Genomic typing and virulence gene profile analysis of Salmonella Derby from different sources.

Salmonella enterica serovar Derby (S. Derby) is one of the most common Salmonella serovars which can infect poultry, swine, and humans. With the reduction of the sequencing cost and the improvement of sequencing technology, whole genome sequencing (WGS) has become an important method for bacterial determination, molecular investigation, and pathogenic tracing analysis. In this study, we investigated S. Derby isolates from different sources in China using in-silico multilocus sequence typing (MLST), core genome MLST (cgMLST) and whole genome MLST (wgMLST) analysis based on WGS. The results showed that 21 S. Derby strains were divided into 3 STs using MLST analysis, including ST40 (n = 19, accounting for 90.48%), ST71 (n = 1, accounting for 4.76%) and ST8016 (n = 1, accounting for 4.76%). cgMLST and wgMLST analysis categorized the tested strains into 13 cgSTs and 21 wgSTs, respectively. The minimum spanning trees of cgMLST and wgMLST both divided these strains into 3 clusters and 4 singletons. In addition, virulence gene profiles of S. Derby isolates were also analyzed, and a total of 174 virulence genes belonged to 8 categories were identified. In summary, we studied genomic typing, phylogenetic relationship and virulence gene profiles of S. Derby strains from different sources in China. These findings were beneficial for the epidemiology and pathogenesis of Salmonella.

Biofilm-forming ability of Salmonella enterica strains of different serotypes isolated from multiple sources in China.

Salmonella is an important zoonotic and foodborne pathogen that can infect humans and animals, causing severe concerns about food safety and a heavy financial burden worldwide. The pathogen can adhere to living and abiotic surfaces by forming biofilms, which increases the risk of transmission and infection. In this study, we investigated the biofilm-forming ability of 243 Salmonella strains of 36 serotypes from different sources in China using microplate crystal violet staining method. The results showed that 99.6% tested strains, with the exception of one strain of S. Thompson, were capable of forming biofilms. The strains with the biofilm-forming ability of strong, medium and weak accounted for 2.88%, 24.28% and 72.43%, respectively. The strains of S. Havana and S. Hvittingfoss had the strongest biofilm-forming ability, with the OD570 of 0.81 ± 0.02 and 0.81 ± 0.38, respectively, while the strains of S. Agona and S. Bovismorbificans had the weakest biofilm-forming ability, with the OD570 of 0.16 ± 0.02 and 0.15 ± 0.00, respectively. Furthermore, statistical analysis results demonstrated that isolation of source had no effect on the biofilm formation ability, while the detection rates of pefABCD and ddhC were positively correlated with the biofilm formation ability of Salmonella. In particular, the detection rate of ddhC gene was more than 60% in the biofilm forming strains. These findings have important guiding significance for the investigation of pathogenesis, as well as the prevention and control of salmonellosis.

Optimization of fermentation, purification, and properties of blue pigment produced from Quambalaria cyanescens QY229.

AIM: Blue pigments have broad applications in foods, cosmetics, and clothing. However, natural blue pigments are rare. At present, the majority of blue pigments for sale are chemically synthetic. Owing to the safety risks of chemical pigments, it is an urgent demand to develop novel natural blue pigments. METHODS AND RESULTS: The fermentation medium and culture conditions of blue pigment produced by Quambalaria cyanescens QY229 were optimized by Plackett-Burman (PB) experimental design and response surface methodology (RSM) for the first time. The stability, bioactivity, and toxicity of the obtained blue pigment were studied after isolation and purification. CONCLUSION: The results showed that the optimal fermentation parameters were 34.61 g·L-1 of peptone concentration, 31.67°C of growing temperature, and 72.33 mL of medium volume in a 250-mL flask, and the yield of blue pigment reached 348.2 ± 7.1 U·mL-1. QY229 blue pigment is stable to light, heat, pH, most metal ions, and additives, and has certain antioxidant and inhibitory activity of α-glucosidase in vitro. QY229 blue pigment at concentrations of 0-1.25 mg·mL-1 was nontoxic to Caenorhabditis elegans in an acute toxicity trial.

Genomic virulence genes profile analysis of Salmonella enterica isolates from animal and human in China from 2004 to 2019.

Salmonella is a momentously zoonotic and food-borne pathogen that seriously threats human and animal health around the world. Salmonella pathogenicity is closely related to its virulence genes profile. However, conventional virulence gene analysis methods cannot truly reveal whole virulence genes carried by Salmonella. In this study, whole genome sequencing in combination with Virulence Factor Database were applied to investigate whole virulence gene profiles of 243 Salmonella isolates from animals and humans in China from 2004 to 2019. The results showed that a total of 670 virulence genes were identified in Salmonella, among them, 319 virulence genes were found in all the Salmonella tested isolates, and 9 virulence genes were unique to Salmonella. The 670 virulence genes were classified into 14 categories according to their functions, and the genes related to adherence, effector delivery system, immune modulation, motility and nutritional/metabolic factors accounted for 84.63%. Relationships between virulence genes and serovars, sequence types indicated that strains belonged to the same serovar or sequence type had similar virulence genes profiles, however, isolates from different sources, years and locations of isolation had variable virulence gene profiles. In addition, copy number of virulence genes and homologous virulence genes shared with other pathogens were also analyzed in this study. In summary, we investigated pan-genomic virulence gene profiles and molecular epidemiology of Salmonella isolates from humans and animals in China from 2004 to 2019. These findings are beneficial for pathogenic monitoring, investigation of virulence evolution as well as prevention and control of Salmonella.

[Assessment of prognostic and influencing factors in elderly patients with multiple organ dysfunction syndrome].

OBJECTIVE: To preliminarily assess the prognosis of patients with multiple organ dysfunction syndrome (MODSE) and analyze their influencing factors. METHODS: The clinical data of 365 MODSE patients admitted into Chinese PLA General Hospital during January 2009 to June 2012 were analyzed retrospectively. According to 28-day outcomes, they were divided into 2 groups (28-day survival and non-survival) while 4 groups according to age. Then these prognosis were evaluated with the current scoring systems ((acute physiology and chronic health evaluation II (APACHEII and III), sample acute physiological score (SAPSII) and multiple organ dysfunction score (MODS)). The predictive powers were compared by receiver operating characteristic (ROC) curve. Finally a binary Logistic regression analysis was performed to evaluate the relevant influencing prognostic factors of MODSE. RESULTS: The mean age was (77.8 ± 9.1) years, mean number of failed organs (3.6 ± 1.2) and a 28-day mortality 45.8%. The ages of non-survival group were older than those of survival group ((78.1 ± 9.1) vs (76.7 ± 11.0) years). The number of organ failures ((4.3 ± 1.1) vs (3.1 ± 1.0)) and scores (APACHEII: (28 ± 7) vs (20 ± 8), APACHE III: (106 ± 27) vs (75 ± 31), SAPSII: (64 ± 16) vs (46 ± 18), MODS: (9 ± 3) vs (6 ± 3)) of non-survivals were significantly higher than that of survivals. The area under ROC curve of these four score systems were 0.790, 0.781, 0.780 and 0.780 respectively. Compared to the above systems, SAPSII had the best performance in sensitivity while APACHEII was more valuable in specificity. All clinical data underwent binary Logistic regression and the results showed that plasma concentration of albumin and mean arterial pressure (MAP) offered beneficial outcomes while age and number of organ failures had unfavorable prognosis. The greater patient age, the higher their mortality. CONCLUSIONS: All four scoring systems have accurate prognostic predictions of MODSE patients. And the predictive power of APACHEII is the best. Plasma concentration of albumin, MAP, age and organ failure number are independent prognostic factors in MODSE patients.

Detection of Antibiotic Resistance and Analysis of Resistance Genes and Virulence Genes of Salmonella Isolated from Human.

Salmonella is a food-borne pathogen that can cause zoonoses. The emergence of drug-resistant strains of Salmonella is of great concern. It is necessary to understand the prevalence of antibiotic resistance, antibiotic resistance genes and virulence genes in human Salmonella. In this study, drug susceptibility test was used to detect and analyze the drug resistance of 24 Salmonella strains collected from human. A multi-drug resistant strain QLUF123 was selected for whole genome sequencing, and its drug resistance genes and virulence genes were analyzed. The results showed that 24 Salmonella strains were resistant to the tested antibiotics, 87.50% of the strains had multi-drug resistance, the resistance rate to ceftazidime, sulfamethoxazole and tilmicosin reached more than 80%. The alignment results based on the whole genome sequence showed that there were multiple types of drug-resistant genes in QLUF123, among which efflux pump system genes were the most abundant, including sdiA, mdtK, baeR and other multidrug-resistant efflux pump system genes. QLUF123 carried 46 kinds of virulence factors and 249 related virulence genes, among which the three functions of secretory system, adhesion and motility accounted for the most virulence genes, accounting for 93.57%. In this study, antibiotic resistance of human Salmonella was detected by drug sensitivity test, and drug resistance and virulence genes in Salmonella were analyzed by whole genome sequencing technology, which is of great significance for scientific treatment and rational drug use of related diseases caused by Salmonella infection.

Genomes-based MLST, cgMLST, wgMLST and SNP analysis of Salmonella Typhimurium from animals and humans.

Salmonella Typhimurium (S. Typhimurium) is an important food-borne and zoonotic pathogen that causes salmonellosis. With the development of whole genome sequencing (WGS), genome-based typing has been widely applied to bacteriology. In this study, we investigated genotyping and phylogenetic clusters of S. Typhimurium isolates from humans and animals in different provinces (including Beijing, Shandong, Guangxi, Shaanxi, Henan, and Shanghai) of China during 2009-2018 using multi locus sequence typing (MLST), core genome MLST (cgMLST), whole genome MLST (wgMLST) and single nucleotide polymorphism (SNP) based on WGS. 29 S. Typhimurium isolates from chicken (n = 22), sick pigeon (n = 2), patients (n = 4) and sick swine (n = 1) were tested. MLST analysis showed S. Typhimurium strains were divided into four STs, namely ST19 (n = 14), ST34 (n = 12), ST128 (n = 2) and ST1544 (n = 1). cgMLST and wgMLST divided 29 strains into 27 cgSTs and 29 wgST, respectively. Phylogenetic clustering showed that all isolates were divided into 4 clusters and 4 singletons. SNP analysis was used to examine MLST, cgMLST, wgMLST analysis. Finally, comparisons of MLST, cgMLST, wgMLST, and SNP were analyzed and the results showed their precision increased in order. In summary, genomic typing and phylogenetic relationships of 29 S. Typhimurium strains from different sources in China were analyzed. These findings were beneficial to investigate molecular pathogenesis, bacterial diversity, and traceability analysis of Salmonella.

Value of soluble TREM-1, procalcitonin, and C-reactive protein serum levels as biomarkers for detecting bacteremia among sepsis patients with new fever in intensive care units: a prospective cohort study.

BACKGROUND: The purpose of this study was to explore the diagnostic value of soluble triggering receptor expressed on myeloid cells 1 (sTREM-1), procalcitonin (PCT), and C-reactive protein (CRP) serum levels for differentiating sepsis from SIRS, identifying new fever caused by bacteremia, and assessing prognosis when new fever occurred. METHODS: We enrolled 144 intensive care unit (ICU) patients: 60 with systemic inflammatory response syndrome (SIRS) and 84 with sepsis complicated by new fever at more than 48 h after ICU admission. Serum sTREM-1, PCT, and CRP levels were measured on the day of admission and at the occurrence of new fever (>38.3°C) during hospitalization. Based on the blood culture results, the patients were divided into a blood culture-positive bacteremia group (33 patients) and blood culture-negative group (51 patients). Based on 28-day survival, all patients, both blood culture-positive and -negative, were further divided into survivor and nonsurvivor groups. RESULTS: On ICU day 1, the sepsis group had higher serum sTREM-1, PCT, and CRP levels compared with the SIRS group (P <0.05). The areas under the curve (AUC) for these indicators were 0.868 (95% CI, 0.798-0.938), 0.729 (95% CI, 0.637-0.821), and 0.679 (95% CI, 0.578-0.771), respectively. With 108.9 pg/ml as the cut-off point for serum sTREM-1, sensitivity was 0.83 and specificity was 0.81. There was no statistically significant difference in serum sTREM-1 or PCT levels between the blood culture-positive and -negative bacteremia groups with ICU-acquired new fever. However, the nonsurvivors in the blood culture-positive bacteremia group had higher levels of serum sTREM-1 and PCT (P <0.05), with a prognostic AUC for serum sTREM-1 of 0.868 (95% CI, 0.740-0.997). CONCLUSIONS: Serum sTREM-1, PCT, and CRP levels each have a role in the early diagnosis of sepsis. Serum sTREM-1, with the highest sensitivity and specificity of all indicators studied, is especially notable. sTREM-1, PCT, and CRP levels are of no use in determining new fever caused by bacteremia in ICU patients, but sTREM-1 levels reflect the prognosis of bacteremia. TRIAL REGISTRATION: ClinicalTrial.gov identifier NCT01410578.

Diagnostic value of urine sCD163 levels for sepsis and relevant acute kidney injury: a prospective study.

BACKGROUND: Sepsis is a common syndrome in critically ill patients and easily leads to the occurrence of acute kidney injury (AKI), with high mortality rates. This study aimed to investigate the diagnostic value of urine soluble CD163 (sCD163) for identification of sepsis, severity of sepsis, and for secondary AKI, and to assess the patients' prognosis. METHODS: We enrolled 20 cases with systemic inflammatory response syndrome (SIRS), 40 cases with sepsis (further divided into 17 sepsis cases and 23 severe sepsis cases) admitted to the intensive care unit (ICU), and 20 control cases. Results for urine sCD163 were recorded on the day of admission to the ICU, and AKI occurrence was noted. RESULTS: On the day of ICU admission, the sepsis group exhibited higher levels of urine sCD163 (74.8 ng/ml; range: 47.9-148.3 ng/ml) compared with those in the SIRS group (31.9 ng/ml; 16.8-48.0, P < 0.001). The area under the curve (AUC) was 0.83 (95% confidence interval [CI]: 0.72-0.94, P < 0.001) the sensitivity was 0.83, and the specificity was 0.75 (based on a cut-off point of 43.0 ng/ml). Moreover, the severe sepsis group appeared to have a higher level of sCD163 compared with that in the sepsis group (76.2; 47.2-167.5 ng/ml vs. 74.2; 46.2-131.6 ng/ml), but this was not significant. For 15 patients with AKI, urine sCD163 levels at AKI diagnosis were significantly higher than those of the remaining 35 sepsis patients upon ICU admission (121.0; 74.6-299.1 ng/ml vs. 61.8; 42.8-128.3 ng/ml, P = 0.049). The AUC for urine sCD163 was 0.688 (95% CI: 0.51-0.87, P = 0.049). Sepsis patients with a poor prognosis showed a higher urine sCD163 level at ICU admission (98.6; 50.3-275.6 ng/ml vs. 68.0; 44.8-114.5 ng/ml), but this was not significant. Patients with AKI with a poor prognosis had higher sCD163 levels than those in patients with a better prognosis (205.9; 38.6-766.0 ng/ml vs. 80.9; 74.9-141.0 ng/ml), but this was not significant. CONCLUSIONS: This study shows, for the first time, the potential value of urine sCD163 levels for identifying sepsis and diagnosing AKI, as well as for assessment of patients' prognosis. TRIAL REGISTRATION: ChiCTR-ONC-10000812.

Prognosis and weaning of elderly multiple organ dysfunction syndrome patients with invasive mechanical ventilation.

BACKGROUND: Elderly multiple organ dysfunction syndrome (MODS) patients receiving invasive mechanical ventilation have poor prognosis in intensive care units (ICUs). We studied the usefulness of four commonly used severity scores and extrapulmonary factors that affected weaning to predict outcome of such patients. METHODS: Clinical data of 197 patients on admission to ICUs (from January 2009 to June 2012) were used retrospectively. The Acute Physiology and Chronic Health Evaluation (APACHE) II, APACHE III, Sample Acute Physiological Score (SAPS) II and MODS scores were calculated. All the patients were grouped into survivors and nonsurvivors according to the prognosis. Patients, who weaned from ventilator (n = 154), were subdivided into a successful weaning group and a failed weaning group. The receiver operating characteristic (ROC) curves and Logistic regression was used for prognostic and weaning assessment. RESULTS: Based on the outcomes, the areas under the ROC of APACHE II, APACHE III, SAPS II, and MODS were 0.837, 0.833, 0.824, and 0.837, respectively. The Logistic regression analysis revealed that the odds ratio (OR) of underlying lung diseases, serum albumin and creatinine, and the number of organ failures was 2.374, 0.920, 1.003, and 1.547. APACHE II scores on admission performed excellent (ROC: 0.921) on the weaning assessments. CONCLUSIONS: APACHE II and MODS systems were marginally better for evaluating the prognosis of elderly MODS patients who received invasive mechanical ventilation. Underlying lung diseases, serum albumin, serum creatinine and the number of organ failures were independent prognostic factors. Using the APACHE II scores on admission before weaning may increase the likelihood of successful weaning. (ClinicalTrial.gov identifier NCT01802983).

Identification of novel biomarkers for sepsis prognosis via urinary proteomic analysis using iTRAQ labeling and 2D-LC-MS/MS.

OBJECTIVES: Sepsis is the major cause of death for critically ill patients. Recent progress in proteomics permits a thorough characterization of the mechanisms associated with critical illness. The purpose of this study was to screen potential biomarkers for early prognostic assessment of patients with sepsis. METHODS: For the discovery stage, 30 sepsis patients with different prognoses were selected. Urinary proteins were identified using isobaric tags for relative and absolute quantitation (iTRAQ) coupled with LC-MS/MS. Mass spec instrument analysis were performed with Mascot software and the International Protein Index (IPI); bioinformatic analyses were used by the algorithm of set and the Gene Ontology (GO) Database. For the verification stage, the study involved another 54 sepsis-hospitalized patients, with equal numbers of patients in survivor and non-survivor groups based on 28-day survival. Differentially expressed proteins were verified by Western Blot. RESULTS: A total of 232 unique proteins were identified. Proteins that were differentially expressed were further analyzed based on the pathophysiology of sepsis and biomathematics. For sepsis prognosis, five proteins were significantly up-regulated: selenium binding protein-1, heparan sulfate proteoglycan-2, alpha-1-B glycoprotein, haptoglobin, and lipocalin; two proteins were significantly down-regulated: lysosome-associated membrane proteins-1 and dipeptidyl peptidase-4. Based on gene ontology clustering, these proteins were associated with the biological processes of lipid homeostasis, cartilage development, iron ion transport, and certain metabolic processes. Urinary LAMP-1 was down-regulated, consistent with the Western Blot validation. CONCLUSION: This study provides the proteomic analysis of urine to identify prognostic biomarkers of sepsis. The seven identified proteins provide insight into the mechanism of sepsis. Low urinary LAMP-1 levels may be useful for early prognostic assessment of sepsis. TRIAL REGISTRATION: ClinicalTrial.gov NCT01493492.

Significance of low serum vitamin D for infection risk, disease severity and mortality in critically ill patients.

BACKGROUND: Hospitalized patients often have higher rate of vitamin D deficiency than healthy people. Vitamin D levels below normal are associated with hospital stay, increased incidence of adverse prognosis and increased mortality of a number of diseases. Whether there is a relationship between vitamin D levels and infection or sepsis in the critically ill is still unclear. This study will explore the relationship between vitamin D levels and risk of infection, assessment for disease severity, and predictor of mortality. METHODS: To evaluate the value of vitamin D in intensive care unit (ICU) cases to sepsis, severity and prognosis assessment, high performance liquid chromatography and tandem mass spectrometry were used to measure the concentrations of vitamin D in sera of critically ill patients. The serum samples were drawn within the first 24 hours of ICU admission. RESULTS: The study included 206 people, 50 healthy controls, 51 ICU control patients and 105 ICU diagnosed with sepsis. Critically ill ICU patients (ICU sepsis and ICU control group) had lower vitamin D concentration than normal people, but septic patients showed no significant reduction of vitamin D concentration when compared with critically ill patients with no positive etiological evidence. For assessment of disease severity, there were very low negative correlations between APACHE II, SAPS II and SOFA scores and vitamin D level. Additionally, patients of different 25-(OH)D levels showed no difference whether in terms of 28-day survival (X(2) = 1.78, P = 0.776) or 90-day survival (X(2) = 4.12, P = 0.389). Multivariate Logistic regression demonstrated that APECHE II and SAPS II scores were independent risk factors to deaths caused by sepsis. CONCLUSION: Clinically, serum concentration of vitamin D is not an indicator for diagnosis and assessment in critically ill patients (ClinicalTrial.gov identifier NCT01636232).

Dynamic changes in serum soluble triggering receptor expressed on myeloid cells-1 (sTREM-1) and its gene polymorphisms are associated with sepsis prognosis.

This study explored the association of sepsis prognosis with dynamic changes in serum soluble triggering receptor expressed on myeloid cells-1 (sTREM-1) and its polymorphisms. We enrolled 80 subjects with sepsis and 80 controls. Serum sTREM-1 was tested on days 1, 3, 5, 7, 10, and 14. PCR sequencing was performed to detect TREM-1 genetic variation on its four exons. sTREM-1 levels were significantly higher in the nonsurvivor than in the survivor group (p < 0.001), and those at each time point were the same (p ≤ 0.001). Of the three tested TREM-1 SNPs (rs144672509, rs2234237, and rs2234246), only rs2234237 (Ser25Thr) was significantly associated with sepsis prognosis in three inheritance models (p < 0.05). However, there was no relationship between TREM-1 polymorphism and dynamic concentration of sTREM-1. Logistic regression showed that sTREM-1, APACHE II, and rs2234237 polymorphism are risk factors for prognosis. Dynamic changes in serum sTREM-1 and rs2234237 polymorphism could be used in sepsis prognosis assessment.