Objective: The aim of this study was to evaluate the effectiveness of metagenomic next-generation sequencing (mNGS) for the diagnosis of Pneumocystis jirovecii Pneumonia (PCP) in critically pediatric patients. Methods: Seventeen critically pediatric patients with PCP and sixty patients diagnosed with non-PCP pneumonia who were admitted in pediatric intensive care unit between June 2018 and July 2021 were enrolled. Conventional methods and mNGS for detecting Pneumocystis jirovecii (P. jirovecii) were compared. The patients’ demographics, comorbidities, laboratory test results, antibiotic treatment response and 30 day mortality were analyzed. Result: The mNGS showed a satisfying diagnostic performance with a sensitivity of 100% in detecting P. jirovecii compared with Gomori methenamine silver staining (5.9%), serum (1,3)-β-D-glucan (86.7%) and and LDH (55.6%). The diagnostic specificity of mNGS for PCP was higher than that of serum BDG (56.7%) and LDH (71.4%). In PCP group, over one thirds’ cases had mixed infections. Compared with survivors, non-survivors had higher stringently mapped read numbers (SMRNs) in bronchoalveolar lavage fluid (BALF) sample (P < 0.05), suggesting SMRNs were closely associated with the severity of response. The detection for P. jirovecii by mNGS both in BALF and blood samples reached a concordance rate of 100%, and the SMRNs in the BALF were remarkably higher than that in blood samples. Initial antimicrobial treatment was modified in 88.2% of PCP patients based on the mNGS results. Conclusion: The mNGS is a potential and efficient technology in diagnosing PCP and shows a satisfying performance in the detection of co-pathogens. Both blood and BALF samples for mNGS are suggested for the presumptive diagnosis of PCP.

Invasive pulmonary aspergillosis (IPA) in patients with diabetes mellitus has high incidence, especially in Type 2 diabetes mellitus (T2DM). The aim of this study was to evaluate the diagnostic efficacy of metagenomic next-generation sequencing (mNGS) for IPA in patients with T2DM. A total of 66 patients with T2DM were included, including 21 IPA and 45 non-IPA patients, from January 2022 to December 2022. The demographic characteristics, comorbidities, laboratory test results, antibiotic treatment response, and 30-day mortality rate of patients were analyzed. The diagnostic accuracy of mNGS and conventional methods was compared, including sensitivity, specificity, positive predictive value and negative predictive value. The sensitivity and specificity of mNGS were 66.7% and 100.0%, respectively, which were significantly higher than those of fluorescence staining (42.1% and 100%), serum 1,3-β-D-glucan detection (38.1% and 90.9%), serum galactomannan detection (14.3% and 94.9%) and BALF galactomannan detection (47.3% and 70.7%). Although the sensitivity of BALF culture (75.0%) was higher than that of mNGS (66.7%), the turnover time of mNGS was significantly shorter than that of traditional culture (1.6 days vs. 5.0 days). The sensitivity of mNGS combined with BALF culture reached 100.0%. In addition, mNGS has a stronger ability to detect co-pathogens with IPA. 47.6% of T2DM patients with IPA were adjusted the initial antimicrobial therapy according to the mNGS results. This is the first study to focus on the diagnostic performance of mNGS in IPA infection in T2DM patients. MNGS can be used as a supplement to conventional methods for the diagnosis of IPA in patients with T2DM.

Background The type VI secretion system (T6SS) has been recognized as a novel virulence factor in Klebsiella pneumoniae. This study investigated the occurrence of T6SS genes in carbapenem-resistant Klebsiella pneumoniae (CRKP) strains during intestinal colonization and evaluated their effect on the development of bloodstream infections. Methods The study encompassed 2,385 patients admitted to the intensive care unit (ICU) and subjected to routine screening for intestinal colonization with CRKP. PFGE was employed on CRKP strains isolated from both the patients’ intestine and blood cultures, confirming their genetic similarity. PCR was employed to detect the presence of carbapenemase genes, T6SS genes, and virulence genes. Quantitative real-time PCR was conducted to assess the expression levels of the core genes associated with the T6SS. The correlation between T6SS expression and sBSI was further investigated. Results Approximately 10% (238/2385) of ICU patients tested positive for CRKP colonization. Among patients who tested positive, 10.1% (24/238) developed CRKP-sBSI. Patients carrying T6SS-positive CRKP isolates were more commonly linked to a history of invasive procedures, antibiotic use, and immunosuppression (P < 0.05), and were strongly associated with 28-day mortality (P < 0.001). It indicated that T6SS-positive CRKP strains exhibited a higher prevalence of virulence genes, such as rmpA and iucA, compared to T6SS-negative ones (P < 0.001). Compared to the strains isolated from simple colonization group, there was a significant increase in the mRNA expression of both hcp and vgrG genes (P < 0.05) of strains from the sBSI group, suggesting the key genes of the T6SS may play a significant role in the occurrence and progression of sBSI caused by CRKP. Conclusion The presence of the T6SS in a CRKP strain from intestinal colonization can serve as a promising predictive marker for sBSI. Conducting screenings for CRKP in patients’ intestinal flora and monitoring T6SS carriage can improve the prevention and management of CRKP bloodstream infections.

Background: Carbapenem-resistant Enterobacteriaceae (CRE) is rapidly becoming a major threat to hospitalized children worldwide. The purpose of this study was to summarize etiological characteristics and identify risk factors relevant to CRE bloodstream infection (BSI) and short-term mortality among pediatric patients in China. Methods: In this study, we included 370 inpatients ≤ 17 years old with BSI caused by Enterobacteriaceae in China from January 2013 to December 2022. By collecting data on demographics, etiological features, and clinical outcomes, we conducted an in-depth analysis. Results: Among the 370 BSI patients with infections caused by Enterobacteriaceae, 35 patients (9.46%) were caused by CRE. Among these CRE strains, Klebsiella pneumoniae (49.46%) was the most important pathogen of BSI in pediatric patients, followed by Escherichia coli (31.62%) and Enterobacter cloacae (5.95%). The most frequent carbapenemase was NDM (23/35, 65.71%), followed by KPC (8/35, 22.86%). The overall 28-day mortality rate of children with an Enterobacteriaceae BSI episode was 1.89% (7/370), of which CRE BSI patients (3/35, 8.57%) were significantly higher than CSE patients (4/335, 1.19%, P < 0.001). Congenital malformation (OR: 8.162, 95% CI: 3.859– 16.680, P < 0.001) and catheter-related (OR: 6.645, 95% CI: 3.159– 13.28, P: < 0.001) were associated with the development of CRE BSI in pediatric patients. A multivariate analysis showed that the infection of CRE (OR 7.758, 95% CI 1.869– 29.62, P = 0.021) were independent risk factors for 28-day mortality of Enterobacteriaceae BSI. When the MIC of any carbapenems was ≥ 8 μg/mL, the mortality rate in the ICU was higher (P < 0.05). Conclusion: Congenital malformation, previous cephalosporin/carbapenems administration, and catheter-related conditions were closely related to the development of CRE BSI. The mortality rate of CRE BSI was higher. NDM was the predominant carbapenemase-producing mechanism in children.

Introduction: The rapid and accurate identification of carbapenemases in Enterobacterales isolates is of paramount importance for the selection of effective antibiotics and the control of hospital-acquired infections. Methods: This study aimed to evaluate the performance of two immunochromatographic methods, NG-Test Carba 5 (Carba 5) and Goldstream Carbapenem-resistant K.N.I.V.O. Detection K-Set (K-Set) for detecting five major carbapenemase (KPC, NDM, IMP, OXA-48-like, and VIM). Carbapenemase genes were confirmed by PCR. Results: In this study, a total of 245 carbapenem-resistant Enterobacterales (CRE) isolates were encompassed, with an overwhelming 96.7% of these strains exhibiting the ability to produce carbapenemase. A total of 58.2% of Klebsiella pneumoniae strains that produce KPC carbapenemase were the most prevalent among carbapenem-resistant Enterobacteriaceae (CRE). NDM-producing Klebsiella pneumoniae accounted for 30.4%. Importantly, NDM-type carbapenemase emerges as the predominant form in Escherichia coli and Enterobacter cloacae strains, accounting for 46 (93.9%) and 20 (83.3%) cases, respectively. The performance of the two methods in carbapenemase detection has demonstrated remarkable outcomes, exhibiting overall specificity and sensitivity exceeding 99%. Specifically, the K-Set accurately detected a unique KPC-carbapenemase in K. pneumoniae, whereas Carba 5 was unable to identify it. This was due to the presence of a novel bla_KPC gene, which harbored a specific point mutation (A to G) at nucleotide position 787, differentiating it from the bla_KPC-33 gene. Conclusion: These two methods, characterized by their simplicity, rapidity, and accuracy, are ideally suited for detecting carbapenemases in routine microbiology laboratories. They serve as a vital foundation for the rational selection of antibiotics in clinical practice.

Background: Elizabethkingia anophelis, an opportunistic pathogen that can cause infections in multiple parts of the human body, has multiple drug resistance and a high mortality rate. However, there have been few reports of infective endocarditis (IE) caused by Elizabethkingia anophelis, which means that diagnosis and treatment face challenges that cannot be ignored. Rapid and accurate identification and drug sensitivity results are needed to make timely treatment adjustments. Case Presentation: An 81-year-old man presented with recurrent fever and increased infection index for more than a month. Based on his clinical symptoms, infection index, reduplicative blood cultures, and results of transesophageal echocardiography, he was ultimately diagnosed with infective endocarditis caused by Elizabethkingia anophelis. The patient had a favorable outcome with a 6-week course of intravenous antibiotic therapy. Conclusion: This is a rare and successfully cured case of IE caused by the pathogen of Elizabethkingia anophelis, which is difficult not only in diagnosis but also in treatment. This case provides a certain referential significance to the treatment of Elizabethkingia anophelis-caused IE in clinical practice.