<clinical_study>
  <!-- This xml conforms to an XML Schema at:
    https://clinicaltrials.gov/ct2/html/images/info/public.xsd -->
  <required_header>
    <download_date>ClinicalTrials.gov processed this data on September 20, 2023</download_date>
    <link_text>Link to the current ClinicalTrials.gov record.</link_text>
    <url>https://clinicaltrials.gov/ct2/show/NCT03252028</url>
  </required_header>
  <id_info>
    <org_study_id>LINPEAS</org_study_id>
    <nct_id>NCT03252028</nct_id>
  </id_info>
  <brief_title>Rapid Test for Detection of the Focus of Infection in Post Neurosurgical Patients.</brief_title>
  <official_title>Rapid Test for Detection of the Focus of Infection in Post Neurosurgical Patients.</official_title>
  <sponsors>
    <lead_sponsor>
      <agency>Fariba Nayeri</agency>
      <agency_class>Other</agency_class>
    </lead_sponsor>
  </sponsors>
  <source>University Hospital, Linkoeping</source>
  <oversight_info>
    <has_dmc>No</has_dmc>
    <is_fda_regulated_drug>No</is_fda_regulated_drug>
    <is_fda_regulated_device>No</is_fda_regulated_device>
  </oversight_info>
  <brief_summary>
    <textblock>
      Background: Due to anatomical restrictions, the inflammatory response to intra-cerebral&#xD;
      bacterial infections exposes swollen brain tissues to pressure and ischemia, resulting in&#xD;
      life-threatening damage. However, diagnosing meningitis in patients after neurosurgery is&#xD;
      complicated, due to brain tissue damage and changes in cerebrospinal fluid (CSF) caused by&#xD;
      surgery. Hepatocyte growth factor (HGF) is a local, acute-phase protein. Previous studies on&#xD;
      community-acquired septic meningitis reported high levels of intrathecal-produced HGF.&#xD;
&#xD;
      Aim: The aim of present study is to evaluate a new platform for qualitative determination of&#xD;
      HGF in body fluids and revealing the site of injury.&#xD;
&#xD;
      Method: Based on a reverse-methachromacy method, strips are prepared. The surface on the&#xD;
      strip changes colour to blue upon contact with HGF.&#xD;
&#xD;
      Plan: CSF, urine and sputum of patients that develop fever post neurosurgery are analysed&#xD;
      with the test and the results compared with conventional diagnostic methods.&#xD;
&#xD;
      Clinical value: A rapid, equipment-free test gives the opportunity to identify the infectious&#xD;
      focus in the infected organ long before culture results are available.&#xD;
    </textblock>
  </brief_summary>
  <detailed_description>
    <textblock>
      Hepatocyte growth factor (HGF) is local acute phase protein with regenerative properties that&#xD;
      become biologic inactive during chronic inflammation and loses the binding affinity to&#xD;
      glycosaminoglycan in the extracellular ma-trix. HGF is excreted into the gastrointestinal&#xD;
      tract and is not detected in normal urine. Nosocomial meningitis can occur when brain&#xD;
      surgical procedures are complicated by infection. Due to underlying CNS disease processes and&#xD;
      CNS devices in situ, the principal agents of nosocomial bacterial meningitis differ from the&#xD;
      agents of community-acquired meningitis. For example, in nosocomial infections, slow growing,&#xD;
      opportunistic microorganisms predominate. The CSF leukocyte profile is affected by&#xD;
      intracerebral haemorrhage, and CSF lactate might be elevated, due to ischemia. Moreover,&#xD;
      altered consciousness can make it difficult to establish a diagnosis in patients on&#xD;
      ventilators that develop fever after neurosurgical operations. In other words, it is often&#xD;
      difficult to determine whether the injured brain has been invaded by environmental bacterial&#xD;
      flora.&#xD;
&#xD;
      Due to the challenges in establishing a diagnosis, and the lack of gold standards, physicians&#xD;
      are motivated to treat suspected infections in serious dis-eases with broad-spectrum&#xD;
      antibiotics. The emerging problems of multiple-resistance bacteria, high costs, and&#xD;
      complications related to new antibiotics have called for diagnostic tests that can minimize&#xD;
      antibiotic consumption.&#xD;
&#xD;
      Currently, the diagnosis of bacterial meningitis remains based on standard methods of direct&#xD;
      microscopy, differential analyses of white blood cells, lactate, and protein, and cultures of&#xD;
      blood and CSF . However, post-neurosurgical infections are difficult to distinguish from the&#xD;
      effects of neurosurgical procedures. Moreover, due to prophylaxis treatments, the cultures&#xD;
      are negative in a large group of patients, and the presence of skin flora, like Coagulase&#xD;
      negative Staphylococcus or Propionbacterium acnes, may indicate either infection or&#xD;
      contamination. Survival from this life-threatening condition depends on a rapid diagnosis and&#xD;
      prompt empirical antibiotic therapy designed to cover the likely pathogens.&#xD;
&#xD;
      Where is the focus of infection? Is it a bacterial (septic) meningitis? Is the broad-spectrum&#xD;
      antibiotic administration indicated? The background of the project: An invasion of bacteria&#xD;
      into the central nervous system (CNS) is followed by a rapidly evolving inflammatory process&#xD;
      that affects the arachnoid space, the pia mater, and the cerebrospinal fluid (CSF). This&#xD;
      condition leads to clinical symptoms of headache, fever, and meningism. The inflammatory&#xD;
      response is caused by the release of various pro-inflammatory cytokines from meningeal cells&#xD;
      into the subarachnoid space. As a result, neutrophils move into the subarachnoid space and&#xD;
      cause pleocytosis in the CSF. The consequences include the breakdown of the blood-brain&#xD;
      barrier, cerebral oedema, reduced cerebral blood flow, focal areas of hypo perfusion,&#xD;
      vascular thrombosis, ischemia, enhanced glucose metabolism via the anaerobic glycolytic&#xD;
      pathway, and enhanced lactate accumulation in the brain and CSF [1]. Survival from this&#xD;
      life-threatening condition depends on a rapid diagnosis and prompt empirical antibiotic&#xD;
      therapy designed to cover the likely pathogens.&#xD;
&#xD;
      Other causes of febrile meningitis include acute viral meningitis and non-pyogenic&#xD;
      meningitis, where the clinical picture is typically sub-acute or chronic. The diagnostic&#xD;
      procedures consist of a lumbar puncture to analyse the CSF for cells and bacteria,&#xD;
      microbiological cultures of blood and CSF, serological tests involving PCR and&#xD;
      antigen-detection, and radiographic techniques [2]. In community-acquired meningitis, a&#xD;
      combination of discriminating values from the CSF analysis can differentiate acute bacterial&#xD;
      meningitis from other, non-ambulatory causes with quite high sensitivity [3].&#xD;
&#xD;
      Nosocomial meningitis can occur when brain surgical procedures are complicated by infection&#xD;
      [2]. Due to underlying CNS disease processes and CNS devices in situ, the principal agents of&#xD;
      nosocomial bacterial meningitis differ from the agents of community-acquired meningitis. The&#xD;
      CSF leukocyte profile is affected by intra-cerebral haemorrhage, and CSF lactate might be&#xD;
      elevated, due to ischemia. Moreover, altered consciousness can make it difficult to establish&#xD;
      a diagnosis in patients on ventilators that develop fever after neurosurgical operations.&#xD;
&#xD;
      Hepatocyte growth factor (HGF) is produced by mesenchymal cells during organ injury. It is&#xD;
      produced as a single-chain precursor protein, and it is activated at the site of injury by&#xD;
      proteolysis cleavage, resulting in a double-chained active form of HGF [4]. Active HGF&#xD;
      stimulates cell division [4] and cell motility, and it promotes normal morphogenic structure&#xD;
      [5] in epithelial cells adjacent to injured areas. Thus, HGF induces regeneration and re-pair&#xD;
      of damaged tissue [6]. High levels of systemic HGF have been detected during injuries caused&#xD;
      by infection [7]. In bacterial meningitis, pneumonia and acute bacterial gastroenteritis,&#xD;
      there is local production of HGF at the site of infection [7-9]. HGF produced locally during&#xD;
      bacterial infection is biologically active [10]. Application of biologically active HGF&#xD;
      promoted healing of chronic leg ulcers in a pilot study [11]. Effective antibiotic therapy&#xD;
      reduces systemic HGF levels during infection [12-13]. HGF might be regarded as a local acute&#xD;
      phase protein with healing properties [14].&#xD;
&#xD;
      The quality of endogenous HGF binding to receptors can be assessed with surface plasmon&#xD;
      resonance (SPR), an optical technique appropriate for clinical studies that can determine the&#xD;
      affinity of a protein for several ligands [15]. SPR-based assessment of HGF binding affinity&#xD;
      for its receptors, c-Met and heparan sulphate proteoglycan (HSPG), could rapidly and&#xD;
      sensitively distinguish HGF variants with different biological activities [16-17].&#xD;
&#xD;
      We have previously studied the concentration of HGF in CSF for patients with community-&#xD;
      acquired meningitis [7]. We have also shown that the presence of biologically active HGF at&#xD;
      the site of injury indicates an acute local inflammation [18]. During a recent study we have&#xD;
      assessed whether HGF concentrations and HGF binding affinity for its receptors might serve as&#xD;
      markers to distinguish between meningitis associated with neurosurgery and other causes of&#xD;
      infection. We determined the concentration and binding affinity of HGF in patients with&#xD;
      either community-acquired meningitis or neurosurgery-associated meningitis, and compared the&#xD;
      results to either patients with Alzheimer's disease or controls with normal CSF. We have&#xD;
      shown that determination of HGF in CSF might be used as an indicator, complementary to&#xD;
      clinical status and routine laboratory findings, for diagnosing bacterial invasion into the&#xD;
      CSF at an early stage of disease [18].&#xD;
&#xD;
      The problem:&#xD;
&#xD;
      Case report: 76 years old woman with high blood pressure was found by her neighbours,&#xD;
      unconscious in her garden 28th October 2009. The CT scan revealed subarachnoid haemorrhage&#xD;
      and neuro-angiography revealed 3 aneurysms. She was admitted to the Department of&#xD;
      neurosurgery and operated 29th October with ventricular drainage and endovascular occlusion&#xD;
      of the aneurysms. She was extubated post-surgery however she suffered from left arm and leg&#xD;
      paralysis and tiredness. Since 30th October she had low grade of fever and slightly increased&#xD;
      CRP and procalcitonin. Cerebrospinal fluid showed 170.000 red and 630 white blood cells and&#xD;
      lactate 5.8. The chest X-ray showed a suspect infiltration. The blood and CSF cultures were&#xD;
      secured. The cultures taken from 5th November revealed significant growth of Enterobacter&#xD;
      cloacae and Staphylococcus aurous in urine and growth of Staphylococcus aurous in the&#xD;
      nasopharynx. She received no antibiotic therapy and the parameters decreased spontaneously&#xD;
      but low grade of fever continued. The CSF blood cells increased to 23000 red and 106 white&#xD;
      blood cells (11th November), and CSF lactate was 4.5 (lumbar drainage). Although CRP was in&#xD;
      the normal range she received meropenen 2g x 3 under suspicion bacterial meningitis. First on&#xD;
      19th November the CSF cultures taken on 11th November revealed growth of Propionibacterium&#xD;
      acnes. The antibiotic regime continued until 23th November. She suffered from disorientation,&#xD;
      fatigue, and had hydrocephalus. She underwent shunt operation 23th November. She died 2nd&#xD;
      December 2009. High concentration of HGF and elevated binding affinity to HSPG was found in&#xD;
      CSF analysis already 2nd November.&#xD;
&#xD;
      Due to the challenges in establishing a diagnosis, and the lack of gold standards, physicians&#xD;
      are motivated to treat suspected infections individually because there is no diagnostic&#xD;
      method that can identify the infections focus in time. PCR for bacterial panel and virus&#xD;
      detection in CSF is sensitive method. However it might detect the colonized bacteria as well.&#xD;
      The presence of bacteria is not the same as infection. Therefore in cases of bacterial&#xD;
      meningitis especially post brain surgery and the foreign bodies that are inserted in the&#xD;
      brain we cannot rely on positive or negative cultures totally. A complementary method (for&#xD;
      the standard available methods at laboratory) with high negative predictive value increases&#xD;
      the specificity of tests and decreases the costs, time and complications such as hospital&#xD;
      acquired infections (HAI) profoundly.&#xD;
&#xD;
      The Method, Reverse Methachromacy:&#xD;
&#xD;
      The N-terminus of HGF (Hairpin Loop) has been identified as the binding site to both c-Met&#xD;
      and HSPG. There is a high correlation between binding of HGF to HSPG and to dextran sulphate&#xD;
      in SPR system (fig 1). Metochromasia is a characteristic colour change that certain aniline&#xD;
      dyes exhibit when bound to chromo trope substances [19].This phenomenon has been widely used&#xD;
      in the study of tissue sections. Methylene blue (O-toluidine) is considered an excellent&#xD;
      metachromatic dye. Upon binding to high molecular weight polysaccharides, such as dextran&#xD;
      sulphate, the colour of the indicator solution changes from blue to red [19].&#xD;
&#xD;
      Combinations of dextran sulphate and O-toluidine in different proportions produce purple-red&#xD;
      coloured solutions of varying intensity. Coated customized filter papers make up surfaces&#xD;
      that are placed on strips. When in con-tact with biological solutions, the sample HGF&#xD;
      competitively replaces O-toluidine for binding to dextran sulphate and the colour of the&#xD;
      surface returns to blue (Fig 2). This platform detects the presence of HSPG-binding proteins&#xD;
      such as HGF in body fluids including expectorant, urine, ulcer secretions, cerebrospinal&#xD;
      fluid, and joint effusion during infection. High amounts of protein or high pH do not cause&#xD;
      non-specific reactions with this device.&#xD;
&#xD;
      The study plan:&#xD;
&#xD;
      Study leader: Amir Ramezani&#xD;
&#xD;
      The Procedure:&#xD;
&#xD;
      CSF specimens, urine and sputum (n=500) are collected from patients (1-99 years old) that&#xD;
      have undergone brain tumour surgery, intracerebral haemorrhage, shunt dysfunction or skull&#xD;
      fracture. The specimens are kept frozen in -20° C until analyses were performed. There are no&#xD;
      exclusion's criteria.&#xD;
&#xD;
      Controls Alzheimer's disease (n=20)&#xD;
&#xD;
        -  CSF samples were collected and kept frozen from patients (N=20) that were diagnosed with&#xD;
           Alzheimer's disease at the Memory Clinic, Skåne University Hospital in Malmö. The&#xD;
           revised DSM-III and NINDS-ADRDA criteria were used for diagnosis of Alzheimer's dementia&#xD;
           (this study group has been included in previous study).&#xD;
&#xD;
        -  Normal CSF&#xD;
&#xD;
        -  This group consists of patients who have undergone lumbar puncture to rule out&#xD;
           meningitis, and they had normal CSF.&#xD;
&#xD;
        -  Analysis of ligand-binding affinity with surface plasmon resonance&#xD;
&#xD;
        -  The biological activity of HGF is analysed with SPR. We measure the binding affinity to&#xD;
           HSPG (Sigma-Aldrich, St. Louis, MO, USA) and to a c-Met recombinant chimera (R&amp;D&#xD;
           Systems), as previously described.&#xD;
&#xD;
        -  Measurements of HGF concentrations in samples A specific enzyme-linked immunosorbent&#xD;
           assay (ELISA) kit (Quantikine Human HGF immunoassay, minimum detectable limit: 0.04&#xD;
           ng/mL; R&amp;D Systems, Minneapolis, MN, USA) was used to determine HGF concentrations in&#xD;
           CSF, sputum and urine according to the manufacturer's instructions. All measurements&#xD;
           were performed with an ELISA reader (Expert96; AsysHitech GmbH, Eugendorf, Austria) at&#xD;
           450 nm, which was calibrated with the recombinant human HGF reference samples and the&#xD;
           standards provided in the kit.&#xD;
&#xD;
        -  Reverse methachromacy The study strip test is performed on all specimens.&#xD;
&#xD;
        -  Routine laboratory assessments&#xD;
&#xD;
        -  CSF is analysed to determine parameter values at the Departments of Clinical Chemistry,&#xD;
           University Hospitals, in Linköping. All cultures, PCR assays, antigen detection assays,&#xD;
           and serological assessments are performed at the Department of Microbiology, University&#xD;
           Hospital, Linköping, Sweden.&#xD;
&#xD;
        -  CSF-cells: Performed by manual phase contrast microscopy (Zeiss) using Jessen Chamber&#xD;
           for counting the number of erythrocytes and leukocytes (polymorphonuclear neutrophils&#xD;
           and monocytes).&#xD;
&#xD;
        -  CSF lactate: Performed using benchtop blood gas analyzer ABL 800 (Radiometer Medical ApS&#xD;
           Denmark).&#xD;
&#xD;
        -  Antigen detection: Antigen (Streptococcus pneumonia, Haemophilus influenza, Neisseria&#xD;
           meningitides group A, group B/Escherichia coli K1, group C and group Y/135,&#xD;
           Streptococcus agalactiae) is detected by latex agglutination method Pastorex Meningitis™&#xD;
           (Bio-Rad, France).&#xD;
&#xD;
        -  CSF culture: Performed in aerobe and anaerobe flasks as well as in Hematin plates.&#xD;
&#xD;
        -  Virus detection PCR: Herpes virus type 1 and 2 (HSV1, HSV 2) and HZV DNA were detected&#xD;
           by PCR. Human enterovirus (HEV) was analysed by automated instrument (Cepheid&#xD;
           GeneXpert).&#xD;
&#xD;
        -  Statistical analysis&#xD;
&#xD;
        -  Because the HGF concentrations and binding affinity data are not normally distributed,&#xD;
           non parametric tests Kruskal-Wallis test followed by the Mann-Whitney U test or the&#xD;
           Wilcoxon matched pairs test are appropriate for analysis. The analysis of test&#xD;
           performance shall be performed manually.&#xD;
&#xD;
      Clinical Evaluation of the Platform in a Dextran Sulphate-Containing Gel Using reverse&#xD;
      methachromacy approach we have studied a total of 656 faeces samples with The sensitivity,&#xD;
      specificity, positive predictive value, and negative predictive value as well as the accuracy&#xD;
      were calculated in the following groups: verified infectious gastroenteritis (n=207) and&#xD;
      other causes of diarrhoea, including IBD (n=268). The test could distinguish acute infectious&#xD;
      gastroenteritis with a sensitivity of 96.6% and specificity 92.4%, and a positive predictive&#xD;
      value of 90.9% and negative predictive value of 97.2%. The accuracy of test was 94.3%. No&#xD;
      significant correlation between the test results and faeces calprotectin were found&#xD;
      (R2=0·056). No significant correlation was observed between the results of the index test and&#xD;
      the presence of blood in the faeces (R2=0·08) [20].&#xD;
&#xD;
      Clinical evaluation of HGF as a marker to distinguish septic meningitis from other causes of&#xD;
      pleocytosis [18] The community-acquired septic meningitis showed significantly higher HGF&#xD;
      concentration (p=0.0133), as well as HGF binding affinity to the c-Met and HSPG receptors&#xD;
      (p=0.0007 and p=0.0009, respectively) compared to nosocomial meningitis. CSF samples from&#xD;
      patients with septic meningitis (including both community-acquired and nosocomial) was&#xD;
      significantly higher in HGF concentrations (p=0.0014), HGF binding to HSPG (p&lt;0.0001), and&#xD;
      HGF binding to c-Met (p&lt;0.0001) compared to samples from patients with aseptic (viral and sub&#xD;
      acute) meningitis. CSF samples from patients with septic meningitis was higher from samples&#xD;
      from the control group (patients with normal CSF) and from patients with Alzheimer's disease&#xD;
      in HGF concentration (p&lt;0.0001, p=0.0010, respectively), HGF binding to HSPG (p&lt;0.0001 and&#xD;
      p=0.9, respectively), and HGF binding to c-Met (both p&lt;0.0001).&#xD;
&#xD;
      Compared to samples from patients that had undergone neurosurgery and had other infectious&#xD;
      diseases, CSF samples from patients with nosocomial meningitis had significantly higher HGF&#xD;
      concentrations (p&lt;0.0001) and HGF binding affinity to c-Met (p&lt;0.0001) and HSPG (p=0.043)&#xD;
      receptors (Figure 3-4).&#xD;
&#xD;
      Figure Legends Fig. 1. Sequence of part of the HGF α-chain. The enlarged codons code for&#xD;
      amino acids important for binding to HSPG and the c-Met receptor.&#xD;
&#xD;
      Fig. 2. The theoretical support for the second approach for a device to de-tect the presence&#xD;
      of HGF in body fluids during an infection. (1) Glucosaminoglycan (GAG) polymer chain with&#xD;
      negative charges. (2) Positive TBO molecules attach to the polymer chain then are&#xD;
      concentrated and form stacks. The colour changes from blue to red. (3) A sample with HGF is&#xD;
      added, which binds to GAG. (4) TBO molecule stacks are destroyed and become freely dissolved&#xD;
      TBO molecules. The colour changes from red to blue.&#xD;
&#xD;
      Figure 3: Flow chart of the selection of CSF specimens from patient and control groups. Extra&#xD;
      samples are the samples taken during stay on ward from patients with nosocomial meningitis.&#xD;
&#xD;
      Figure 4: Properties of HGF derived from different CSF samples were analysed by using surface&#xD;
      plasmon and ELISA techniques. (A) Binding to c-Met receptors; (B) Binding to HSPG receptors;&#xD;
      (C) HGF concentrations (median). *p&lt;0.05, **p&lt;0.01, ***p&lt;0.001, ****p&lt;0.0001.&#xD;
&#xD;
      The rapid evolution and spread of antimicrobial-resistant bacteria in parallel with&#xD;
      insufficient development of new active drugs seriously affect future anti-infective therapy&#xD;
      of bacterial infections, especially those due to Gram-negative rods. Experts in the field&#xD;
      estimate that by the next decade the world will have witnessed the wide dissemination of&#xD;
      untreatable (or next-to untreatable) infections, both within and beyond hospitals. To avoid&#xD;
      or at least attempt to retard this crisis, many researchers have dedicated their efforts to&#xD;
      elucidate factors favouring the emergence and global spread of antimicrobial resistant&#xD;
      bacteria. An effective strategy to limit the emergence of multiple resistant strains might&#xD;
      include directing the antibiotic therapy by identifying cases that should or should not be&#xD;
      treated with antibiotics, avoiding wide-spectrum antibiotic administration, and promoting&#xD;
      more specific treatments.&#xD;
&#xD;
      What is needed is a simple, accurate, cost-effective and feasible test that answers to&#xD;
      crucial questions.&#xD;
&#xD;
      In order to assess the clinical relevance and the performance of such a test there is not a&#xD;
      better environment than the Swedish healthcare that provides the relevant and modern&#xD;
      assessment of diseases and evidence-based therapy for nearly all patients. Such an&#xD;
      environment is an opportunity to perform the patient inclusion and evaluation of the study&#xD;
      test in comparison to the most reliable standard tools such as cultures, blood tests and&#xD;
      X-rays. These resources are not available automatically in any other centre with such&#xD;
      reliable, non-commercial and impartial properties.&#xD;
&#xD;
      dnr 2015-429-32&#xD;
    </textblock>
  </detailed_description>
  <overall_status>Unknown status</overall_status>
  <last_known_status>Recruiting</last_known_status>
  <start_date type="Actual">December 2, 2015</start_date>
  <completion_date type="Anticipated">December 1, 2020</completion_date>
  <primary_completion_date type="Anticipated">December 1, 2019</primary_completion_date>
  <study_type>Observational</study_type>
  <has_expanded_access>No</has_expanded_access>
  <study_design_info>
    <observational_model>Cohort</observational_model>
    <time_perspective>Prospective</time_perspective>
  </study_design_info>
  <primary_outcome>
    <measure>detection of infections in intensive care unit</measure>
    <time_frame>4 years</time_frame>
    <description>analyse of different body fluids to detect the focus of infections post neurosurgical fever</description>
  </primary_outcome>
  <enrollment type="Anticipated">150</enrollment>
  <condition>Infection, Bacterial</condition>
  <condition>Antibiotic Resistant Infection</condition>
  <condition>Diagnostic Self Evaluation</condition>
  <condition>Meningitis, Bacterial</condition>
  <condition>Nosocomial Infection</condition>
  <biospec_retention>Samples With DNA</biospec_retention>
  <biospec_descr>
    <textblock>
      Samples from CSF, Sputum, Urine, Blood&#xD;
    </textblock>
  </biospec_descr>
  <eligibility>
    <study_pop>
      <textblock>
        all post-neurosurgical patients in neuro intensive care unit university hospital Linkoping,&#xD;
        Sweden&#xD;
      </textblock>
    </study_pop>
    <sampling_method>Probability Sample</sampling_method>
    <criteria>
      <textblock>
        Inclusion Criteria:&#xD;
&#xD;
          -  all post neurosurgical patients with suspect infection or fever&#xD;
&#xD;
        Exclusion Criteria:&#xD;
      </textblock>
    </criteria>
    <gender>All</gender>
    <minimum_age>N/A</minimum_age>
    <maximum_age>N/A</maximum_age>
    <healthy_volunteers>Accepts Healthy Volunteers</healthy_volunteers>
  </eligibility>
  <overall_contact>
    <last_name>Fariba Nayeri, MD</last_name>
    <phone>+46702080804</phone>
    <email>fariba.nayeri@regionostergotland.se</email>
  </overall_contact>
  <overall_contact_backup>
    <last_name>Annette Theodorsson, Professor</last_name>
    <phone>+46734607538</phone>
    <email>annette.theodorsson@liu.se</email>
  </overall_contact_backup>
  <location>
    <facility>
      <name>Neurosurgery dept. University Hospital</name>
      <address>
        <city>Linkoping</city>
        <state>Ostergotland</state>
        <zip>581 85</zip>
        <country>Sweden</country>
      </address>
    </facility>
    <status>Recruiting</status>
    <contact>
      <last_name>Amir Ramezani, MD</last_name>
      <phone>+46705534710</phone>
      <email>amir.ramezani@regionostergotland.se</email>
    </contact>
  </location>
  <location>
    <facility>
      <name>Neurosurgery department, University Hospital</name>
      <address>
        <city>Linkoping</city>
        <state>Ostergotland</state>
        <zip>58185</zip>
        <country>Sweden</country>
      </address>
    </facility>
    <status>Recruiting</status>
    <contact>
      <last_name>Amir Ramezani, MD</last_name>
    </contact>
  </location>
  <location_countries>
    <country>Sweden</country>
  </location_countries>
  <verification_date>August 2017</verification_date>
  <study_first_submitted>August 13, 2017</study_first_submitted>
  <study_first_submitted_qc>August 13, 2017</study_first_submitted_qc>
  <study_first_posted type="Actual">August 17, 2017</study_first_posted>
  <last_update_submitted>August 18, 2017</last_update_submitted>
  <last_update_submitted_qc>August 18, 2017</last_update_submitted_qc>
  <last_update_posted type="Actual">August 21, 2017</last_update_posted>
  <responsible_party>
    <responsible_party_type>Sponsor-Investigator</responsible_party_type>
    <investigator_affiliation>University Hospital, Linkoeping</investigator_affiliation>
    <investigator_full_name>Fariba Nayeri</investigator_full_name>
    <investigator_title>MD, PhD, associated professor</investigator_title>
  </responsible_party>
  <keyword>Hepatocyte growth factor, Nosocomial infections, VAP, Meningitis, In vitro, point of care, rapid test</keyword>
  <condition_browse>
    <!-- CAUTION:  The following MeSH terms are assigned with an imperfect algorithm            -->
    <mesh_term>Infections</mesh_term>
    <mesh_term>Communicable Diseases</mesh_term>
    <mesh_term>Cross Infection</mesh_term>
    <mesh_term>Bacterial Infections</mesh_term>
    <mesh_term>Meningitis, Bacterial</mesh_term>
    <mesh_term>Meningitis</mesh_term>
  </condition_browse>
  <!-- Results have not yet been posted for this study                                          -->
</clinical_study>


download date: ClinicalTrials.gov processed this data on September 20, 2023
link text: Link to the current ClinicalTrials.gov record.
url: https://clinicaltrials.gov/ct2/show/NCT03252028
org study id: LINPEAS
nct id: NCT03252028
lead sponsor: 
has dmc: No
is fda regulated drug: No
is fda regulated device: No
Background: Due to anatomical restrictions, the inflammatory response to intra-cerebral
      bacterial infections exposes swollen brain tissues to pressure and ischemia, resulting in
      life-threatening damage. However, diagnosing meningitis in patients after neurosurgery is
      complicated, due to brain tissue damage and changes in cerebrospinal fluid (CSF) caused by
      surgery. Hepatocyte growth factor (HGF) is a local, acute-phase protein. Previous studies on
      community-acquired septic meningitis reported high levels of intrathecal-produced HGF.

      Aim: The aim of present study is to evaluate a new platform for qualitative determination of
      HGF in body fluids and revealing the site of injury.

      Method: Based on a reverse-methachromacy method, strips are prepared. The surface on the
      strip changes colour to blue upon contact with HGF.

      Plan: CSF, urine and sputum of patients that develop fever post neurosurgery are analysed
      with the test and the results compared with conventional diagnostic methods.

      Clinical value: A rapid, equipment-free test gives the opportunity to identify the infectious
      focus in the infected organ long before culture results are available.
Hepatocyte growth factor (HGF) is local acute phase protein with regenerative properties that
      become biologic inactive during chronic inflammation and loses the binding affinity to
      glycosaminoglycan in the extracellular ma-trix. HGF is excreted into the gastrointestinal
      tract and is not detected in normal urine. Nosocomial meningitis can occur when brain
      surgical procedures are complicated by infection. Due to underlying CNS disease processes and
      CNS devices in situ, the principal agents of nosocomial bacterial meningitis differ from the
      agents of community-acquired meningitis. For example, in nosocomial infections, slow growing,
      opportunistic microorganisms predominate. The CSF leukocyte profile is affected by
      intracerebral haemorrhage, and CSF lactate might be elevated, due to ischemia. Moreover,
      altered consciousness can make it difficult to establish a diagnosis in patients on
      ventilators that develop fever after neurosurgical operations. In other words, it is often
      difficult to determine whether the injured brain has been invaded by environmental bacterial
      flora.

      Due to the challenges in establishing a diagnosis, and the lack of gold standards, physicians
      are motivated to treat suspected infections in serious dis-eases with broad-spectrum
      antibiotics. The emerging problems of multiple-resistance bacteria, high costs, and
      complications related to new antibiotics have called for diagnostic tests that can minimize
      antibiotic consumption.

      Currently, the diagnosis of bacterial meningitis remains based on standard methods of direct
      microscopy, differential analyses of white blood cells, lactate, and protein, and cultures of
      blood and CSF . However, post-neurosurgical infections are difficult to distinguish from the
      effects of neurosurgical procedures. Moreover, due to prophylaxis treatments, the cultures
      are negative in a large group of patients, and the presence of skin flora, like Coagulase
      negative Staphylococcus or Propionbacterium acnes, may indicate either infection or
      contamination. Survival from this life-threatening condition depends on a rapid diagnosis and
      prompt empirical antibiotic therapy designed to cover the likely pathogens.

      Where is the focus of infection? Is it a bacterial (septic) meningitis? Is the broad-spectrum
      antibiotic administration indicated? The background of the project: An invasion of bacteria
      into the central nervous system (CNS) is followed by a rapidly evolving inflammatory process
      that affects the arachnoid space, the pia mater, and the cerebrospinal fluid (CSF). This
      condition leads to clinical symptoms of headache, fever, and meningism. The inflammatory
      response is caused by the release of various pro-inflammatory cytokines from meningeal cells
      into the subarachnoid space. As a result, neutrophils move into the subarachnoid space and
      cause pleocytosis in the CSF. The consequences include the breakdown of the blood-brain
      barrier, cerebral oedema, reduced cerebral blood flow, focal areas of hypo perfusion,
      vascular thrombosis, ischemia, enhanced glucose metabolism via the anaerobic glycolytic
      pathway, and enhanced lactate accumulation in the brain and CSF [1]. Survival from this
      life-threatening condition depends on a rapid diagnosis and prompt empirical antibiotic
      therapy designed to cover the likely pathogens.

      Other causes of febrile meningitis include acute viral meningitis and non-pyogenic
      meningitis, where the clinical picture is typically sub-acute or chronic. The diagnostic
      procedures consist of a lumbar puncture to analyse the CSF for cells and bacteria,
      microbiological cultures of blood and CSF, serological tests involving PCR and
      antigen-detection, and radiographic techniques [2]. In community-acquired meningitis, a
      combination of discriminating values from the CSF analysis can differentiate acute bacterial
      meningitis from other, non-ambulatory causes with quite high sensitivity [3].

      Nosocomial meningitis can occur when brain surgical procedures are complicated by infection
      [2]. Due to underlying CNS disease processes and CNS devices in situ, the principal agents of
      nosocomial bacterial meningitis differ from the agents of community-acquired meningitis. The
      CSF leukocyte profile is affected by intra-cerebral haemorrhage, and CSF lactate might be
      elevated, due to ischemia. Moreover, altered consciousness can make it difficult to establish
      a diagnosis in patients on ventilators that develop fever after neurosurgical operations.

      Hepatocyte growth factor (HGF) is produced by mesenchymal cells during organ injury. It is
      produced as a single-chain precursor protein, and it is activated at the site of injury by
      proteolysis cleavage, resulting in a double-chained active form of HGF [4]. Active HGF
      stimulates cell division [4] and cell motility, and it promotes normal morphogenic structure
      [5] in epithelial cells adjacent to injured areas. Thus, HGF induces regeneration and re-pair
      of damaged tissue [6]. High levels of systemic HGF have been detected during injuries caused
      by infection [7]. In bacterial meningitis, pneumonia and acute bacterial gastroenteritis,
      there is local production of HGF at the site of infection [7-9]. HGF produced locally during
      bacterial infection is biologically active [10]. Application of biologically active HGF
      promoted healing of chronic leg ulcers in a pilot study [11]. Effective antibiotic therapy
      reduces systemic HGF levels during infection [12-13]. HGF might be regarded as a local acute
      phase protein with healing properties [14].

      The quality of endogenous HGF binding to receptors can be assessed with surface plasmon
      resonance (SPR), an optical technique appropriate for clinical studies that can determine the
      affinity of a protein for several ligands [15]. SPR-based assessment of HGF binding affinity
      for its receptors, c-Met and heparan sulphate proteoglycan (HSPG), could rapidly and
      sensitively distinguish HGF variants with different biological activities [16-17].

      We have previously studied the concentration of HGF in CSF for patients with community-
      acquired meningitis [7]. We have also shown that the presence of biologically active HGF at
      the site of injury indicates an acute local inflammation [18]. During a recent study we have
      assessed whether HGF concentrations and HGF binding affinity for its receptors might serve as
      markers to distinguish between meningitis associated with neurosurgery and other causes of
      infection. We determined the concentration and binding affinity of HGF in patients with
      either community-acquired meningitis or neurosurgery-associated meningitis, and compared the
      results to either patients with Alzheimer's disease or controls with normal CSF. We have
      shown that determination of HGF in CSF might be used as an indicator, complementary to
      clinical status and routine laboratory findings, for diagnosing bacterial invasion into the
      CSF at an early stage of disease [18].

      The problem:

      Case report: 76 years old woman with high blood pressure was found by her neighbours,
      unconscious in her garden 28th October 2009. The CT scan revealed subarachnoid haemorrhage
      and neuro-angiography revealed 3 aneurysms. She was admitted to the Department of
      neurosurgery and operated 29th October with ventricular drainage and endovascular occlusion
      of the aneurysms. She was extubated post-surgery however she suffered from left arm and leg
      paralysis and tiredness. Since 30th October she had low grade of fever and slightly increased
      CRP and procalcitonin. Cerebrospinal fluid showed 170.000 red and 630 white blood cells and
      lactate 5.8. The chest X-ray showed a suspect infiltration. The blood and CSF cultures were
      secured. The cultures taken from 5th November revealed significant growth of Enterobacter
      cloacae and Staphylococcus aurous in urine and growth of Staphylococcus aurous in the
      nasopharynx. She received no antibiotic therapy and the parameters decreased spontaneously
      but low grade of fever continued. The CSF blood cells increased to 23000 red and 106 white
      blood cells (11th November), and CSF lactate was 4.5 (lumbar drainage). Although CRP was in
      the normal range she received meropenen 2g x 3 under suspicion bacterial meningitis. First on
      19th November the CSF cultures taken on 11th November revealed growth of Propionibacterium
      acnes. The antibiotic regime continued until 23th November. She suffered from disorientation,
      fatigue, and had hydrocephalus. She underwent shunt operation 23th November. She died 2nd
      December 2009. High concentration of HGF and elevated binding affinity to HSPG was found in
      CSF analysis already 2nd November.

      Due to the challenges in establishing a diagnosis, and the lack of gold standards, physicians
      are motivated to treat suspected infections individually because there is no diagnostic
      method that can identify the infections focus in time. PCR for bacterial panel and virus
      detection in CSF is sensitive method. However it might detect the colonized bacteria as well.
      The presence of bacteria is not the same as infection. Therefore in cases of bacterial
      meningitis especially post brain surgery and the foreign bodies that are inserted in the
      brain we cannot rely on positive or negative cultures totally. A complementary method (for
      the standard available methods at laboratory) with high negative predictive value increases
      the specificity of tests and decreases the costs, time and complications such as hospital
      acquired infections (HAI) profoundly.

      The Method, Reverse Methachromacy:

      The N-terminus of HGF (Hairpin Loop) has been identified as the binding site to both c-Met
      and HSPG. There is a high correlation between binding of HGF to HSPG and to dextran sulphate
      in SPR system (fig 1). Metochromasia is a characteristic colour change that certain aniline
      dyes exhibit when bound to chromo trope substances [19].This phenomenon has been widely used
      in the study of tissue sections. Methylene blue (O-toluidine) is considered an excellent
      metachromatic dye. Upon binding to high molecular weight polysaccharides, such as dextran
      sulphate, the colour of the indicator solution changes from blue to red [19].

      Combinations of dextran sulphate and O-toluidine in different proportions produce purple-red
      coloured solutions of varying intensity. Coated customized filter papers make up surfaces
      that are placed on strips. When in con-tact with biological solutions, the sample HGF
      competitively replaces O-toluidine for binding to dextran sulphate and the colour of the
      surface returns to blue (Fig 2). This platform detects the presence of HSPG-binding proteins
      such as HGF in body fluids including expectorant, urine, ulcer secretions, cerebrospinal
      fluid, and joint effusion during infection. High amounts of protein or high pH do not cause
      non-specific reactions with this device.

      The study plan:

      Study leader: Amir Ramezani

      The Procedure:

      CSF specimens, urine and sputum (n=500) are collected from patients (1-99 years old) that
      have undergone brain tumour surgery, intracerebral haemorrhage, shunt dysfunction or skull
      fracture. The specimens are kept frozen in -20° C until analyses were performed. There are no
      exclusion's criteria.

      Controls Alzheimer's disease (n=20)

        -  CSF samples were collected and kept frozen from patients (N=20) that were diagnosed with
           Alzheimer's disease at the Memory Clinic, Skåne University Hospital in Malmö. The
           revised DSM-III and NINDS-ADRDA criteria were used for diagnosis of Alzheimer's dementia
           (this study group has been included in previous study).

        -  Normal CSF

        -  This group consists of patients who have undergone lumbar puncture to rule out
           meningitis, and they had normal CSF.

        -  Analysis of ligand-binding affinity with surface plasmon resonance

        -  The biological activity of HGF is analysed with SPR. We measure the binding affinity to
           HSPG (Sigma-Aldrich, St. Louis, MO, USA) and to a c-Met recombinant chimera (R&D
           Systems), as previously described.

        -  Measurements of HGF concentrations in samples A specific enzyme-linked immunosorbent
           assay (ELISA) kit (Quantikine Human HGF immunoassay, minimum detectable limit: 0.04
           ng/mL; R&D Systems, Minneapolis, MN, USA) was used to determine HGF concentrations in
           CSF, sputum and urine according to the manufacturer's instructions. All measurements
           were performed with an ELISA reader (Expert96; AsysHitech GmbH, Eugendorf, Austria) at
           450 nm, which was calibrated with the recombinant human HGF reference samples and the
           standards provided in the kit.

        -  Reverse methachromacy The study strip test is performed on all specimens.

        -  Routine laboratory assessments

        -  CSF is analysed to determine parameter values at the Departments of Clinical Chemistry,
           University Hospitals, in Linköping. All cultures, PCR assays, antigen detection assays,
           and serological assessments are performed at the Department of Microbiology, University
           Hospital, Linköping, Sweden.

        -  CSF-cells: Performed by manual phase contrast microscopy (Zeiss) using Jessen Chamber
           for counting the number of erythrocytes and leukocytes (polymorphonuclear neutrophils
           and monocytes).

        -  CSF lactate: Performed using benchtop blood gas analyzer ABL 800 (Radiometer Medical ApS
           Denmark).

        -  Antigen detection: Antigen (Streptococcus pneumonia, Haemophilus influenza, Neisseria
           meningitides group A, group B/Escherichia coli K1, group C and group Y/135,
           Streptococcus agalactiae) is detected by latex agglutination method Pastorex Meningitis™
           (Bio-Rad, France).

        -  CSF culture: Performed in aerobe and anaerobe flasks as well as in Hematin plates.

        -  Virus detection PCR: Herpes virus type 1 and 2 (HSV1, HSV 2) and HZV DNA were detected
           by PCR. Human enterovirus (HEV) was analysed by automated instrument (Cepheid
           GeneXpert).

        -  Statistical analysis

        -  Because the HGF concentrations and binding affinity data are not normally distributed,
           non parametric tests Kruskal-Wallis test followed by the Mann-Whitney U test or the
           Wilcoxon matched pairs test are appropriate for analysis. The analysis of test
           performance shall be performed manually.

      Clinical Evaluation of the Platform in a Dextran Sulphate-Containing Gel Using reverse
      methachromacy approach we have studied a total of 656 faeces samples with The sensitivity,
      specificity, positive predictive value, and negative predictive value as well as the accuracy
      were calculated in the following groups: verified infectious gastroenteritis (n=207) and
      other causes of diarrhoea, including IBD (n=268). The test could distinguish acute infectious
      gastroenteritis with a sensitivity of 96.6% and specificity 92.4%, and a positive predictive
      value of 90.9% and negative predictive value of 97.2%. The accuracy of test was 94.3%. No
      significant correlation between the test results and faeces calprotectin were found
      (R2=0·056). No significant correlation was observed between the results of the index test and
      the presence of blood in the faeces (R2=0·08) [20].

      Clinical evaluation of HGF as a marker to distinguish septic meningitis from other causes of
      pleocytosis [18] The community-acquired septic meningitis showed significantly higher HGF
      concentration (p=0.0133), as well as HGF binding affinity to the c-Met and HSPG receptors
      (p=0.0007 and p=0.0009, respectively) compared to nosocomial meningitis. CSF samples from
      patients with septic meningitis (including both community-acquired and nosocomial) was
      significantly higher in HGF concentrations (p=0.0014), HGF binding to HSPG (p<0.0001), and
      HGF binding to c-Met (p<0.0001) compared to samples from patients with aseptic (viral and sub
      acute) meningitis. CSF samples from patients with septic meningitis was higher from samples
      from the control group (patients with normal CSF) and from patients with Alzheimer's disease
      in HGF concentration (p<0.0001, p=0.0010, respectively), HGF binding to HSPG (p<0.0001 and
      p=0.9, respectively), and HGF binding to c-Met (both p<0.0001).

      Compared to samples from patients that had undergone neurosurgery and had other infectious
      diseases, CSF samples from patients with nosocomial meningitis had significantly higher HGF
      concentrations (p<0.0001) and HGF binding affinity to c-Met (p<0.0001) and HSPG (p=0.043)
      receptors (Figure 3-4).

      Figure Legends Fig. 1. Sequence of part of the HGF α-chain. The enlarged codons code for
      amino acids important for binding to HSPG and the c-Met receptor.

      Fig. 2. The theoretical support for the second approach for a device to de-tect the presence
      of HGF in body fluids during an infection. (1) Glucosaminoglycan (GAG) polymer chain with
      negative charges. (2) Positive TBO molecules attach to the polymer chain then are
      concentrated and form stacks. The colour changes from blue to red. (3) A sample with HGF is
      added, which binds to GAG. (4) TBO molecule stacks are destroyed and become freely dissolved
      TBO molecules. The colour changes from red to blue.

      Figure 3: Flow chart of the selection of CSF specimens from patient and control groups. Extra
      samples are the samples taken during stay on ward from patients with nosocomial meningitis.

      Figure 4: Properties of HGF derived from different CSF samples were analysed by using surface
      plasmon and ELISA techniques. (A) Binding to c-Met receptors; (B) Binding to HSPG receptors;
      (C) HGF concentrations (median). *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001.

      The rapid evolution and spread of antimicrobial-resistant bacteria in parallel with
      insufficient development of new active drugs seriously affect future anti-infective therapy
      of bacterial infections, especially those due to Gram-negative rods. Experts in the field
      estimate that by the next decade the world will have witnessed the wide dissemination of
      untreatable (or next-to untreatable) infections, both within and beyond hospitals. To avoid
      or at least attempt to retard this crisis, many researchers have dedicated their efforts to
      elucidate factors favouring the emergence and global spread of antimicrobial resistant
      bacteria. An effective strategy to limit the emergence of multiple resistant strains might
      include directing the antibiotic therapy by identifying cases that should or should not be
      treated with antibiotics, avoiding wide-spectrum antibiotic administration, and promoting
      more specific treatments.

      What is needed is a simple, accurate, cost-effective and feasible test that answers to
      crucial questions.

      In order to assess the clinical relevance and the performance of such a test there is not a
      better environment than the Swedish healthcare that provides the relevant and modern
      assessment of diseases and evidence-based therapy for nearly all patients. Such an
      environment is an opportunity to perform the patient inclusion and evaluation of the study
      test in comparison to the most reliable standard tools such as cultures, blood tests and
      X-rays. These resources are not available automatically in any other centre with such
      reliable, non-commercial and impartial properties.

      dnr 2015-429-32
observational model: Cohort
time perspective: Prospective
measure: detection of infections in intensive care unit
time frame: 4 years
description: analyse of different body fluids to detect the focus of infections post neurosurgical fever
Samples from CSF, Sputum, Urine, Blood
study pop: 
sampling method: Probability Sample
criteria: 
gender: All
minimum age: N/A
maximum age: N/A
healthy volunteers: Accepts Healthy Volunteers
last name: Fariba Nayeri, MD
phone: +46702080804
email: fariba.nayeri@regionostergotland.se
last name: Annette Theodorsson, Professor
phone: +46734607538
email: annette.theodorsson@liu.se
facility: 
status: Recruiting
contact: 
facility: 
status: Recruiting
contact: 
country: Sweden
responsible party type: Sponsor-Investigator
investigator affiliation: University Hospital, Linkoeping
investigator full name: Fariba Nayeri
investigator title: MD, PhD, associated professor
mesh term: Infections
mesh term: Communicable Diseases
mesh term: Cross Infection
mesh term: Bacterial Infections
mesh term: Meningitis, Bacterial
mesh term: Meningitis