coli isolates belonged to the same clone (ST410) and they were completely resistant to all ß-lactams, as well as carbapenems. Two blaNDM-5-harboring plasmids belonged to the same incompatibility group, IncFIA/B, and consisted of 79,613 bp and 111,890 bp with 87 and 130 coding sequences, respectively. The genetic structures of the two blaNDM-5-bearing plasmids, which were distinct from the blaNDM-5-bearing plasmids from the Klebsiella pneumoniae isolates previously transmitted from the United Arab Emirates (UAE) to South Korea, differed from each other. While pCC1702-NDM-5 showed high degree of identity with the plasmid from a multidrug-resistant isolate of Citrobacter fruendii P5571 found in China, pCC1706-NDM-5 was very similar to the plasmid from a multidrug-resistant isolate of E. coli AMA1176 found in Denmark. pCC1706-OXA-181, which was a 51 kb, self-transmissible IncX3 plasmid, was identical to the E. coli plasmids pAMA1167-OXA-181 from Denmark and pOXA-181-WCHEC14828 from China. Plasmids harboring blaNDM-5 in E. coli isolates might not be transferred from K. pneumoniae isolates co-producing NDM-5 and OXA-181. They probably originated from multiple sources.