Tandem gene duplications contributed to high-level azole resistance in a rapidly expanding Candida tropicalis population.

Invasive diseases caused by the globally distributed commensal yeast Candida tropicalis are associated with mortality rates of greater than 50%. Notable increases of azole resistance have been observed in this species, particularly within Asia-Pacific regions. Here, we carried out a genetic population study on 1571 global C. tropicalis isolates using multilocus sequence typing (MLST). In addition, whole-genome sequencing (WGS) analysis was conducted on 629 of these strains, comprising 448 clinical invasive strains obtained in this study and 181 genomes sourced from public databases. We found that MLST clade 4 is the predominant azole-resistant clone. WGS analyses demonstrated that dramatically increasing rates of azole resistance are associated with a rapid expansion of cluster AZR, a sublineage of clade 4. Cluster AZR isolates exhibited a distinct high-level azole resistance, which was induced by tandem duplications of the ERG11A395T gene allele. Ty3/gypsy-like retrotransposons were found to be highly enriched in this population. The alarming expansion of C. tropicalis cluster AZR population underscores the urgent need for strategies against growing threats of antifungal resistance.

Emergence of Antifungal Resistant Subclades in the Global Predominant Phylogenetic Population of Candida albicans.

Candida albicans remains the most common species causing invasive candidiasis. In this study, we present the population structure of 551 global C. albicans strains. Of these, the antifungal susceptibilities of 370 strains were tested. Specifically, 66.6% of the azole-nonsusceptible (NS)/non-wild-type (NWT) strains that were tested belonged to Clade 1. A phylogenetic analysis, a principal components analysis, the population structure, and a loss of heterozygosity events revealed two nested subclades in Clade 1, namely, Clade 1-R and Clade 1-R-α, that exhibited higher azole-NS/NWT rates (75.0% and 100%, respectively). In contrast, 6.4% (21/326) of the non-Clade 1-R isolates were NS/NWT to at least 1 of 4 azoles. Notably, all of the Clade 1-R-α isolates were pan-azole-NS/NWT that carried unique A114S and Y257H double substitutions in Erg11p and had the overexpression of ABC-type efflux pumps introduced by the substitution A736V in transcript factor Tac1p. It is worth noting that the Clade 1-R and Clade 1-R-α isolates were from different cities that are distributed over a large geographic span. Our study demonstrated the presence of specific phylogenetic subclades that are associated with antifungal resistance among C. albicans Clade 1, which calls for public attention on the monitoring of the future spread of these clones. IMPORTANCE Invasive candidiasis is the most common human fungal disease among hospitalized patients, and Candida albicans is the predominant pathogen. Considering the large number of infected cases and the limited alternative therapies, the azole-resistance of C. albicans brings a huge clinical threat. Here, our study suggested that antifungal resistance in C. albicans could also be associated with phylogenetic lineages. Specifically, it was revealed that more than half of the azole-resistant C. albicans strains belonged to the same clade. Furthermore, two nested subclades of the clade exhibited extremely high azole-resistance. It is worth noting that the isolates of two subclades were from different cities that are distributed over a large geographic span in China. This indicates that the azole-resistant C. albicans subclades may develop into serious public health concerns.

PgtE Enzyme of Salmonella enterica Shares the Similar Biological Roles to Plasminogen Activator (Pla) in Interacting With DEC-205 (CD205), and Enhancing Host Dissemination and Infectivity by Yersinia pestis.

Yersinia pestis, the cause of plague, is a newly evolved Gram-negative bacterium. Through the acquisition of the plasminogen activator (Pla), Y. pestis gained the means to rapidly disseminate throughout its mammalian hosts. It was suggested that Y. pestis utilizes Pla to interact with the DEC-205 (CD205) receptor on antigen-presenting cells (APCs) to initiate host dissemination and infection. However, the evolutionary origin of Pla has not been fully elucidated. The PgtE enzyme of Salmonella enterica, involved in host dissemination, shows sequence similarity with the Y. pestis Pla. In this study, we demonstrated that both Escherichia coli K-12 and Y. pestis bacteria expressing the PgtE-protein were able to interact with primary alveolar macrophages and DEC-205-transfected CHO cells. The interaction between PgtE-expressing bacteria and DEC-205-expressing transfectants could be inhibited by the application of an anti-DEC-205 antibody. Moreover, PgtE-expressing Y. pestis partially re-gained the ability to promote host dissemination and infection. In conclusion, the DEC-205-PgtE interaction plays a role in promoting the dissemination and infection of Y. pestis, suggesting that Pla and the PgtE of S. enterica might share a common evolutionary origin.

Anti-SARS-CoV-2 IgG responses are powerful predicting signatures for the outcome of COVID-19 patients.

Introduction: The COVID-19 global pandemic is far from ending. There is an urgent need to identify applicable biomarkers for early predicting the outcome of COVID-19. Growing evidences have revealed that SARS-CoV-2 specific antibodies evolved with disease progression and severity in COIVD-19 patients. Objectives: We assumed that antibodies may serve as biomarkers for predicting the clinical outcome of hospitalized COVID-19 patients on admission. Methods: By taking advantage of a newly developed SARS-CoV-2 proteome microarray, we surveyed IgG responses against 20 proteins of SARS-CoV-2 in 1034 hospitalized COVID-19 patients on admission and followed till 66 days. The microarray results were further correlated with clinical information, laboratory test results and patient outcomes. Cox proportional hazards model was used to explore the association between SARS-CoV-2 specific antibodies and COVID-19 mortality. Results: Nonsurvivors (n = 955) induced higher levels of IgG responses against most of non-structural proteins than survivors (n = 79) on admission. In particular, the magnitude of IgG antibodies against 8 non-structural proteins (NSP1, NSP4, NSP7, NSP8, NSP9, NSP10, RdRp, and NSP14) and 2 accessory proteins (ORF3b and ORF9b) possessed significant predictive power for patient death, even after further adjustments for demographics, comorbidities, and common laboratory biomarkers for disease severity (all with p trend < 0.05). Additionally, IgG responses to all of these 10 non-structural/accessory proteins were also associated with the severity of disease, and differential kinetics and serum positive rate of these IgG responses were confirmed in COVID-19 patients of varying severities within 20 days after symptoms onset. The area under curves (AUCs) for these IgG responses, determined by computational cross-validations, were between 0.62 and 0.71. Conclusions: Our findings might have important implications for improving clinical management of COVID-19 patients.

Antifungal Susceptibility Profiles and Resistance Mechanisms of Clinical Diutina catenulata Isolates With High MIC Values.

Diutina catenulata (Candida catenulata) is an ascomycete yeast species widely used in environmental and industrial research and capable of causing infections in humans and animals. At present, there are only a few studies on D. catenulata, and further research is required for its more in-depth characterization and analysis. Eleven strains of D. catenulata collected from China Hospital Invasive Fungal Surveillance Net (CHIF-NET) and the CHIF-NET North China Program were identified using matrix-assisted laser desorption ionization-time of flight mass spectrometry and internal transcribed spacer sequencing. The antifungal susceptibility of the Diutina catenulata strains was tested using the Clinical and Laboratory Standards Institute broth microdilution method and Sensititre YeastOne™. Furthermore, ERG11 and FKS1 were sequenced to determine any mutations related to azole and echinocandin resistance in D. catenulata. All isolates exhibited low minimum inhibitory concentration (MIC) values for itraconazole (0.06-0.12 µg/ml), posaconazole (0.06-0.12 µg/ml), amphotericin B (0.25-1 µg/ml), and 5-flucytosine (range, <0.06-0.12 µg/ml), whereas four isolates showed high MICs (≥4 µg/ml) for echinocandins. Strains with high MIC values for azoles showed common ERG11 mutations, namely, F126L/K143R. In addition, L139R mutations may be linked to high MICs of fluconazole. Two amino acid alterations reported to correspond to high MIC values of echinocandin, namely, F621I (F641) and S625L (S645), were found in the hot spot 1 region of FKS1. In addition, one new amino acid alteration, I1348S (I1368), was found outside of the FKS1 hot spot 2 region, and its contribution to echinocandin resistance requires future investigation. Diutina catenulata mainly infects patients with a weak immune system, and the high MIC values for various antifungals exhibited by these isolates may represent a challenge to clinical treatment.

Loss of the virulence plasmid by Shigella sonnei promotes its interactions with CD207 and CD209 receptors.

Introduction. Shigella sonnei, the cause of bacillary dysentery, belongs to Gram-negative enteropathogenic bacteria. S. sonnei contains a 210 kb virulence plasmid that encodes an O-antigen gene cluster of LPSs. However, this virulence plasmid is frequently lost during replication. It is well-documented that after losing the O-antigen and becoming rough strains, the Gram-negative bacteria may express an LPS core on its surface. Previous studies have suggested that by using the LPS core, Gram-negative bacteria can interact with several C-type lectin receptors that are expressed on antigen-presenting cells (APCs).Hypothesis/Gap Statement. S. sonnei by losing the virulence plasmid may hijack APCs via the interactions of LPS-CD209/CD207.Aim. This study aimed to investigate if the S. sonnei rough strain, by losing the virulence plasmid, interacted with APCs that express C-type lectins of human CD207, human CD209a and mouse CD209b.Methodology. SDS-PAGE silver staining was used to examine the O-antigen expression of S. sonnei WT and its rough strain. Invasion assays and inhibition assays were used to examine the ability of S. sonnei WT and its rough strain to invade APCs and investigate whether CD209 and CD207 are receptors for phagocytosis of rough S. sonnei. Animal assays were used to observe the dissemination of S. sonnei.Results. S. sonnei did not express O-antigens after losing the virulence plasmid. The S. sonnei rough strain invades with APCs, including human dendritic cells (DCs) and mouse macrophages. CD209 and CD207 are receptors for phagocytosis of rough S. sonnei. Expression of the O-antigen reduces the ability of the S. sonnei rough strain to be disseminated to mesenteric lymph nodes and spleens.Conclusion. This work demonstrated that S. sonnei rough strains - by losing the virulence plasmid - invaded APCs through interactions with CD209 and CD207 receptors.

Antibody dynamics to SARS-CoV-2 in asymptomatic COVID-19 infections.

BACKGROUND: The missing asymptomatic COVID-19 infections have been overlooked because of the imperfect sensitivity of the nucleic acid testing (NAT). Globally understanding the humoral immunity in asymptomatic carriers will provide scientific knowledge for developing serological tests, improving early identification, and implementing more rational control strategies against the pandemic. MEASURE: Utilizing both NAT and commercial kits for serum IgM and IgG antibodies, we extensively screened 11 766 epidemiologically suspected individuals on enrollment and 63 asymptomatic individuals were detected and recruited. Sixty-three healthy individuals and 51 mild patients without any preexisting conditions were set as controls. Serum IgM and IgG profiles were further probed using a SARS-CoV-2 proteome microarray, and neutralizing antibody was detected by a pseudotyped virus neutralization assay system. The dynamics of antibodies were analyzed with exposure time or symptoms onset. RESULTS: A combination test of NAT and serological testing for IgM antibody discovered 55.5% of the total of 63 asymptomatic infections, which significantly raises the detection sensitivity when compared with the NAT alone (19%). Serum proteome microarray analysis demonstrated that asymptomatics mainly produced IgM and IgG antibodies against S1 and N proteins out of 20 proteins of SARS-CoV-2. Different from strong and persistent N-specific antibodies, S1-specific IgM responses, which evolved in asymptomatic individuals as early as the seventh day after exposure, peaked on days from 17 days to 25 days, and then disappeared in two months, might be used as an early diagnostic biomarker. 11.8% (6/51) mild patients and 38.1% (24/63) asymptomatic individuals did not produce neutralizing antibody. In particular, neutralizing antibody in asymptomatics gradually vanished in two months. CONCLUSION: Our findings might have important implications for the definition of asymptomatic COVID-19 infections, diagnosis, serological survey, public health, and immunization strategies.

Distinct effects of asthma and COPD comorbidity on disease expression and outcome in patients with COVID-19.

BACKGROUND: The impacts of chronic airway diseases on coronavirus disease 2019 (COVID-19) are far from understood. OBJECTIVE: To explore the influence of asthma and chronic obstructive pulmonary disease (COPD) comorbidity on disease expression and outcomes, and the potential underlying mechanisms in COVID-19 patients. METHODS: A total of 961 hospitalized COVID-19 patients with a definite clinical outcome (death or discharge) were retrospectively enrolled. Demographic and clinical information were extracted from the medical records. Lung tissue sections from patients suffering from lung cancer were used for immunohistochemistry study of angiotensin-converting enzyme II (ACE2) expression. BEAS-2B cell line was stimulated with various cytokines. RESULTS: In this cohort, 21 subjects (2.2%) had COPD and 22 (2.3%) had asthma. After adjusting for confounding factors, COPD patients had higher risk of developing severe illness (OR: 23.433; 95% CI 1.525-360.135; P < .01) and acute respiratory distress syndrome (OR: 19.762; 95% CI 1.461-267.369; P = .025) than asthmatics. COPD patients, particularly those with severe COVID-19, had lower counts of CD4+ T and CD8+ T cells and B cells and higher levels of TNF-α, IL-2 receptor, IL-10, IL-8, and IL-6 than asthmatics. COPD patients had increased, whereas asthmatics had decreased ACE2 protein expression in lower airways, compared with that in control subjects without asthma and COPD. IL-4 and IL-13 downregulated, but TNF-α, IL-12, and IL-17A upregulated ACE2 expression in BEAS-2B cells. CONCLUSION: Patients with asthma and COPD likely have different risk of severe COVID-19, which may be associated with different ACE2 expression.

A national survey on fungal infection diagnostic capacity in the clinical mycology laboratories of tertiary care hospitals in China.

BACKGROUND/PURPOSE: As the incidence of fungal infections in China increases, the demand for rapid and accurate diagnosis of mycoses is growing. Yet, information on current diagnostic capacity is scarce. METHODS: An online survey was conducted in February 2018 to collect information on mycology testing from tertiary care hospitals across China. Responses from 348 hospitals were analyzed, and a scoring system was designed and employed to assess the overall diagnostic capacity. RESULTS: Most of the surveyed hospitals did not have separate laboratory space, manpower, or equipment dedicated for fungal testing. Conventional staining methods were widely available (>70%), whereas GMS and fluorescent staining were less common. Fungal identification services were offered mostly with chromogenic medium, morphological characterization or automated identification systems, other than more advanced methods such as MALDI-TOF MS and DNA sequencing. Fungal serology testing was available in 81.1%, with G test being the most often used. Though 91.8% of the respondents had the ability to perform antifungal susceptibility testing for yeasts, less than 13% conducted such testing for molds. The percentage of laboratories participating in External Quality Assessment programs and research was 57.5% and 32.5%, respectively. The average score for the 348 surveyed hospitals was 37.2 (out of a maximum of 89 points), with only 15 hospitals scoring >60, suggesting a general lack of high-quality mycology laboratories. CONCLUSIONS: The overall clinical testing capacity for fungal infection in China is insufficient. More investment and training efforts are warranted to establish centers of excellence and promote access to high-quality diagnostic services.