Etiological diagnostic performance of probe capture-based targeted next-generation sequencing in bloodstream infection.

Background: A rapid and precise etiological diagnosis is crucial for the effective treatment of bloodstream infection (BSI). In this study, the performance of probe capture-based targeted next-generation sequencing (tNGS) was compared to that of blood culture and metagenomic next-generation sequencing (mNGS) in detecting potential pathogens in patients with BSI. Methods: A total of 80 patients with suspected BSI were prospectively enrolled from 24 November 2023 to 30 December 2023 at Zhongshan Hospital, Shanghai, China. All 80 participants underwent simultaneous blood culture, blood mNGS, and blood tNGS after admission when febrile, and the results were compared. Results: Among the 80 participants, 11 were clinically diagnosed with noninfectious fever, and 69 were diagnosed with BSI. Blood tNGS had a higher sensitivity for the diagnosis of BSI than blood culture (91.3% vs. 23.2%, P<0.001) and blood mNGS (91.3% vs. 69.6%, P=0.001). There was no significant difference in specificity between blood mNGS and tNGS (81.8% vs. 100.0%, P=0.13). Blood tNGS demonstrated a faster turnaround time than blood culture and blood mNGS. In 22 (31.9%) patients with BSI, targeted adjustment of the anti-infectious therapy according to the blood tNGS results resulted in clinical improvement. Conclusions: Blood tNGS may be a promising tool for detecting potential pathogens in patients with BSI. The application of blood tNGS for BSI could guide anti-infectious treatment strategies and might improve clinical outcomes.

Accurate etiological diagnosis of Mycoplasma hominis mediastinitis in immunocompetent patients using metagenomic next-generation sequencing: a case series and literature review.

Background: As a culture-independent method, metagenomic next-generation sequencing (mNGS) is widely used in microbiological diagnosis with advantages in identifying potential pathogens, guiding antibiotic therapy, and improving clinical prognosis, especially in culture-negative cases. Mycoplasma hominis (M. hominis) mediastinitis is a rare and severe disease for which etiological diagnosis is important but challenging. The application of mNGS in the etiological diagnosis of mediastinitis has seldom been studied. Methods: By searching the electronic medical history retrieval system with "Mycoplasma hominis" and "mediastinitis", seven patients diagnosed with M. hominis mediastinitis were reviewed in Zhongshan Hospital, Fudan University, Shanghai from 9 December 2020 to 14 February 2023. Microbiological cultures and mNGS were conducted for blood, abscess, and/or mediastinal fluid. Adjustment of the antibiotic therapy due to mNGS was assessed. A literature review was conducted in the PubMed database beginning in 1970 for M. hominis infection and mediastinitis. Results: For the seven patients, cultures of blood, abscess, and mediastinal fluid were negative whereas mNGS identified M. hominis in serum, abscess, and/or mediastinal fluid and was used to guide specific antibiotic therapy. The stringent mapped reads number of genera (SMRNG), stringent mapped reads number of species (SMRN), and coverage rate of M. hominis detection by mNGS were significantly higher in body fluid (abscess or mediastinal fluid) than in serum. All seven patients had underlying heart diseases and underwent previous cardiac surgery. The most common symptoms were fever and sternal pain. After detection of M. hominis, antibiotics were adjusted to quinolones or doxycycline except for one patient, whose diagnosis was clarified after death. Two patients died. Literature review since 1970 identified 30 cases of extra-genital infection caused by M. hominis. Including our seven new cases, 2 (5.4%) were neonates and 35 (94.6%) were adults. Thirty (81.1%) cases were postoperative infection and 15 (40.5%) had implanted devices. Five patients (13.5%) died. Conclusions: mNGS might be a promising technology in the detection of fastidious pathogens such as M. hominis. Accurate etiological diagnosis by mNGS could guide antibiotic therapy and facilitate clinical management.

Mechanism of benzoxazinoids affecting the growth and development of Fusarium oxysporum f. sp. fabae.

Continuous cropping of faba bean (Vicia faba L.) has led to a high incidence of wilt disease. The implementation of an intercropping system involving wheat and faba bean can effectively control the propagation of faba bean wilt disease. To investigate the mechanisms of wheat in mitigating faba bean wilt disease in a wheat-faba bean intercropping system. A comprehensive investigation was conducted to assess the temporal variations in Fusarium oxysporum f. sp. fabae (FOF) on the chemotaxis of benzoxazinoids (BXs) and wheat root through indoor culture tests. The effects of BXs on FOF mycelial growth, spore germination, spore production, and electrical conductivity were examined. The influence of BXs on the ultrastructure of FOF was investigated through transmission electron microscopy. Eukaryotic mRNA sequencing was utilized to analyze the differentially expressed genes in FOF upon treatment with BXs. FOF exhibited a significant positive chemotactic effect on BXs in wheat roots and root secretions. BXs possessed the potential to exert significant allelopathic effects on the mycelial growth, spore germination, and sporulation of FOF. In addition, BXs demonstrated a remarkable ability to disrupt the structural integrity and stability of the membrane and cell wall of the FOF mycelia. BXs possessed the capability of posing threats to the integrity and stability of the cell membrane and cell wall. This ultimately resulted in physiological dysfunction, effectively inhibiting the regular growth and developmental processes of the FOF.

Benzoxazinoids secreted by wheat root weaken the pathogenicity of Fusarium oxysporum f. sp. fabae by inhibiting linoleic acid and nucleotide metabolisms.

KEY MESSAGE: The regulatory action of BXs secreted by wheat on the pathogenicity of FOF causing Fusarium wilt in faba bean were analyzed. DIMBOA and MBOA weakened the pathogenicity of FOF. A large number of pathogenic bacteria in continuous cropping soil infect faba bean plants, leading to the occurrence of wilt disease, which restricts their production. Faba bean-wheat intercropping is often used to alleviate this disease. This study investigates the effect of benzoxazinoids (BXs) secreted by wheat root on the pathogenicity of Fusarium oxysporum f. sp. Fabae (FOF) and underlying molecular mechanisms. The effects of DIMBOA(2,4-dihydroxy-7-methoxy-1,4-benzoxazine-4-one) and MBOA(6-methoxybenzoxazolin-2-one) on the activity of cell-wall-degrading enzymes in FOF(cellulase, pectinase, amylase, and protease), FOF Toxin (fusaric acid, FA) content were investigated through indoor culture experiments. The effect of BXs on the metabolic level of FOF was analyzed by metabonomics to explore the ecological function of benzoxazines intercropping control of Fusarium wilt in faba bean. The results show that the Exogenous addition of DIMBOA and MBOA decreased the activity of plant-cell-wall-degrading enzymes and fusaric acid content and significantly weakened the pathogenicity of FOF. DIMBOA and MBOA significantly inhibited the pathogenicity of FOF, and metabolome analysis showed that DIMBOA and MBOA reduced the pathogenicity of FOF by down-regulating related pathways such as nucleotide metabolism and linoleic acid metabolism, thus effectively controlling the occurrence of Fusarium wilt in faba bean.

Fever clinic construction and management targeted to prevention and control of healthcare-associated respiratory viral infections in Jiangsu, China: A cross-sectional observational study.

To analyze the post-COVID-19 construction and management of fever clinics targeted to prevention and control of healthcare-associated respiratory viral infections in medical institutions at all levels in China, and to provide a basis for promoting their standardized construction, we conducted this survey on the construction of fever clinics in 429 medical institutions of Jiangsu Province from July to December 2020. Contents of the questionnaire included the general situation of medical institutions, the construction status and future construction plans of fever clinics. We find the construction rate of fever clinic in medical institutions of Jiangsu province was 75.3%. All construction indicators, quality management systems and processes fail to fully meet the requirements of documents and standards. Jiangsu province actively promotes the construction of fever clinic layout, but there is still a gap with the construction standard. As a result, it is necessary to further promote standardized construction of fever clinic, and necessary financial input should be increased to expand all constructions of fever clinic in primary medical institutions.

Oral Simnotrelvir for Adult Patients with Mild-to-Moderate Covid-19.

BACKGROUND: Simnotrelvir is an oral 3-chymotrypsin-like protease inhibitor that has been found to have in vitro activity against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and potential efficacy in a phase 1B trial. METHODS: In this phase 2-3, double-blind, randomized, placebo-controlled trial, we assigned patients who had mild-to-moderate coronavirus disease 2019 (Covid-19) and onset of symptoms within the past 3 days in a 1:1 ratio to receive 750 mg of simnotrelvir plus 100 mg of ritonavir or placebo twice daily for 5 days. The primary efficacy end point was the time to sustained resolution of symptoms, defined as the absence of 11 Covid-19-related symptoms for 2 consecutive days. Safety and changes in viral load were also assessed. RESULTS: A total of 1208 patients were enrolled at 35 sites in China; 603 were assigned to receive simnotrelvir and 605 to receive placebo. Among patients in the modified intention-to-treat population who received the first dose of trial drug or placebo within 72 hours after symptom onset, the time to sustained resolution of Covid-19 symptoms was significantly shorter in the simnotrelvir group than in the placebo group (180.1 hours [95% confidence interval {CI}, 162.1 to 201.6] vs. 216.0 hours [95% CI, 203.4 to 228.1]; median difference, -35.8 hours [95% CI, -60.1 to -12.4]; P = 0.006 by Peto-Prentice test). On day 5, the decrease in viral load from baseline was greater in the simnotrelvir group than in the placebo group (mean difference [±SE], -1.51±0.14 log10 copies per milliliter; 95% CI, -1.79 to -1.24). The incidence of adverse events during treatment was higher in the simnotrelvir group than in the placebo group (29.0% vs. 21.6%). Most adverse events were mild or moderate. CONCLUSIONS: Early administration of simnotrelvir plus ritonavir shortened the time to the resolution of symptoms among adult patients with Covid-19, without evident safety concerns. (Funded by Jiangsu Simcere Pharmaceutical; ClinicalTrials.gov number, NCT05506176.).

Proteomic analysis of the faba bean-wheat intercropping system in controlling the occurrence of faba bean fusarium wilt due to stress caused by Fusarium oxysporum f. sp. fabae and benzoic acid.

BACKGROUND: In faba bean, continuous cropping severely affects plant growth and increases the incidence of fusarium wilt due to the accumulation of pathogens and autotoxic substances. The intercropping of faba bean and wheat is commonly used to alleviate the occurrence of fusarium wilt in the faba bean. OBJECTIVE: To investigate the role of Fusarium oxysporum f. sp. Fabae(FOF) and benzoic acid in the occurrence of faba bean fusarium wilt and unravel the potential mechanism of intercropping in alleviating its occurrence. METHODS: Hydroponic experiment was carried out using monocropping faba bean (M) and intercropping faba bean and wheat (I) patterns under FOF alone stress (M + F, I + F), FOF and benzoic acid double stress (M + F + B, I + F + B). The growth of faba bean seedlings under FOF and benzoic acid dual stresses were analyzed as well as the protein expression profile of monocropping and intercropping faba bean roots. RESULT: Under FOF stress, the growth of faba bean seedlings was inhibited, and the inhibitory effect was enhanced under the dual stress of FOF and benzoic acid. However, faba bean-wheat intercropping alleviated the inhibitory effect of FOF and benzoic acid on faba bean growth. In faba bean, the up-regulated protein was involved in different functions, such as redox, hydrogen peroxide decomposition, and metabolic processes under FOF stress (M + F, I + F) compared to the control. Compared with FOF stress (M + F, I + F), under the dual stress of FOF and benzoic acid (M + F + B, I + F + B), the up-regulated protein in faba bean were involved in intracellular redox balance, defense, and maintenance of cell integrity. Compared with monocropping (M, M + F, M + F + B), the up-regulated protein function of intercropping(I, I + F, I + F + B) was mainly involved in the biosynthesis of secondary metabolites, redox balance, biological carbon fixation of photosynthesis, and so on. KEGG enrichment analysis results showed that intercropping increased ethylene and jasmonic acid synthesis and other related pathways to improve resistance against fusarium wilt in the faba bean. CONCLUSION: The growth of faba bean was inhibited under FOF stress and the inhibitory effect was enhanced under the dual stress of FOF and benzoic acid, which promoted the occurrence of faba bean fusarium wilt. This might be due to the down-regulation of energy and cytoplasmic matrix proteins under FOF and benzoic acid stress. The faba bean wheat intercropping alleviated the inhibition of FOF and benzoic acid stress by up-regulating the biosynthesis of secondary metabolites, redox homeostasis, photosynthetic carbon fixation, and other related proteins. Besides, it also promoted the biosynthesis of ethylene, and jasmonic acid, improved the resistance of faba bean plants, and alleviated the occurrence of faba bean fusarium wilt. This provides a theoretical basis for the determination of jasmonic acid and ethylene content.

Epidemiology of Clinically Significant Aspergillus Species from a Large Tertiary Hospital in Shanghai, China, for the Period of Two Years.

Background: Aspergillus species are becoming a major public health concern worldwide due to the increase in the incidence of aspergillosis and emergence of antifungal resistance. In this study, we surveyed all Aspergillus species isolated from aspergillosis patients in Zhongshan Hospital Fudan University, Shanghai, China, from 2019 to 2021. Methods: We characterized the susceptibility profiles of these Aspergillus species to medical azoles (voriconazole, itraconazole and posaconazole) using YeastOneTM broth microdilution system. To determine the underlying antifungal resistance mechanisms in azole-resistant A. fumigatus (ARAf) isolates, we characterized mutations in the cyp51A gene. Genotypic diversity of sampled A. fumigatus was investigated using CSP-typing. Results: A total of 112 Aspergillus isolates (81 A. fumigatus, 17 A. flavus, 5 A. niger, 2 A. terreus, 2 A. lentulus, 2 A. oryzae, 1 A. nidulans, 1 A. versicolor and 1 A. sydowii) from 105 patients diagnosed with aspergillosis (including proven or probable invasive aspergillosis, chronic pulmonary aspergillosis, allergic bronchopulmonary aspergillosis and cutaneous aspergillosis) were obtained. Eight isolates (7 A. fumigatus and 1 A. niger) from seven patients were either azole non-susceptible or non-wild type. Azole non-susceptible or non-wild type rate was 7.1%/isolate and 6.7%/patient analysed. Four ARAf harbored TR34/L98H mutation, whereas one carried TR46/Y121F/T289A allele. The 81 A. fumigatus isolates were spread across 8 CSP types with t01 to be the predominant type (53.1%). ARAf isolates were distributed over CSP types t01, t02, t04A and t11. Conclusion: Results from this study provided us with an understanding of the antifungal resistance and related characteristics of Aspergillus species in Eastern China. Further comparisons of our results with those in other countries reflect potential clonal expansion of A. fumigatus in our region. Further surveillance study is warranted to guide antifungal therapy and for epidemiological purposes.

Anxiety and depression among patients with non-tuberculous mycobacterial disease in Shanghai: a cross-sectional study.

Objective: To understand the mental health status and its influencing factors among patients with non-tuberculous mycobacterial disease and to provide a reference for medical staff to formulate scientific and feasible intervention strategies. Methods: A total of 114 patients diagnosed with non-tuberculous mycobacillosis during hospitalization in the Department of Infection from September 2020 to April 2021 were selected as the research participants. Participants' mental health status and related factors were evaluated using a self-made general patient information questionnaire, self-rating Anxiety Scale (SAS), and self-rating Depression Scale (SDS). Results: Among 114 patients with non-tuberculous mycosis, 61 (53.51%) exhibited depressive symptoms, and the SDS score was 51.15 ± 13.04, which was higher than the national norm of 41.88 ± 10.57 (p < 0.05); further, 39 patients (34.21%) showed anxiety symptoms, and the SAS score was 45.75 ± 10.81, which was significantly higher than the national norm of 29.78 ± 10.07 (p < 0.05). Body mass index and monthly household income had significant effects on depression in patients with non-tuberculous mycobacterial disease (p < 0.05). Educational level had a significant effect on the anxiety state of patients with non-tuberculous mycobacterial disease (p < 0.05). Conclusion: Patients with non-tuberculous mycobacterial disease are prone to depression and anxiety. Nurses should pay attention to it in clinical work for the timely identification of and intervention for anxiety and depression and intervene.

Safety and immunogenicity of a modified COVID-19 mRNA vaccine, SW-BIC-213, as a heterologous booster in healthy adults: an open-labeled, two-centered and multi-arm randomised, phase 1 trial.

BACKGROUND: We assessed the safety and immunogenicity of a core-shell structured lipopolyplex (LPP) based COVID-19 mRNA vaccine, SW-BIC-213, as a heterologous booster in healthy adults. METHODS: We conducted an open-labeled, two-centered, and three-arm randomised phase 1 trial. Healthy adults, who had completed a two-dose of inactivated COVID-19 vaccine for more than six months, were enrolled and randomized to receive a booster dose of COVILO (inactivated vaccine) (n = 20) or SW-BIC-213-25µg (n = 20), or SW-BIC-213-45µg (n = 20). The primary study endpoint was adverse events within 30 days post-boosting. The secondary endpoint was the titers of binding antibodies and neutralizing antibodies against the wild-type (WT) of SARS-CoV-2 as well as variants of concern in serum. The exploratory endpoint was the cellular immune responses. This trial was registered with http://www.chictr.org.cn (ChiCTR2200060355). FINDINGS: Between Jun 6 and Jun 22, 2022, 60 participants were enrolled and randomized to receive a booster dose of SW-BIC-213 (25 µg, n = 20, or 45 µg, n = 20) or COVILO (n = 20). The baseline demographic characteristics of the participants at enrollment were similar among the treatment groups. For the primary outcome, injection site pain and fever were more common in the SW-BIC-213 groups (25 µg and 45 µg). Grade 3 fever was reported in 25% (5/20) of participants in the SW-BIC-213-45µg group but was resolved within 48 h after onset. No fatal events or adverse events leading to study discontinuation were observed. For secondary and exploratory outcomes, SW-BIC-213 elicited higher and longer humoral and cellular immune responses than that in the COVILO group. INTERPRETATION: SW-BIC-213, a core-shell structured lipopolyplex (LPP) based mRNA vaccine, was safe, tolerable, and immunogenic as a heterologous booster in healthy Chinese adults. FUNDING: Shanghai Municipal Government, the Science and Technology and Economic Commission of Shanghai Pudong New Area, and mRNA Innovation and Translation Center of Shanghai.

Association between the overall burden of comorbidity and Ct values among the older patients with Omicron infection: Mediated by inflammation.

Objectives: To investigate the associations between the overall burden of comorbidity, inflammatory indicators in plasma and Ct values among the elderly with COVID-19. Methods: We conducted a retrospective observational study. The results of each nucleic acid test of during hospitalization were obtained. Linear regression models assessed the associations between the overall burden of comorbidity, inflammatory indicators in plasma and Ct values among the elderly. A causal mediation analysis was performed to assess the mediation effects of inflammatory indicators on the association between the overall burden of comorbidity and Ct values. Results: A total of 767 COVID-19 patients aged ≥ 60 years were included between April 2022 and May 2022. Patients with a high burden of comorbidity had significantly lower Ct values of the ORF gene than subjects with a low burden of comorbidity (median, 24.81 VS 26.58, P < 0.05). Linear regression models showed that a high burden of comorbidity was significantly associated with higher inflammatory responses, including white blood cell count, neutrophil count and C-reactive protein. Also, white blood cell count, neutrophil count, C-reactive protein and the overall burden of comorbidity assessed by age-adjusted Charlson comorbidity index were independent risk factors for the Ct values. A mediation analysis detected the mediation effect of white blood cells on the association between the burden of comorbidity and Ct values, with the indirect effect estimates of 0.381 (95% CI: 0.166, 0.632, P < 0.001). Similarly, the indirect effect of C-reactive protein was -0.307 (95% CI: -0.645, -0.064, P = 0.034). White blood cells and C-reactive protein significantly mediated the relationship between the burden of comorbidity and Ct values by 29.56% and 18.13% of the total effect size, respectively. Conclusions: Inflammation mediated the association between the overall burden of comorbidity and Ct values among elderly with COVID-19, which suggests that combined immunomodulatory therapies could reduce the Ct values for such patients with a high burden of comorbidity.

Guidelines for the diagnosis, treatment, prevention and control of infections caused by carbapenem-resistant gram-negative bacilli.

The dissemination of carbapenem-resistant Gram-negative bacilli (CRGNB) is a global public health issue. CRGNB isolates are usually extensively drug-resistant or pandrug-resistant, resulting in limited antimicrobial treatment options and high mortality. A multidisciplinary guideline development group covering clinical infectious diseases, clinical microbiology, clinical pharmacology, infection control, and guideline methodology experts jointly developed the present clinical practice guidelines based on best available scientific evidence to address the clinical issues regarding laboratory testing, antimicrobial therapy, and prevention of CRGNB infections. This guideline focuses on carbapenem-resistant Enterobacteriales (CRE), carbapenem-resistant Acinetobacter baumannii (CRAB), and carbapenem-resistant Pseudomonas aeruginosa (CRPA). Sixteen clinical questions were proposed from the perspective of current clinical practice and translated into research questions using PICO (population, intervention, comparator, and outcomes) format to collect and synthesize relevant evidence to inform corresponding recommendations. The grading of recommendations, assessment, development and evaluation (GRADE) approach was used to evaluate the quality of evidence, benefit and risk profile of corresponding interventions and formulate recommendations or suggestions. Evidence extracted from systematic reviews and randomized controlled trials (RCTs) was considered preferentially for treatment-related clinical questions. Observational studies, non-controlled studies, and expert opinions were considered as supplementary evidence in the absence of RCTs. The strength of recommendations was classified as strong or conditional (weak). The evidence informing recommendations derives from studies worldwide, while the implementation suggestions combined the Chinese experience. The target audience of this guideline is clinician and related professionals involved in management of infectious diseases.

Guidelines for the diagnosis, treatment, prevention and control of infections caused by carbapenem-resistant gram-negative bacilli [Pré-print]

The dissemination of carbapenem-resistant Gram-negative bacilli (CRGNB) is a global public health issue. CRGNB isolates are usually extensively drug-resistant or pandrug-resistant, resulting in limited antimicrobial treatment options and high mortality. A multidisciplinary guideline development group covering clinical infectious diseases, clinical microbiology, clinical pharmacology, infection control, and guideline methodology experts jointly developed the present clinical practice guidelines based on best available scientific evidence to address the clinical issues regarding laboratory testing, antimicrobial therapy, and prevention of CRGNB infections. This guideline focuses on carbapenem-resistant Enterobacteriales (CRE), carbapenem-resistant Acinetobacter baumannii (CRAB), and carbapenem-resistant Pseudomonas aeruginosa (CRPA). Sixteen clinical questions were proposed from the perspective of current clinical practice and translated into research questions using PICO (population, intervention, comparator, and outcomes) format to collect and synthesize relevant evidence to inform corresponding recommendations. The grading of recommendations, assessment, development and evaluation (GRADE) approach was used to evaluate the quality of evidence, benefit and risk profile of corresponding interventions and formulate recommendations or suggestions. Evidence extracted from systematic reviews and randomized controlled trials (RCTs) was considered preferentially for treatment-related clinical questions. Observational studies, non-controlled studies, and expert opinions were considered as supplementary evidence in the absence of RCTs. The strength of recommendations was classified as strong or conditional (weak). The evidence informing recommendations derives from studies worldwide, while the implementation suggestions combined the Chinese experience. The target audience of this guideline is clinician and related professionals involved in management of infectious diseases.

Contamination of personal protective equipment and environmental surfaces in Fangcang shelter hospitals.

BACKGROUND: Fangcang shelter hospitals emerged as a new public health concept after COVID-19. Data regarding contamination of Fangcang shelter environments remains scarce. This study aims to investigate the extent of SARS-CoV-2 contamination on personal protective equipment and surfaces in Fangcang hospitals. METHODS: Between March and May 2022, during wave of omicron variant, a prospective study was conducted in 2 Fangcang hospitals in Shanghai, China. Swabs of personal protective equipment worn and environmental surfaces of contaminated areas, doffing rooms, and potentially contaminated areas were collected. SARS-CoV-2 RNA was detected by reverse transcription quantitative polymerase chain reaction. If viral RNA was detected, sampling was repeated after cleaning and disinfection. RESULTS: A total of 602 samples were collected. 13.3% of the personal protective equipment were contaminated. Positive rate was higher in the contaminated areas (48.4%) than in the doffing rooms (11.7%) and the potentially contaminated areas (0; P<.05). Contamination was highest in patient occupied areas (67.5%). After cleaning, samples taken at previously contaminated surfaces are all negative. CONCLUSIONS: SARS-CoV-2 RNA contamination is prevalent in Fangcang hospitals and healthcare workers are under risk of infection. Potentially contaminated areas and surfaces after cleaning and disinfection are negative, underlying the importance of infection control policy.

Evaluation of respiratory samples in etiology diagnosis and microbiome characterization by metagenomic sequencing.

BACKGROUND: The application of clinical mNGS for diagnosing respiratory infections improves etiology diagnosis, however at the same time, it brings new challenges as an unbiased sequencing method informing all identified microbiomes in the specimen. METHODS: Strategy evaluation and metagenomic analysis were performed for the mNGS data generated between March 2017 and October 2019. Diagnostic strengths of four specimen types were assessed to pinpoint the more appropriate type for mNGS diagnosis of respiratory infections. Microbiome complexity was revealed between patient cohorts and infection types. A bioinformatic pipeline resembling diagnosis results was built based upon multiple bioinformatic parameters. RESULTS: The positive predictive values (PPVs) for mNGS diagnosing of non-mycobacterium, Nontuberculous Mycobacteria (NTM), and Aspergillus were obviously higher in bronchoalveolar lavage fluid (BALF) demonstrating the potency of BALF in mNGS diagnosis. Lung tissues and sputum were acceptable for diagnosis of the Mycobacterium tuberculosis (MTB) infections. Interestingly, significant taxonomy differences were identified in sufficient BALF specimens, and unique bacteriome and virome compositions were found in the BALF specimens of tumor patients. Our pipeline showed comparative diagnostic strength with the clinical microbiological diagnosis. CONCLUSIONS: To achieve reliable mNGS diagnosis result, BALF specimens for suspicious common infections, and lung tissues and sputum for doubtful MTB infections are recommended to avoid the false results given by the complexed respiratory microbiomes. Our developed bioinformatic pipeline successful helps mNGS data interpretation and reduces manual corrections for etiology diagnosis.

Prolonged Use of Surgical Masks and Respirators Affects the Protection and Comfort for Healthcare Workers.

This study explored the ideal period for wearing masks to prevent the physiological and psychological problems associated with long-term face mask use during respiratory infections by healthcare workers. Breathing simulators, surgical masks (SM) and medical respirators (PM) were prepared for two to eight hours. Changes in the comfort of masks (facial skin temperature, breathing resistance, and moisture permeability) and protection (filtration efficiency, resistance to blood penetration, and colony count) were assessed. The results demonstrated that the masks offered efficient liquid-particle filtering even after eight hours of use. However, the number of bacterial colonies using PM and SM grew significantly after two and four hours, respectively. Concerning comfort, the inspiratory resistance of masks rose dramatically after two hours, whereas the moisture permeability declined considerably after four hours. In addition, skin temperature had a significant increase within two hours, which may result in facial discomfort. When conditions permitted, the hospital staff was instructed to replace their masks every two hours.

Diagnostic accuracy of metagenomic next-generation sequencing for cryptococcosis in immunocompetent and immunocompromised patients.

Objective: To compare the diagnostic accuracy of metagenomic next-generation sequencing (mNGS) for cryptococcosis in patients with different immune statuses with that of conventional detection. Methods: A total of 1442 specimens including 71 specimens from patients with cryptococcosis were analyzed in the study. The chi square test was used to screen the sensitivity and specificity of different detection methods for different specimen types. One-way ANOVA was used to compare the mNGS results with age, CD4, lymphocytes, IFN, IL-6, IL-2 and serum antigen assay. Results: The sensitivity of mNGS was 44.29% in Cryptococcus infection cases. The positive rate of mNGS results for bronchoalveolar lavage fluid (BALF, 87.50%) from immunocompromised patients was higher than that of BALF from immunocompetent patients (40.00%, p=0.04). The sensitivity of the serum Cryptococcus capsular antigen assay was 80.00% in immunocompetent patients and 96.42% in immunocompromised patients (p = 0.049). A positive rate of detection of Cryptococcus from mNGS was higher when cryptococcal antigen ≥1:160 (p=0.022) in immunocompromised patients. A positive rate of detection of Cryptococcus from mNGS was higher when lymphocyte counts were lower in both immunocompetent patients(p=0.017) and in immunocompromised patients(p=0.029). Conclusions: The sensitivity of mNGS is lower than that of serum cryptococcal antigen assay and histopathology in immunocompetent patients. However, BALF detection is recommend for immunocompromised patients compared with tissue and CSF. The positive mNGS result was correlated with lower lymphocyte counts, higher IL-2 and higher serum antigen assay in immunocompromised patients.

Characterization of Phage Resistance and Their Impacts on Bacterial Fitness in Pseudomonas aeruginosa.

The emergence and spread of antibiotic resistance pose serious environmental and health challenges. Attention has been drawn to phage therapy as an alternative approach to combat antibiotic resistance with immense potential. However, one of the obstacles to phage therapy is phage resistance, and it can be acquired through genetic mutations, followed by consequences of phenotypic variations. Therefore, understanding the mechanisms underlying phage-host interactions will provide us with greater detail on how to optimize phage therapy. In this study, three lytic phages (phipa2, phipa4, and phipa10) were isolated to investigate phage resistance and the potential fitness trade-offs in Pseudomonas aeruginosa. Specifically, in phage-resistant mutants phipa2-R and phipa4-R, mutations in conferring resistance occurred in genes pilT and pilB, both essential for type IV pili (T4P) biosynthesis. In the phage-resistant mutant phipa10-R, a large chromosomal deletion of ~294 kb, including the hmgA (homogentisate 1,2-dioxygenase) and galU (UTP-glucose-1-phosphate uridylyltransferase) genes, was observed and conferred phage phipa10 resistance. Further, we show examples of associated trade-offs in these phage-resistant mutations, e.g., impaired motility, reduced biofilm formation, and increased antibiotic susceptibility. Collectively, our study sheds light on resistance-mediated genetic mutations and their pleiotropic phenotypes, further emphasizing the impressive complexity and diversity of phage-host interactions and the challenges they pose when controlling bacterial diseases in this important pathogen. IMPORTANCE Battling phage resistance is one of the main challenges faced by phage therapy. To overcome this challenge, detailed information about the mechanisms of phage-host interactions is required to understand the bacterial evolutionary processes. In this study, we identified mutations in key steps of type IV pili (T4P) and O-antigen biosynthesis leading to phage resistance and provided new evidence on how phage predation contributed toward host phenotypes and fitness variations. Together, our results add further fundamental knowledge on phage-host interactions and how they regulate different aspects of Pseudomonas cell behaviors.

Incidence and outcomes of patients with COVID-19 associated pulmonary aspergillosis (CAPA) in intensive care units: a systematic review and meta-analysis of 31 cohort studies.

BACKGROUND: We aimed to identify studies systematically that describe the incidence and outcome of COVID-19-related pulmonary aspergillosis (CAPA). METHODS: We searched ScienceDirect, PubMed, CNKI, and MEDLINE (OVID) from December 31, 2019 to November 20, 2021 for all eligible studies. Random-model was used to reported the incidence, all-cause case fatality rate (CFR) and 95% confidence intervals (CIs). The meta-analysis was registered with PROSPERO (CRD42021242179). RESULTS: In all, thirty-one cohort studies were included in this study. A total of 3,441 patients with severe COVID-19 admitted to an intensive care unit (ICU) were investigated and 442 cases of CAPA were reported (30 studies). The pooled incidence rate of CAPA was 0.14 (95% CI: 0.11-0.17, I2=0.0%). Twenty-eight studies reported 287 deceased patients and 269 surviving patients. The pooled CFR of CAPA was 0.52 (95% CI: 0.47-0.56, I2=3.9%). Interestingly, patients with COVID19 would develop CAPA at 7.28 days after mechanical ventilation (range, 5.48-9.08 days). No significant publication bias was detected in this meta-analysis. DISCUSSION: Patients with COVID-19 admitted to an ICU might develop CAPA and have high all-cause CFR. We recommend conducting prospective screening for CAPA among patients with severe COVID-19, especially for those who receive mechanical ventilation over 7 days.

Evaluation of the EUCAST Rapid Antimicrobial Susceptibility Test for Enterobacterales-Containing Blood Cultures in China.

Bloodstream infection (BSI) is defined by the presence of microbes in the bloodstream and has high mortality. Early antimicrobial therapy is key to treating BSI patients. Because of potential antimicrobial resistance, rapid evaluation for the most suitable antimicrobial therapy is important for appropriate treatment. In China, the current workflow of microbiological diagnosis in BSI involves blood culture, species identification, and antimicrobial susceptibility testing, which takes around 3 days. However, this delay could lead to worse symptoms. To rapidly and accurately assess antimicrobial susceptibility, in this study, we applied EUCAST rapid antimicrobial susceptibility testing (RAST) to determine the antimicrobial susceptibilities of the most frequently detected Enterobacterales sampled in China, including Escherichia coli and Klebsiella pneumoniae. Based on EUCAST guidelines, we evaluated its efficiencies with six commercially available antimicrobials, including imipenem (10 µg), meropenem (10 µg), ciprofloxacin (5 µg), levofloxacin (5 µg), amikacin (30 µg), and trimethoprim-sulfamethoxazole (1.25/23.75 µg), with bacterium-spiked blood cultures. In addition, we developed potential breakpoints for a recently introduced antimicrobial, 30/20 µg ceftazidime-avibactam, which has high potential for treating multidrug-resistant Enterobacterales. Our results showed that EUCAST RAST is a reliable method for rapidly determining the antimicrobial susceptibilities of BSI-causing bacteria in China, with an overall categorical agreement rate at 8 h of ≥90%. The breakpoints developed in this study can categorize the isolates sampled in this study with an accuracy of 93%. Results from our experiments can be applied to clinically determine the microbial susceptibility of BSI-causing bacteria within 8 h and benefit clinical diagnostics for BSI patients.