In vivo functional immunoprotection correlates for vaccines against invasive bacteria.

Vaccination has significantly reduced the incidence of invasive infections caused by several bacterial pathogens, including Streptococcus pneumoniae, Haemophilus influenzae, and Neisseria meningitidis. However, no vaccines are available for many other invasive pathogens. A major hurdle in vaccine development is the lack of functional markers to quantify vaccine immunity in eliminating pathogens during the process of infection. Based on our recent discovery of the liver as the major organ of vaccine-induced clearance of blood-borne virulent bacteria, we here describe a new vaccine evaluation system that quantitatively characterizes the key features of effective vaccines in shuffling virulent bacteria from the blood circulation to the liver resident macrophage Kupffer cells (KCs) and sinusoidal endothelial cells (LSECs) in mouse septic infection model. This system consists of three related correlates or assays: pathogen clearance from the bloodstream, pathogen trapping in the liver, and pathogen capture by KCs/LSECs. These readouts were consistently associated with the serotype-specific immunoprotection levels of the 13-valent pneumococcal polysaccharide conjugate vaccine (PCV13) against lethal infection of S. pneumoniae, a major invasive Gram-positive pathogen of community-acquired infections in humans. Furthermore, the reliability and sensitivity of these correlates in reflecting vaccine efficacy were verified with whole cell vaccines of Klebsiella pneumoniae and Escherichia coli, two major Gram-negative pathogens in hospital-acquired invasive infections. This system may be used as effective readouts to evaluate the immunoprotective potential of vaccine candidates in the preclinical phase by filling the current technical gap in vaccine evaluation between the conventional in vitro approaches (e.g. antibody production and pathogen neutralization/opsonophagocytosis) and survival of immunized animals.

Microbiological characteristics of bloodstream infections in a reference hospital in northeastern Brazil

Abstract Routine blood culture is used for the detection of bloodstream infections by aerobic and anaerobic bacteria and by common pathogenic yeasts. A retrospective study was conducted in a public hospital in Maceió-AL, by collecting data of all medical records with positive blood cultures. Out of the 2,107 blood cultures performed, 17% were positive with Staphylococcus coagulase negative (51.14%), followed by Staphylococcus aureus (11.21%) and Klebsiella pneumoniae (6.32%). Gram-positive bacteria predominated among positive blood cultures, highlighting the group of Staphylococcus coagulase-negative. While Gram-negative bacteria had a higher number of species among positive blood cultures.

Resumo A cultura sanguínea de rotina é usada para a detecção de infecções na corrente sanguínea por bactérias aeróbias e anaeróbias e por leveduras patogênicas comuns. Estudo retrospectivo realizado em hospital público de Maceió-AL, por meio da coleta de dados de todos os prontuários com culturas sanguíneas positivas. Das 2.107 culturas sanguíneas realizadas, 17% foram positivas com Staphylococcus coagulase negativo (51,14%), seguido por Staphylococcus aureus (11,21%) e Klebsiella pneumoniae (6,32%). As bactérias Gram-positiva predominaram entre as culturas de sangue positivas, destacando-se o grupo das Staphylococcus coagulase-negativo. Enquanto as bactérias Gram-negativas apresentaram um número maior de espécies entre as culturas de sangue positivas.

Microbiological characteristics of bloodstream infections in a reference hospital in northeastern Brazil / Características microbiológicas de infecções da corrente sanguínea em hospital de referência no nordeste do Brasil

Abstract Routine blood culture is used for the detection of bloodstream infections by aerobic and anaerobic bacteria and by common pathogenic yeasts. A retrospective study was conducted in a public hospital in Maceió-AL, by collecting data of all medical records with positive blood cultures. Out of the 2,107 blood cultures performed, 17% were positive with Staphylococcus coagulase negative (51.14%), followed by Staphylococcus aureus (11.21%) and Klebsiella pneumoniae (6.32%). Gram-positive bacteria predominated among positive blood cultures, highlighting the group of Staphylococcus coagulase-negative. While Gram-negative bacteria had a higher number of species among positive blood cultures.

Resumo A cultura sanguínea de rotina é usada para a detecção de infecções na corrente sanguínea por bactérias aeróbias e anaeróbias e por leveduras patogênicas comuns. Estudo retrospectivo realizado em hospital público de Maceió-AL, por meio da coleta de dados de todos os prontuários com culturas sanguíneas positivas. Das 2.107 culturas sanguíneas realizadas, 17% foram positivas com Staphylococcus coagulase negativo (51,14%), seguido por Staphylococcus aureus (11,21%) e Klebsiella pneumoniae (6,32%). As bactérias Gram-positiva predominaram entre as culturas de sangue positivas, destacando-se o grupo das Staphylococcus coagulase-negativo. Enquanto as bactérias Gram-negativas apresentaram um número maior de espécies entre as culturas de sangue positivas.

Clinical Diagnosis Application of Metagenomic Next-Generation Sequencing of Plasma in Suspected Sepsis.

Purpose: We analyzed the clinical concordance of mNGS test results from blood samples and improved the clinical efficiency of mNGS in the diagnosis of suspected sepsis pathogens. Patients and Methods: In this study, 99 samples of suspected blood flow infection were included for plasma mNGS, and the correlation between mNGS results and blood culture results, serum inflammatory indices, clinical symptoms and antibiotic treatment was analyzed, as well as the comparison with the detection rate of BALF pathogens, as well as the classification of different pathogens in the mNGS results were analyzed. Results: The mNGS pathogen detection rate was higher than that of traditional blood culture (83.02% vs 35.82%). The rate of the mNGS results being consistent with the clinical diagnosis was also higher than that of traditional blood culture (58.49% vs 20.75%). This study shows that bacteria and fungi are the main pathogens in sepsis, and viral sepsis is very rare. In this study, 32% of sepsis patients were secondary to pneumonia. Compared with the pathogen detection rate using alveolar lavage fluid, the detection rate from plasma mNGS was 62.5%. Samples were also easy to sample, noninvasive, and more convenient for clinical application. Conclusion: This study shows that compared with blood culture, the detection rate of mNGS pathogen that meets the diagnosis of sepsis is higher. We need a combination of multiple indicators to monitor the early diagnosis and treatment of sepsis.

Influencing factors of blood infection and death risk of Acinetobacter baumannii in elderly patients / 公共卫生与预防医学

Objective To investigate the death prognosis and risk factors of extensively drug-resistant Acinetobacter baumannii in hospitalized elderly patients with hematological infection, so as to facilitate clinical prevention and treatment. Methods The elderly (> 80 years old) hospitalized patients with hematological infection in our hospital from 2015 to 2021 were selected for analysis. Firstly, 314 patients with extensively drug-resistant Acinetobacter baumannii hematological infection were distinguished by etiological analysis. A total of 98 cases of death were detected during hospitalization (later referred to as the observation group). By comparing with the surviving patients (216 cases) (later referred to as the control group), the general data of patients with XDRAB infection were collected, and the risk of death and its influencing factors were analyzed. Results In the study, the proportion of patients in the observation group who used catheters was higher, the catheter retention time was longer, the acute physiology and chronic health status II scores were higher, and the proportion of patients who lost self-care ability was also higher. Conclusion The death of blood borne infection of extensively drug-resistant Acinetobacter baumannii in elderly patients is affected by many factors. Among them, patients who use catheters for a long time, have poor self-care ability and lose self-care ability have a higher risk of death, which is worthy of clinical attention.

Influence of seamless nursing model of humanistic care on nursing quality and blood infection risk of neurosurgical ICU patients and its correlation with APACHE II score.

This research aims to explore the influence of seamless nursing mode of humanistic care on nursing quality and blood infection risk of ICU patients in neurosurgery, and the model of correlation with APACHE II score. 110 ICU patients are grouped into control set and study set, which are, respectively, given the previous routine nursing and the seamless management based on humanistic care to compare the two sets in the following aspects: nursing quality, blood infection rate, anxiety and depression extension before and after nursing, nursing satisfaction and APACHE II score, and to figure out the correlation between patient nursing quality score, and to compare blood infection and APACHE II score. Comparison and statistical analysis are used to disclose the influence and the correlation. The results show that there is not only a large negative correlation between nursing quality scores and APACHE II scores, but also a large negative correlation between the risk of blood infection and APACHE II score.

Automated characterisation of neutrophil activation phenotypes in ex vivo human Candida blood infections.

Rapid identification of pathogens is required for early diagnosis and treatment of life-threatening bloodstream infections in humans. This requirement is driving the current developments of molecular diagnostic tools identifying pathogens from human whole blood after successful isolation and cultivation. An alternative approach is to determine pathogen-specific signatures from human host immune cells that have been exposed to pathogens. We hypothesise that activated immune cells, such as neutrophils, may exhibit a characteristic behaviour - for instance in terms of their speed, dynamic cell morphology - that allows (i) identifying the type of pathogen indirectly and (ii) providing information on therapeutic efficacy. In this feasibility study, we propose a method for the quantitative assessment of static and morphodynamic features of neutrophils based on label-free time-lapse imaging data. We investigate neutrophil activation phenotypes after confrontation with fungal pathogens and isolation from a human whole-blood assay. In particular, we applied a machine learning supported approach to time-lapse microscopy data from different infection scenarios and were able to distinguish between Candida albicans and C. glabrata infection scenarios with test accuracies well above 75%, and to identify pathogen-free samples with accuracy reaching 100%. These results significantly exceed the test accuracies achieved using state-of-the-art deep neural networks to classify neutrophils by their morphodynamics.

From environmental adaptation to host survival: Attributes that mediate pathogenicity of Candida auris.

Candida species are a major cause of invasive fungal infections. While Candida albicans, C. glabrata, C. parapsilosis, and C. tropicalis are the most dominant species causing life-threatening candidiasis, C. auris recently emerged as a new species causing invasive infections with high rates of clinical treatment failures. To mimic initial phases of systemic Candida infections with dissemination via the bloodstream and to elucidate the pathogenic potential of C. auris, we used an ex vivo whole blood infection model. Similar to other clinically relevant Candida spp., C. auris is efficiently killed in human blood, but showed characteristic patterns of immune cell association, survival rates, and cytokine induction. Dual-species transcriptional profiling of C. auris-infected blood revealed a unique C. auris gene expression program during infection, while the host response proofed similar and conserved compared to other Candida species. C. auris-specific responses included adaptation and survival strategies, such as counteracting oxidative burst of immune cells, but also expression of potential virulence factors, (drug) transporters, and cell surface-associated genes. Despite comparable pathogenicity to other Candida species in our model, C. auris-specific transcriptional adaptations as well as its increased stress resistance and long-term environmental survival, likely contribute to the high risk of contamination and distribution in a nosocomial setting. Moreover, infections of neutrophils with pre-starved C. auris cells suggest that environmental preconditioning can have modulatory effects on the early host interaction. In summary, we present novel insights into C. auris pathogenicity, revealing adaptations to human blood and environmental niches distinctive from other Candida species.

Antibiotics Act with vB_AbaP_AGC01 Phage against Acinetobacter baumannii in Human Heat-Inactivated Plasma Blood and Galleria mellonella Models.

Increasing multidrug resistance has led to renewed interest in phage-based therapy. A combination of the bacteriophages and antibiotics presents a promising approach enhancing the phage therapy effectiveness. First, phage candidates for therapy should be deeply characterized. Here we characterize the bacteriophage vB_AbaP_AGC01 that poses antibacterial activity against clinical Acinetobacter baumannii strains. Moreover, besides genomic and phenotypic analysis our study aims to analyze phage-antibiotic combination effectiveness with the use of ex vivo and in vivo models. The phage AGC01 efficiently adsorbs to A. baumannii cells and possesses a bacteriolytic lifecycle resulting in high production of progeny phages (317 ± 20 PFU × cell-1). The broad host range (50.27%, 93 out of 185 strains) against A. baumannii isolates and the inability of AGC01 to infect other bacterial species show its high specificity. Genomic analysis revealed a high similarity of the AGC01 genome sequence with that of the Friunavirus genus from a subfamily of Autographivirinae. The AGC01 is able to significantly reduce the A. baumannii cell count in a human heat-inactivated plasma blood model (HIP-B), both alone and in combination with antibiotics (gentamicin (GEN), ciprofloxacin (CIP), and meropenem (MER)). The synergistic action was observed when a combination of phage treatment with CIP or MER was used. The antimicrobial activity of AGC01 and phage-antibiotic combinations was confirmed using an in vivo larva model. This study shows the greatest increase in survival of G. mellonella larvae when the combination of phage (MOI = 1) and MER was used, which increased larval survival from 35% to 77%. Hence, AGC01 represents a novel candidate for phage therapy. Additionally, our study suggests that phages and antibiotics can act synergistically for greater antimicrobial effect when used as combination therapy.

Real-Time Monitoring of Bacteria Clearance From Blood in a Murine Model.

Bloodstream infections, especially those that are antibiotic resistant, pose a significant challenge to health care leading to increased hospitalization time and patient mortality. There are different facets to this problem that make these diseases difficult to treat, such as the difficulty to detect bacteria in the blood and the poorly understood mechanism of bacterial invasion into and out of the circulatory system. However, little progress has been made in developing techniques to study bacteria dynamics in the bloodstream. Here, we present a new approach using an in vivo flow cytometry platform for real-time, noninvasive, label-free, and quantitative monitoring of the lifespan of green fluorescent protein-expressing Staphylococcus aureus and Pseudomonas aeruginosa in a murine model. We report a relatively fast average rate of clearance for S. aureus (k = 0.37 ± 0.09 min-1 , half-life ~1.9 min) and a slower rate for P. aeruginosa (k = 0.07 ± 0.02 min-1 , half-life ~9.6 min). We also observed what appears to be two stages of clearance for S. aureus, while P. aeruginosa appeared only to have a single stage of clearance. Our results demonstrate that an advanced research tool can be used for studying the dynamics of bacteria cells directly in the bloodstream, providing insight into the progression of infectious diseases in circulation. © 2019 International Society for Advancement of Cytometry.

Severe acute pancreatitis with blood infection by Candida glabrata complicated severe agranulocytosis: a case report.

BACKGROUND: Blood infection with Candida glabrata often occurs in during severe acute pancreatitis (SAP). It complicate severe agranulocytosis has not been reported. CASE PRESENTATION: We present a case where a SAP patient presenting with a sudden hyperpyrexia was treated for 19 days. We monitored her routine blood panel and CRP levels once or twice daily. The results showed that WBC count decreased gradually. And the lowest level of WBC was appeared at the 21st day of treatment. WBC 0.58 × 109/L(4.0-10.0 × 109/L), neutrophils 0.1 × 109/L [2.20%] (2.5-7.5 × 109/L). During treatment, Candida glabrata was identified as the infecting agent through blood culture, drainage tubes culture and gene detection. During anti-infection therapy, the patient had severe agranulocytosis. With control of the infection, her WBC and granulocyte counts gradually returned to the normal range. CONCLUSIONS: Blood infection with Candida glabrata can complicate severe agranulocytosis.

Predictive Virtual Infection Modeling of Fungal Immune Evasion in Human Whole Blood.

Bloodstream infections by the human-pathogenic fungi Candida albicans and Candida glabrata increasingly occur in hospitalized patients and are associated with high mortality rates. The early immune response against these fungi in human blood comprises a concerted action of humoral and cellular components of the innate immune system. Upon entering the blood, the majority of fungal cells will be eliminated by innate immune cells, i.e., neutrophils and monocytes. However, recent studies identified a population of fungal cells that can evade the immune response and thereby may disseminate and cause organ dissemination, which is frequently observed during candidemia. In this study, we investigate the so far unresolved mechanism of fungal immune evasion in human whole blood by testing hypotheses with the help of mathematical modeling. We use a previously established state-based virtual infection model for whole-blood infection with C. albicans to quantify the immune response and identified the fungal immune-evasion mechanism. While this process was assumed to be spontaneous in the previous model, we now hypothesize that the immune-evasion process is mediated by host factors and incorporate such a mechanism in the model. In particular, we propose, based on previous studies that the fungal immune-evasion mechanism could possibly arise through modification of the fungal surface by as of yet unknown proteins that are assumed to be secreted by activated neutrophils. To validate or reject any of the immune-evasion mechanisms, we compared the simulation of both immune-evasion models for different infection scenarios, i.e., infection of whole blood with either C. albicans or C. glabrata under non-neutropenic and neutropenic conditions. We found that under non-neutropenic conditions, both immune-evasion models fit the experimental data from whole-blood infection with C. albicans and C. glabrata. However, differences between the immune-evasion models could be observed for the infection outcome under neutropenic conditions with respect to the distribution of fungal cells across the immune cells. Based on these predictions, we suggested specific experimental studies that might allow for the validation or rejection of the proposed immune-evasion mechanism.

Diagnostic value of procalcitonin in distinguishing Gram-negative bacterial blood stream infection from Gram-positive bacterial infection in early stage of sepsis / 中华急诊医学杂志

Objective To investigate the diagnostic values of procalcitonin (PCT),high sensitive C-reactive protein (hs-CRP),white blood cell (WBC)and percentage of neutrocyte (NEU％)in Gramnegative and Gram-positive bacterial blood stream infection in early stage of sepsis in order to investigate the correlation between PCT and APACHE lⅡ score as well as between PCT and SOFA score,and the prognostic value in assessment of Gram-negative and Gram-positive bacterial blood stream infection.Methods Clinical data of patients admitted to ICU from January 2012 through December 2014 were retrospectively analyzed.A total of 124 sepsis patients with blood stream infection were checked with PCT,hs-CRP,WBC and NEU％ tests,and APACHE Ⅱ score and SOFA score were calculated.The differences in APACHE Ⅱ score and SOFA score were compared between Gram-negative group (n =41) and Gram-positive group (n =83).The correlation between PCT and APACHE Ⅱ score as well as between PCT and SOFA score was analyzed.The differences in diagnostic values of PCT,hs-CRP,WBC and NEU％ between Gram-negative group and Grampositive group were analyzed by using receiver operating characteristic (ROC) curve and it was plotted to assess the prognostic values of PCT,hs-CRP,WBC and NEU％ for septic patients with blood stream infection.Results Compared with Gram-positive group,the levels of PCT [.55.32 (22.01,97.11) vs.2.13 (0.27,5.27)] (P ＜0.01),hs-CRP [105.09 (69.97,186.12) vs.70.54 (42.37,138.63)] (P=0.508),NEU％ [88.30 (75.79,93.52) vs.55.32 (22.01,97.11)] (P=0.302) were higher but WBC was lower [13.59 (10.74,17.58) vs.13.73 (11.32,20.90)] (P=0.058) in Gram-negative group.The ROC curve analysis of PCT showed the area under the curve (AUC) was 0.867 (95％ CI:0.789-0.946).When the optimal cutoff point of PCT was 17.48 ng/mL,the largest Youden's index was found to be 0.661 with 76.9％ sensitivity and 89.2％ specificity.Between two groups,there were significant differences in APACHE Ⅱ score and SOFA score (27.46 ± 9.60 vs.23.67 ± 7.74,P =0.020;8.05 ±3.38 vs.6.59-±3.45,P =0.028).There was significant difference in diagnostic value between PCT and SOFA (r =0.536,P =0.036) in Gram-negative group but no significant difference in Gram-positive group.Conclusions Higher PCT levels are found in Gram-negative group and it can play a role in differntiation between the Gram-negative group and Gram-positive group rather than hs-CRP,WBC and NEU％.PCT can be a better indicator for evaluation of severity of sepsis as well as for prognosis of sepsis patients with Gram-negative bacterium infection.

Clinical study analysis on ulinastatin for treating systemic inflammatory response in patients with serious blood infection / 国际检验医学杂志

Objective To investigate the clinical effect of ulinastatin treating systemic inflammatory response in the patients with serious blood infection .Methods The patients with serious blood infection in our hospital were selected and divided into the control group and observation group .The control group was treated with the conventional treatment ,while on this basis the observation group was added with ulinastatin .The body temperature ,heart rate ,respiratory rate ,WBC ,Scr ,CRP ,PCT ,TNF-α,APACHE Ⅱscore and mortality rate were compared between the two groups .Results The body temperature ,heart rate ,respiratory rate and A-PACHE Ⅱ score in the observation group were lower than those in the control group ,moreover ,the levels of WBC ,CRP ,PCT , TNF-αand Scr ,and mortality rate in the observation group were lower than those in the control group ,the differences were statisti-cally significant (P<0 .05) .Conclusion Uinastatin has good effect for treating systemic inflammatory response in the patients with serious blood infection ,which can decrease the mortality and is worth clinical promotion .

High risk factors of blood infection in voluntary donors in China: A single-centre retrospective study / 中华器官移植杂志

Objective To analyze the high risk factors of blood infection in Chinese citizens' organ donation,provide the basic evidence for early protection,increase the success rate of donor distribution,and expand the Chinese organ donation pool.Methods A retrospective study was performed on 70 cases of donation recruited during October 2014 to January 2016.The incidence of blood infection in these donors was analyzed.The univariate analysis and multivariate logistic regression analysis were used to find out the high risk factors influencing the donor blood infection.Finally,the donor blood infection assessment model and the receiver operating characteristic (ROC) curve were established to assess the sensitivity and specificity.Results The overall infection rate was 64.3％ (45/70).The pulmonary,blood,and urinary tract infection rate was 42.9％,31.4％ and 1.4％ respectively.The total length of hospital stay (＞10 days) (P =0.017),oxygenation index (＜ 233.5 ± 107.0) (P =0.046),aspartate aminotransferase (＞196.9 ± 329.1 U/L) (P =0.044),and valley alanine aminotransferase (＞95.0 ± 78.1 U/L) (P =0.026) were four risk factors for predicting the donor blood infection.The multivariate logistic regression analysis revealed that the total length of stay ＞10 days along with the donors' oxygenation index (＜233.5 ± 107.0) was independent risk factor for predicting the blood infection.The donor blood infection model was:0.193 + 1.753 hospital stay (＞10 days)-0.007 oxygenation index.The sensitivity and specificity were 0.682 and 0.75 (P ＜0.001) respectively.Conclusion For a long-term stay in ICU,the rate of blood infection for donors was much higher,at this time,the most effective antibiotics should be chosen.Besides,improving donor oxygenation index and liver function can reduce the incidence of infection.

Clinical characteristics and drug resistance of 176 strains of candida isolated from children with blood infections / 中国小儿急救医学

Objective To analyze the pathogen distribution and drug resistance of candida isolated from children with blood infections in our hospital,and to provide reference for clinical effective prevention and treatment.Methods The blood specimens of pediatric patients were collected between January 2009 and December 2015,and were cultured using BacT/ALERT 3D and BD9140 instruments.The candida were separated with Sobaurandps agar culture medium,and identified with chromogenic medium,API 20CAUX test strips or VITEK-2 compact YST card.The minimal inhibitory concentration of 5 drugs were determined by ATB FUNGUS 3 system.Results In 176 cases,92 strains (52.3％) were from neonatal ward,and 46 strains (26.1％) were from PICU.In newborn group,85 strains were isolated from premature,which contained the low and very low birth weight infants (37 strains),pneumonia(20 strains),neonatal respiratory distress syndrome(9 strains).In PICU,the strains were commonly isolated from children with severe infection.Among 176 strains of candida,71 strains (40.3％) were C.albicans,62 strains (35.2％) were C.parapsilosis,16 strains(9.1％) were C.glabrata,9 strains(5.1％) were C.tropicalis,and 18 strains(10.2％) belonged to other candida.Conclusion Candida blood infections can happen at all age of chlidren.The most common strains detected from blood were C.albicans,followed by C.parapsilosis.Most of these strains are susceptible to antifungal drugs,such as fluconazole,except C.glabrata.The sensitive rates to commonly used antifungal drug are more than 93％.The selection of antifungal drugs should be based on the species of strains.

Bottom-up modeling approach for the quantitative estimation of parameters in pathogen-host interactions.

Opportunistic fungal pathogens can cause bloodstream infection and severe sepsis upon entering the blood stream of the host. The early immune response in human blood comprises the elimination of pathogens by antimicrobial peptides and innate immune cells, such as neutrophils or monocytes. Mathematical modeling is a predictive method to examine these complex processes and to quantify the dynamics of pathogen-host interactions. Since model parameters are often not directly accessible from experiment, their estimation is required by calibrating model predictions with experimental data. Depending on the complexity of the mathematical model, parameter estimation can be associated with excessively high computational costs in terms of run time and memory. We apply a strategy for reliable parameter estimation where different modeling approaches with increasing complexity are used that build on one another. This bottom-up modeling approach is applied to an experimental human whole-blood infection assay for Candida albicans. Aiming for the quantification of the relative impact of different routes of the immune response against this human-pathogenic fungus, we start from a non-spatial state-based model (SBM), because this level of model complexity allows estimating a priori unknown transition rates between various system states by the global optimization method simulated annealing. Building on the non-spatial SBM, an agent-based model (ABM) is implemented that incorporates the migration of interacting cells in three-dimensional space. The ABM takes advantage of estimated parameters from the non-spatial SBM, leading to a decreased dimensionality of the parameter space. This space can be scanned using a local optimization approach, i.e., least-squares error estimation based on an adaptive regular grid search, to predict cell migration parameters that are not accessible in experiment. In the future, spatio-temporal simulations of whole-blood samples may enable timely stratification of sepsis patients by distinguishing hyper-inflammatory from paralytic phases in immune dysregulation.

Development of droplet digital PCR for the detection of Babesia microti and Babesia duncani.

Babesia spp. are obligate protozoan parasites of red blood cells. Transmission to humans occurs through bites from infected ticks or blood transfusion. Infections with B. microti account for the majority of the reported cases of human babesiosis in the USA. A lower incidence is caused by the more recently described species B. duncani. The current gold standard for detection of Babesia is microscopic examination of blood smears. Recent PCR-based assays, including real-time PCR, have been developed for B. microti. On the other hand, molecular assays that detect and distinguish between B. microti and B. duncani infections are lacking. Closely related species of Babesia can be differentiated due to sequence variation within the internal transcribed spacer (ITS) regions of nuclear ribosomal RNAs. In the present study, we targeted the ITS regions of B. microti and B. duncani to develop sensitive and species-specific droplet digital PCR (ddPCR) assays. The assays were shown to discriminate B. microti from B. duncani and resulted in limits of detection of ~10 gene copies. Moreover, ddPCR for these species were useful in DNA extracted from blood of experimentally infected hamsters, detecting infections of low parasitemia that were negative by microscopic examination. In summary, we have developed sensitive and specific quantitative ddPCR assays for the detection of B. microti and B. duncani in blood. Our methods could be used as sensitive approaches to monitor the progression of parasitemia in rodent models of infection as well as serve as suitable molecular tests in blood screening.

Analysis on pathogenic distribution and bacterial drug resistance of clinical blood infections / 国际检验医学杂志

Objective To understand the pathogenic distribution and drug resistance in the patients with blood infections in our hospital to provide reference for the empirical medication of blood infections .Methods The cases of blood infections in our hospital during the period 2012-2014 were performed the retrospective investigation .The BD Bactec blood culture system was adopted to conduct the blood culture .The bacterial strain identification and susceptibility test was conducted by using the Microscan Walkaway 40 and the data were analyzed by adopting the WHONET5 .6 software .Results The top four of department distribution in blood infections were the digestive system department ,lung diseases department ,orthopedic department and surgery department ,account‐ing for 25 .20% ,19 .60% ,14 .70% and 10 .50% respectively .The pathogens were mainly Gram negative bacteria ,and the top three were Escherichia coli ,Klebsiella pneumoniae and non fermenting bacteria ,accounting for 44 .10% ,13 .30% and 7 .69% respective‐ly .The top three of Gram positive bacteria were coagulase negative staphylococci ,Staphylococcus aureus and Enterococcus ,account‐ing for 12 .58% ,9 .09% and 7 .69% respectively .The positive rates of ESBL in Escherichia coli and Klebsiella pneumoniae were 45 .5% and 60 .8% respectively .The detection rate of methicillin resistant coagulase negative staphylococci (MRCNS) was 55 .5% , which of methicillin-resistant staphylococcus aureus (MRSA) was 58 .0% .Conclusion The cases of blood infections in our hospi‐tal come from different wards areas ,and the digestive system department is in the majority mostly .The pathogenic bacteria are dom‐inated by Gram negative bacteria ,and the treatment should rationally use the antibacterial drugs according to the bacteria drug re‐sistance situation .

Enhanced nucleic acid amplification with blood in situ by wire-guided droplet manipulation (WDM).

There are many challenges facing the use of molecular biology to provide pertinent information in a timely, cost effective manner. Wire-guided droplet manipulation (WDM) is an emerging format for conducting molecular biology with unique characteristics to address these challenges. To demonstrate the use of WDM, an apparatus was designed and assembled to automate polymerase chain reaction (PCR) on a reprogrammable platform. WDM minimizes thermal resistance by convective heat transfer to a constantly moving droplet in direct contact with heated silicone oil. PCR amplification of the GAPDH gene was demonstrated at a speed of 8.67 s/cycle. Conventional PCR was shown to be inhibited by the presence of blood. WDM PCR utilizes molecular partitioning of nucleic acids and other PCR reagents from blood components, within the water-in-oil droplet, to increase PCR reaction efficiency with blood in situ. The ability to amplify nucleic acids in the presence of blood simplifies pre-treatment protocols towards true point-of-care diagnostic use. The 16s rRNA hypervariable regions V3 and V6 were amplified from Klebsiella pneumoniae genomic DNA with blood in situ. The detection limit of WDM PCR was 1 ng/µL or 10(5)genomes/µL with blood in situ. The application of WDM for rapid, automated detection of bacterial DNA from whole blood may have an enormous impact on the clinical diagnosis of infections in bloodstream or chronic wound/ulcer, and patient safety and morbidity.