Laboratory aids in the diagnosis of invasive candidiasis

The laboratory diagnosis of candidiasis continues to be problematic; however, there have been several advances in the past decade which promise to enhance our ability to identify patients at high risk for infection and/or to document invasive candidiasis in critically ill and immunocompromised patients. The introduction of commercially available biphasic blood culture medium and subsequently the lysiscentrifugation procedure has markedly improved the ability of laboratories to detect fungemia. Although serologic methods have not been very successful in diagnosing candidiasis in immunocompromised patients, several antigen detection methods are now under investigation. In addition, detection of fungal metabolites such as D-arabinitol remains promising. Finally, application of the techniques of molecular biology for typing and detection of fungal pathogens has expanded our understanding of candidal infections and may offer the most sensitive and specific means of diagnosing invasive candidiasis.     

Particularidades clínicas del paciente crítico con infección fúngica invasora

BackgroundInvasive fungal infection (IFI) is an entity that encompasses different types of infections caused by different types of those fungi pathogenic for humans. In the setting of critically ill patients with multiple and oftenconcurrent risk factors and comorbidities the most common are those caused by the Candida and Aspergillus species. Among the characteristics of IFI in critically ill patients, three aspects can be highlighted: those related to the host (e.g.: risk factors, clinical severity), those related with the pathogen (sensitivity, virulence), or those concerning antifungal treatment (spectrum, features PK / PD, safety, interactions). The fungus that most often causes an IFI in critically ill patients is Candida; the most common type infections are candidemia, Candida peritonitis and catheter-related infections. In recent years new antifungal treatments have expanded the therapeutic options, with echinocandins as a clear choice, often the first in the latest guidelines in critically ill patients with IFI.Case reportWe report the case of a critically ill patient having the most common risk factors, multiple organ dysfunction and development of an IFI. The complexity of establishing an antifungal treatment from the moment of its inception, its setting, and the considerations of the different therapeutic possibilities according to organ dysfunction of the patient are discussed. The antifungal treatment options mentioned in the current guidelines and recommendations are also evaluated.ConclusionsThe most common fungal infection in critically ill patients is invasive candidiasis, with candidemia or candida peritonitis being the most frequent clinical presentations. Candins have brought new possibilities for treating these complex patients due to their good safety profile and clinical efficacy.     

Early Prediction of Sepsis Incidence in Critically Ill Patients Using Specific Genetic Polymorphisms

Several diagnostic methods for the evaluation and monitoring were used to find out the pro-inflammatory status, as well as incidence of sepsis in critically ill patients. One such recent method is based on investigating the genetic polymorphisms and determining the molecular and genetic links between them, as well as other sepsis-associated pathophysiologies. Identification of genetic polymorphisms in critical patients with sepsis can become a revolutionary method for evaluating and monitoring these patients. Similarly, the complications, as well as the high costs associated with the management of patients with sepsis, can be significantly reduced by early initiation of intensive care.     

Does hemodynamic monitoring complement conventional methods of assessment in the critically ill cardiac patient?

Conventional clinical methods give qualitative information about left ventricular function in the critically ill patient. However, the information gathered from the physical examination and noninvasive methods is subject to important pitfalls with respect to both its nonspecificity and interobserver variability. The advent of hemodynamic monitoring has highlighted these difficulties and provided more quantitative information that is relevant in both diagnosis and treatment of the critically ill patient.     

Hsp72 Is a Novel Biomarker to Predict Acute Kidney Injury in Critically Ill Patients

Background and Objectives

Acute kidney injury (AKI) complicates the course of disease in critically ill patients. Efforts to change its clinical course have failed because of the fail in the early detection. This study was designed to assess whether heat shock protein (Hsp72) is an early and sensitive biomarker of acute kidney injury (AKI) compared with kidney injury molecule (Kim-1), neutrophil gelatinase-associated lipocalin (NGAL), and interleukin-18 (IL-18) biomarkers.

Methods

A total of 56 critically ill patients fulfilled the inclusion criteria. From these patients, 17 developed AKI and 20 were selected as controls. In AKI patients, Kim-1, IL-18, NGAL, and Hsp72 were measured from 3 days before and until 2 days after the AKI diagnosis and in no-AKI patients at 1, 5 and 10 days after admission. Biomarker sensitivity and specificity were determined. To validate the results obtained with ROC curves for Hsp72, a new set of critically ill patients was included, 10 with AKI and 12 with no-AKI patients.

Results

Urinary Hsp72 levels rose since 3 days before the AKI diagnosis in critically ill patients; this early increase was not seen with any other tested biomarkers. Kim-1, IL-18, NGAL, and Hsp72 significantly increased from 2 days before AKI and remained elevated during the AKI diagnosis. The best sensitivity/specificity was observed in Kim-1 and Hsp72: 83/95% and 100/90%, respectively, whereas 1 day before the AKI diagnosis, the values were 100/100% and 100/90%, respectively. The sensibility, specificity and accuracy in the validation test for Hsp72 were 100%, 83.3% and 90.9%, respectively.

Conclusions

The biomarker Hsp72 is enough sensitive and specific to predict AKI in critically ill patients up to 3 days before the diagnosis.     

危重症患者预后的危险因素及相关指标的预后预测价值分析

目的:探讨危重症患者预后的危险因素,并分析相关指标对患者预后的预测价值。方法:对2016年4月至2018年4月上海交通大学附属第六人民医院救治的5585例ICU危重症患者病例进行回顾性分析,收集患者一般资料、初始乳酸水平、24 h乳酸水平、24 h乳酸清除率以及APACHEII评分等,采用多因素logistic回归分析危重症患者预后的影响因素,ROC曲线评估多项指标预测患者预后的价值。结果:共纳入1465例危重症患者,多因素logistic回归分析显示,住ICU平均时间长、APACHEII评分高、初始乳酸水平大于4 mmol/L、24 h乳酸清除率低、术后肾功能异常是危重症患者住院期间死亡的危险因素(P0.05),24 h乳酸清除率、APACHEII评分及初始乳酸水平三者联合评估预测患者住院死亡率的ROC曲线下面积(ROC-AUC)为0.900,高于单独检测。结论:住ICU时间、APACHEII评分、初始乳酸水平、24 h乳酸清除率、术后肾功能是危重症患者住院期间死亡的影响因素,初始乳酸水平、24 h乳酸清除率和APACHEII评分联合使用可以更好地预测危重症患者的短期预后。     

Blood Lactate/ATP Ratio,as an Alarm Index and Real-Time Biomarker in Critical Illness

Objective

The acute physiology, age and chronic health evaluation (APACHE) II score and other related scores have been used for evaluation of illness severity in the intensive care unit (ICU), but there is still a need for real-time and sensitive prognostic biomarkers. Recently, alarmins from damaged tissues have been reported as alarm-signaling molecules. Although ATP is a member of the alarmins and its depletion in tissues closely correlates with multiple-organ failure, blood ATP level has not been evaluated in critical illness. To identify real-time prognostic biomarker of critical illness, we measured blood ATP levels and the lactate/ATP ratio (ATP-lactate energy risk score, A-LES) in critically ill patients.

Methods and Results

Blood samples were collected from 42 consecutive critically ill ICU patients and 155 healthy subjects. The prognostic values of blood ATP levels and A-LES were compared with APACHE II score. The mean ATP level (SD) in healthy subjects was 0.62 (0.19) mM with no significant age or gender differences. The median ATP level in severely ill patients at ICU admission was significantly low at 0.31 mM (interquartile range 0.25 to 0.44) than the level in moderately ill patient at 0.56 mM (0.38 to 0.70) (P<0.01). Assessment with ATP was further corrected by lactate and expressed as A-LES. The median A-LES was 2.7 (2.1 to 3.3) in patients with satisfactory outcome at discharge but was significantly higher in non-survivors at 38.9 (21.0 to 67.9) (P<0.01). Receiver operating characteristic analysis indicated that measurement of blood ATP and A-LES at ICU admission are as useful as APACHE II score for prediction of mortality.

Conclusion

Blood ATP levels and A-LES are sensitive prognostic biomarkers of mortality at ICU admission. In addition, A-LES provided further real-time evaluation score of illness severity during ICU stay particularly for critically ill patients with APACHE II scores of ≥20.0.     

Assessment of body cell mass at bedside in critically ill patients

Critical illness affects body composition profoundly, especially body cell mass (BCM). BCM loss reflects lean tissue wasting and could be a nutritional marker in critically ill patients. However, BCM assessment with usual isotopic or tracer methods is impractical in intensive care units (ICUs). We aimed to modelize the BCM of critically ill patients using variables available at bedside. Fat-free mass (FFM), bone mineral (Mo), and extracellular water (ECW) of 49 critically ill patients were measured prospectively by dual-energy X-ray absorptiometry and multifrequency bioimpedance. BCM was estimated according to the four-compartment cellular level: BCM = FFM - (ECW/0.98) - (0.73 × Mo). Variables that might influence the BCM were assessed, and multivariable analysis using fractional polynomials was conducted to determine the relations between BCM and these data. Bootstrap resampling was then used to estimate the most stable model predicting BCM. BCM was 22.7 ± 5.4 kg. The most frequent model included height (cm), leg circumference (cm), weight shift (Δ) between ICU admission and body composition assessment (kg), and trunk length (cm) as a linear function: BCM (kg) = 0.266 × height + 0.287 × leg circumference + 0.305 × Δweight - 0.406 × trunk length - 13.52. The fraction of variance explained by this model (adjusted r(2)) was 46%. Including bioelectrical impedance analysis variables in the model did not improve BCM prediction. In summary, our results suggest that BCM can be estimated at bedside, with an error lower than ±20% in 90% subjects, on the basis of static (height, trunk length), less stable (leg circumference), and dynamic biometric variables (Δweight) for critically ill patients.     

Tratamiento antifúngico individualizado en el paciente crítico con infección fúngica invasora

Invasive candidiasis (IC) is the most common invasive fungal infection (IFI) affecting critically ill patients, followed by invasive pulmonary aspergillosis (IPA). International guidelines provide different recommendations for a first-line antifungal therapy and, in most of them, echinocandins are considered the first-line treatment for IC, and triazoles are so for the treatment of IPA. However, liposomal amphotericin B (L-AmB) is still considered a second-line therapy for both clinical entities. Although in the last decade the management of IFI has improved, several controversies persist. The antifungal drugs currently available may have a suboptimal activity, or be wrongly used in certain IFI involving critically ill patients. The aim of this review is to analyze when to provide individualized antifungal therapy to critically ill patients suffering from IFI, emphasizing the role of L-AmB. Drug-drug interactions, the clinical status, infectious foci (peritoneal candidiasis is discussed), the fungal species involved, and the need of monitoring the concentration of the antifungal drug in the patient are considered.     

Evaluation of the Broad-Range PCR/ESI-MS Technology in Blood Specimens for the Molecular Diagnosis of Bloodstream Infections

Background

Rapid identification of the etiological agent in bloodstream infections is of vital importance for the early administration of the most appropriate antibiotic therapy. Molecular methods may offer an advantage to current culture-based microbiological diagnosis. The goal of this study was to evaluate the performance of IRIDICA, a platform based on universal genetic amplification followed by mass spectrometry (PCR/ESI-MS) for the molecular diagnosis of sepsis-related pathogens directly from the patient’s blood.

Methods

A total of 410 whole blood specimens from patients admitted to Emergency Room (ER) and Intensive Care Unit (ICU) with clinical suspicion of sepsis were tested with the IRIDICA BAC BSI Assay (broad identification of bacteria and Candida spp.). Microorganisms grown in culture and detected by IRIDICA were compared considering blood culture as gold standard. When discrepancies were found, clinical records and results from other cultures were taken into consideration (clinical infection criterion).

Results

The overall positive and negative agreement of IRIDICA with blood culture in the analysis by specimen was 74.8% and 78.6%, respectively, rising to 76.9% and 87.2% respectively, when compared with the clinical infection criterion. Interestingly, IRIDICA detected 41 clinically significant microorganisms missed by culture, most of them from patients under antimicrobial treatment. Of special interest were the detections of one Mycoplasma hominis and two Mycobacterium simiae in immunocompromised patients. When ICU patients were analyzed separately, sensitivity, specificity, positive and negative predictive values compared with blood culture were 83.3%, 78.6%, 33.9% and 97.3% respectively, and 90.5%, 87.2%, 64.4% and 97.3% respectively, in comparison with the clinical infection criterion.

Conclusions

IRIDICA is a promising technology that offers an early and reliable identification of a wide variety of pathogens directly from the patient’s blood within 6h, which brings the opportunity to improve management of septic patients, especially for those critically ill admitted to the ICU.     

白斑综合症病毒实时荧光LAMP检测方法的建立及应用

研究利用ESE-Quant tube scanner检测平台, 建立了一套基于环介导等温扩增技术(Loop-Mediated Isothermal Amplification, LAMP)的实时荧光检测方法, 用于白斑综合征病毒(White Spot Syndrome Virus, WSSV)的检测; 并在此基础上, 与巢式PCR、Real-time PCR和其他已发表的4种LAMP方法在检测灵敏度、实际应用方面进行比较. 结果显示, 研究建立的实时荧光LAMP检测方法在63℃恒温反应30min可检测到最低为105倍稀释的基因组DNA模板, 与Real-time PCR检测方法的灵敏度相当, 高于巢式PCR和其他已发表的4种LAMP方法的检测灵敏度; 而且特异性较好, 与传染性皮下及造血组织坏死病毒等5种常见对虾病原DNA均无交叉反应. 通过构建质粒进一步进行灵敏度测试显示, 本研究建立的实时荧光LAMP检测方法最低检测限度为24个拷贝质粒DNA, 检出时间亦为30min. 通过对66份待检样品的检测结果显示, 实时荧光LAMP检测方法的检出阳性率为7.57%, 准确率为100%, 高于其他WSSV的检测方法. 因此, 研究建立的WSSV实时荧光LAMP检测方法, 操作简单, 反应速度快, 特异性好, 灵敏度高, 成本低廉, 可以直观、实时地观察反应的进行情况, 适合对虾养殖现场及诊断实验室的WSSV快速检测.       

Cytokine storm intervention in the early stages of COVID-19 pneumonia

Clinical intervention in patients with corona virus disease 2019 (COVID-19) has demonstrated a strong upregulation of cytokine production in patients who are critically ill with SARS-CoV2-induced pneumonia. In a retrospective study of 41 patients with COVID-19, most patients with SARS-CoV-2 infection developed mild symptoms, whereas some patients later developed aggravated disease symptoms, and eventually passed away because of multiple organ dysfunction syndrome (MODS), as a consequence of a severe cytokine storm. Guidelines for the diagnosis and treatment of SARS-CoV-2 infected pneumonia were first published January 30^th, 2020; these guidelines recommended for the first time that cytokine monitoring should be applied in severely ill patients to reduce pneumonia related mortality. The cytokine storm observed in COVID-19 illness is also an important component of mortality in other viral diseases, including SARS, MERS and influenza. In view of the severe morbidity and mortality of COVID-19 pneumonia, we review the current understanding of treatment of human coronavirus infections from the perspective of a dysregulated cytokine and immune response.     

Aspergillus in the Intensive Care Unit

Invasive aspergillosis (IA), the most life-threatening form of aspergillosis, has become a major opportunistic fungal disease in immunocompromised patients. In high-risk patients with hematologic malignancies, IA appears to decline with the use of mold-active antifungal prophylaxis, but the situation is less clear in other patient groups at risk for IA, and precise epidemiologic data from patients treated in intensive care units (ICUs) are lacking. Most Aspergillus culture isolates from nonsterile body sites do not represent disease, but isolation of Aspergillus in critically ill patients is a marker of poor prognosis and is associated with high mortality regardless of invasion or colonization. This review presents current information on epidemiology, risk factors, and diagnosis, and discusses treatment options for patients with IA in the ICU.     

Comparison of the Therapeutic Effectiveness of Sustained Low-Efficiency Dialysis (SLED) with Continuous Blood Purification (CBP) in Critically Ill Patients

The differences in therapeutic effectiveness between sustained low-efficiency dialysis (SLED) and continuous blood purification (CBP) were investigated. In order to assess the different treatment methods, 56 critically ill patients were divided into two groups, the CBP group and the SLED group. A comparison was made between all the biochemical indicators, in-hospital duration, hemodynamic parameters, acute physiology and chronic health evaluation (APACHE-II), the survival, and the mortality rates. After treatment, the levels of serum creatine kinase isozyme MB (CK-MB), creatine kinase, creatinine, glutamate-oxalacetate transaminase (AST), glutamate-pyruvate transaminase (ALT), APACHE II score on the 1st, 2nd, and 7th day in both the treatment groups were lower than that before the treatment (P < 0.05). There are no statistical differences in in-hospital duration, biochemical indicators, APACHE II score, hemodynamic parameters, the survival rate and the mortality rate between the two groups (P > 0.05). It was concluded that SLED has similar hemodynamic stability with CBP and the two methods have similar treatment effects in critically ill patients. However, we noticed that SLED can be relatively economical and convenient for critically ill patients in clinical practice.     

Epidemiología de las candidiasis invasoras en población pediátrica y adulta

BackgroundInvasive candidiasis (IC) is the most frequent fungal disease in children and adults.AimsTo critically review and update the current epidemiology of Candida spp. disease in neonates, children and adults (critically ill patients and in oncohematologic patients and in solid organ transplant recipients).MethodsWe searched the PubMed/Medline, discussing the current data.Results and conclusionsIC is associated with high attributable morbimortality and increased healthcare costs. In the last decades the incidence of invasive Candida spp. disease has increased in critically ill patients, has decreased in oncohematologic patients, although currently the involvement of non-albicans Candida species in the etiology of this disease is increasing steadily.     

Immunoproteomics based identification of thioredoxin reductase GliT and novel <Emphasis Type="Italic">Aspergillus fumigatus</Emphasis> antigens for serologic diagnosis of invasive aspergillosis

Background  

There has been a rising incidence of invasive aspergillosis (IA) in critically ill patients, even in the absence of an apparent predisposing immunodeficiency. The diagnosis of IA is difficult because clinical signs are not sensitive and specific, and serum galactomannan has relatively low sensitivity in this group of patients. Therefore, more prompt and accurate disease markers for early diagnosis are needed. To establish disease markers demands a thorough knowledge of fungal antigens which may be detected in the serum or other body fluids of patients. Herein we report novel immunodominant antigens identified from extracellular proteins of Aspergillus fumigatus.     

Kinetic assay for the determination of the oxidative stress biomarker, advanced oxidation protein products (AOPP) in the human blood plasma

A number of human diseases and pathological conditions were found to be associated with increased oxidative stress. In the literature several techniques are available for the assessment of oxidative stress, but most of them are not applicable for a routine medical laboratory due to the complex methodology and/or financial reasons. We report here on a simple, inexpensive, kinetic assay for the determination of the oxidative stress biomarker, advanced oxidation protein products (AOPP) in the human blood plasma. METHODS: This study involved 70 patients (47M/23F; mean age: 64.6 y; range: 16-85) admitted to our Department with a wide range of cardiovascular and peripheral vascular diseases. Three critically ill patients were assigned for monitoring purposes. Plasma AOPP were simultaneously determined using an end-point assay as reference method and by a kinetic method developed in our laboratory. Plasma fibrinogen concentration was measured according to the Clauss method. RESULTS: There was a highly significant correlation (r2 = 0.588; p < 0.0001) between AOPP concentration (reference method) and AOPP reactivity (kinetic method). Both AOPP concentration and AOPP reactivity also significantly correlated with plasma fibrinogen concentration (r2 = 0.780; p < 0.0001; r2 = 0.564; p < 0.0001). The three representative cases presented appear to support the relevance of our novel method in the monitoring of critically ill patients. CONCLUSIONS: This simple and inexpensive kinetic assay can be widely used in any routine laboratory interested in oxidative stress research. It is especially recommended for monitoring critically ill or other patients.     

Recommendations for preventing transmission of HIV in health care settings.

The initial resuscitation and stabilization provided to a critically ill or injured child is often an important determinant of outcome. Before transfer to a tertiary care facility the initial care may be provided by physicians unaccustomed to managing critically ill children. The authors outline the unique aspects of resuscitation and stabilization of the critically ill child and give guidelines for the initial management of diseases affecting the central nervous system and respiratory tract (the most frequent indications for transfer to a tertiary care facility) and other, less frequent but important problems. In many situations it is worth while to enlist the expertise of the tertiary care centre, either by telephone consultation or by dispatch of a specially trained transport team.     

Pyrosequencing for the rapid and efficient quantification of allele-specific expression

We have developed a rapid and sensitive quantitative assay for the measurement of individual allelic ratios. This assay minimizes time and labor, the need for special restriction endonuclease enzymes for polymorphic sites, and avoids heteroduplex formation seen with traditional quantitative PCR-based methods. It has improved sensitivity compared to other methods and is capable of distinguishing 1% differences in allelic expression. This assay, termed Pyrosequencing for Imprinted Expression (PIE), involves the use of an intron-crossing PCR primer to generate the first PCR product. We applied the assay to analyze Insulin-like Growth Factor-2 (IGF2) imprinting in both human and mouse prostate tissues.