Consecutive Cerebrospinal Fluid Samples Research Articles

Globicatella sanguinis meningitis in a post head trauma patient: first case report from Asia

Globicatella sanguinis is a rare isolate in clinical samples. We present a case of meningitis in a 70-year-old male patient after a head injury operation. Three consecutive cerebrospinal fluid samples obtained from the patient identified Globicatella sanguinis based on morphology, biochemical profile, and Vitek-2 identification. The patient recovered after antibiotic treatment with vancomycin. This is the first case report of Globicatella sanguinis from Asia from a case of meningitis.

Differentiating patterns of oligoclonal banding in the cerebrospinal fluid improves diagnostic utility for multiple sclerosis

Detection of oligoclonal bands (OCBs) in the cerebrospinal fluid (CSF) is an important adjunct to the diagnosis of multiple sclerosis (MS) and clinically isolated syndromes (CIS). OCBs are considered present if two or more extra IgG bands are present in CSF in comparison to a simultaneously collected serum sample. However, using isoelectric focusing and immunofixation with anti-IgG, we observed two distinct banding patterns, one in which the bands were numerous and prominent (which we termed 'delta') and a much more subtle pattern, with fewer, more indistinct bands ('theta'). To perform a prospective study to determine the diagnostic implications of the two OCB patterns for multiple sclerosis. Over a 2-year period, 56 consecutive CSF samples with OCBs were identified. Clinical information and radiological data were collected and correlated with the two banding patterns. : Of the 56 positive CSF samples, 46 (82%) demonstrated a delta pattern, and 10 (18%), a theta pattern. The delta pattern was associated with MS/CIS in 34 of 46 (74%) samples, compared with zero of the 10 theta samples (0%, p < 0.001). Exclusion of the theta pattern samples increased the positive predictive value of OCB testing from 61% to 74% for MS/CIS. The diagnostic significance of oligoclonal bands in CSF for MS/CIS can be improved by restricting interpretation to the delta banding pattern alone.

Performance Evaluation of the Sysmex XE-5000 Hematology Analyzer for White Blood Cell Analysis in Cerebrospinal Fluid

The newest generation hematology analyzer, Sysmex XE-5000 (Sysmex Corporation, Kobe, Japan) is equipped with an improved body fluid analysis mode. To evaluate the applicability of the XE-5000 analyzer to white blood cell (WBC) analysis in cerebrospinal fluid (CSF). A total of 425 routinely collected, consecutive CSF samples were included in the study. For a comparison of total WBC counts, the results of routine chamber counts were grouped into categories of 0 to 5 (n = 330), >5 to 10 (n = 36), >10 to 50 (n = 39), >50 to 200 (n = 15), and >200 (n = 5) WBC/µL. Microscopic differential counts were performed using cytospins from 276 samples. Results were grouped according to the percent content of polymorphonuclear (PMN) cells, 0% to 25% (n = 263), >25% to 50% (n = 7), >50% to 75% (n = 3), and >75% to 100% (n = 3) of WBC. Corresponding results of XE-5000 analysis were matched to these particular count categories. For total WBC counts, the proportions of samples correctly classified by the XE-5000 from the percentage groups described above were 88%, 47%, 72%, 93%, and 100%, respectively. After the two lowest count categories were combined into one range of 0 to 10 WBC/µL, matches increased to 95%. For PMN counts in the 0% to 25% group, 37% of samples were misclassified by the XE-5000. Conversely, for samples with microscopic PMN counts of more than 25%, there was a trend toward underestimation by the XE-5000. Mismatches were most pronounced in samples with fewer than 10 WBC/µL. The Sysmex XE-5000 hematology analyzer yields valid total CSF cell counts and may be considered an acceptable alternative to the traditional chamber method, even for samples with low WBC counts. However, it cannot be recommended as a suitable alternative for manual differential cytologic workup.

Sporobolomyces roseus in the cerebrospinal fluid of an immunocompetent patient – to treat or not to treat?

We present the case of an immunocompetent male who presented with symptoms of meningitis. Yeasts were seen in two consecutive cerebrospinal fluid samples, which were identified by PCR as Sporobolomyces roseus. This yeast is rarely encountered in clinical settings, and has only previously been seen to cause infection in immunocompromised patients. This case highlights the challenges presented by the identification of an unusual pathogen in an unexpected clinical setting.

Neutrophils in cerebrospinal fluid without pleocytosis

Neutrophils in cerebrospinal fluid (CSF) samples are commonly considered a pathological feature; however, there is little information on the frequency and significance of these cells in CSF samples without pleocytosis. Therefore, the frequency and possible clinical significance of neutrophils in CSF was investigated. In a retrospective study comprising 1556 consecutive CSF samples, cytologies and patient records were reviewed. Five hundred thirty-eight CSF samples without pleocytosis were identified. Neutrophils were detected in 35.5% of these samples. The presence of neutrophils was associated with sepsis (P < 0.01), recent epileptic seizure (P < 0.0001), and blood contamination (P < 0.01). Amongst patients without CSF pleocytosis, CNS infections were not more frequent if neutrophils were present. Neutrophils are frequently observed in CSF with normal leukocyte counts. As sepsis but not CNS infection occurred more frequently in these patients, we conclude that in the absence of CSF pleocytosis, neutrophils are not indicative of CNS infections.

Evaluation of the MODS Culture Technique for the Diagnosis of Tuberculous Meningitis

BackgroundTuberculous meningitis (TBM) is a devastating condition. The rapid instigation of appropraite chemotherapy is vital to reduce morbidity and mortality. However rapid diagnosis remains elusive; smear microscopy has extremely low sensitivity on cerebrospinal fluid (CSF) in most laboratories and PCR requires expertise with advanced infrastructure and has sensitivity of only around 60% under optimal conditions. Neither technique allows for the microbiological isolation of M. tuberculosis and subsequent drug susceptibility testing. We evaluated the recently developed microscopic observation drug susceptibility (MODS) assay format for speed and accuracy in diagnosing TBM.Methodology/Principal FindingsTwo hundred and thirty consecutive CSF samples collected from 156 patients clinically suspected of TBM on presentation at a tertiary referal hospital in Vietnam were enrolled into the study over a five month period and tested by Ziehl-Neelsen (ZN) smear, MODS, Mycobacterial growth Indicator tube (MGIT) and Lowenstein-Jensen (LJ) culture. Sixty-one samples were from patients already on TB therapy for >1day and 19 samples were excluded due to untraceable patient records. One hundred and fifty samples from 137 newly presenting patients remained. Forty-two percent (n = 57/137) of patients were deemed to have TBM by clinical diagnostic and microbiological criteria (excluding MODS). Sensitivity by patient against clinical gold standard for ZN smear, MODS MGIT and LJ were 52.6%, 64.9%, 70.2% and 70.2%, respectively. Specificity of all microbiological techniques was 100%. Positive and negative predictive values for MODS were 100% and 78.7%, respectively for HIV infected patients and 100% and 82.1% for HIV negative patients. The median time to positive was 6 days (interquartile range 5–7), significantly faster than MGIT at 15.5 days (interquartile range 12–24), and LJ at 24 days (interquartile range 18–35 days) (P<0.01).ConclusionsWe have shown MODS to be a sensitive, rapid technique for the diagnosis of TBM with high sensitivity, ease of performance and low cost (0.53 USD/sample).

A screening assay to detect antigen-specific antibodies within cerebrospinal fluid

Identification of the aetiology of central nervous system infections requires the detection of either the organism or a microbe-specific immune response within the brain or cerebrospinal fluid. We describe a screening assay to detect herpes simplex virus, varicella zoster virus, cytomegalovirus, measles and Toxoplasma gondii specific antibodies in cerebrospinal fluid. Antigen-specific immunoblotting of oligoclonal IgG and IgM was used to confirm the presence of antibody. Of 51 consecutive cerebrospinal fluid samples received by the laboratory from patients with suspected central nervous system infection 18 (35%) were screen positive for one or more antigen. In only 7 of these were antigen-specific oligoclonal IgG or IgM bands confirmed. The assay provides a simple, cheap assay to screen for microbial-specific antibody in the cerebrospinal fluid samples of patients with suspected neurological infections.

Modulation of sFas indicates apoptosis in human herpes simplex encephalitis

Herpes simplex encephalitis (HSE) is the most common cause of non-epidemic, acute and fatal viral encephalitis. A pronounced mortality and morbidity remains in HSE despite antiviral treatment. There is evidence of a vigorous intrathecal immune activity in acute phases of HSE and of persistently increased activity at follow-ups after years. The role of apoptosis of neuronal cells in HSE patients as a mechanism of damage has been brought up lately. We hypothesize that the severity and the progression of the cerebral injury resulting from HSE can be evaluated by quantitative measurement of a compartment of immune activation molecules i.e. soluble Fas (sFas) involved in apoptosis through the Fas/Fas Ligand pathway. Consecutive cerebrospinal fluid (CSF) samples from a prospectively followed cohort, included in an antiviral treatment trial in HSE, were enrolled for quantitative measurement of sFas using commercial capture ELISA. In total, CSF samples from 49 patients with HSE, 63 patients with non-HSE encephalitis and 18 healthy individuals were studied. High levels of sFas were expressed in CSF samples collected between days 0–45 after neurological onset in 41 / 49 (84%) HSE patients, whereas only 21 / 63 (33%) of non-HSE patients and none of 18 healthy controls demonstrated measurable levels of sFas. Following the consecutive CSF sFas levels over the time and considering the clinical state of patients at admission, their neurological or lethal outcome at 12 months, and antiviral treatment, we observed that HSE patients with severe neurological sequels revealed an increase in changes of CSF sFas as compared to patients with mild or moderate neurological outcome (57.6 ± 55.6 pg/ml, n = 10 versus 26.3 ± 97.5 pg/ml, n = 14; P = 0.008). Also HSE patients undergoing vidarabine treatment expressed significantly higher levels of changes of CSF sFas when compared to acyclovir-treated patients (63.7 ± 52.8 pg/ml, n = 9 versus 26.1 ± 98.4 pg/ml, n = 14; P = 0.003). Interestingly, regardless of the clinical state at admission, and subsequent disease progression of the HSE patients, we could not observe any significant differences in the CSF sFas levels during the first 7 days of neurological symptoms. These observations underline the role of immunological response throughout the course of HSV infection in the brain and the role of the Fas/FasL pathway in particular in disease progression of HSE. The findings further enforce the need of expanding the knowledge of the pathogenesis of HSE to direct to more effective, in particular not only antiviral but also anti-apoptotic or anti-inflammatory treatment.

Delayed craniospinal irradiation for a first isolated central nervous relapse of acute lymphoblastic leukemia: report on 14 cases.

Children developing an isolated central nervous system (CNS) relapse as first recurrence of their acute lymphoblastic leukemia (ALL) are considered to have a systemic relapse as well. They are mostly treated with intensive chemotherapy and craniospinal irradiation. In most treatment schedules, irradiation is given early after induction treatment. Because craniospinal irradiation affects a large portion of hematopoietic bone marrow systemically, treatment is often delayed owing to aplasias. Also, dose reductions are frequently needed. Children receiving simultaneously irradiation and chemotherapy are prone to (often severe) neurotoxicity. This study reports on children with a first isolated CNS relapse of their ALL receiving chemotherapy for 40 weeks. Treatment ends with the administration of irradiation given after cessation of chemotherapy. Fourteen children, with blasts and >5 cells/mm(3) in two consecutive samples of cerebrospinal fluid and a blast percentage <5% in their bone marrow were treated according to an intensive systemic and site-specific chemotherapy. Craniospinal irradiation was administered after cessation chemotherapy. Event-free-survival was 57% (confidence interval 35-89%), freedom from relapse was 61.5%; follow-up ranges from 2.0 to 15.1 years (median 11.7 years). One child died from septicemia during induction. Five children experienced a second relapse and died from their malignancy. Two children [with a t(9;22) or a rearranged MLL gene] relapsed prior to radiotherapy. Outcome was related to duration of first remission, age at relapse, and identification as a high-risk patient at initial diagnosis. No neurologic complications were noted during and after treatment. Delayed irradiation for isolated CNS relapse in children with ALL gives favorable survival rates, without significant toxicity. Neurotoxicity was absent.

Laboratory diagnosis of common viral infections of the central nervous system by using a single multiplex PCR screening assay.

A multiplex PCR assay that detects the four commonest causes of viral meningitis and encephalitis in the United Kingdom (herpes simplex virus [HSV] type 1 [HSV-1], HSV type 2 [HSV-2], varicella-zoster virus [VZV], and enteroviruses) was developed, and its sensitivity was compared with those of similar assays described previously for this application. Compared to the previous assays, this single multiplex PCR assay had higher molecular sensitivities for the detection for each of the viruses and improved utility for routine use in a diagnostic laboratory. The assay was used to test a series of 1,683 consecutive cerebrospinal fluid (CSF) samples between June 1997 and March 1998 inclusively. Viral nucleic acid was detected in 138 (8.2%) of the CSF samples, including enteroviruses in 51 samples, HSV-2 in 33 samples, VZV in 28 samples, and HSV-1 in 25 samples. Compared to the accepted relative incidence of viral etiologies, aseptic meningitis due to HSV-2 infection was high, and in adult female patients with symptoms of aseptic meningitis, HSV-2 was the virus most commonly detected in the CSF.

Elevated levels of antibody to myelin oligodendrocyte glycoprotein is not specific for patients with multiple sclerosis.

To evaluate the presence and specificity of anti-myelin oligodendrocyte glycoprotein (MOG) antibody in the cerebrospinal fluid and plasma of patients with multiple sclerosis (MS). Case-control study of patients with clinically definite MS compared with patients with other neurologic diseases (ONDs) of the central nervous system and control subjects. Referral center in the Department of Neurology of Hadassah University Hospital, greater Jerusalem area, Israel. Consecutive cerebrospinal fluid samples from 31 patients with MS, 31 patients with ONDs, and 28 healthy controls; and plasma samples from 33 patients with MS, 28 patients with ONDs, and 31 healthy controls were taken from the cerebrospinal fluid and plasma bank of the Department of Neurology, Hadassah University Hospital. Levels and frequencies of anti-MOG antibody in patients with MS, as defined by enzyme-linked immunosorbent assay. Cerebrospinal fluid levels of antibodies to MOG and to myelin basic protein were significantly higher in patients with MS (P<.001 and P = .001, respectively) and patients with ONDs (P = .005 and P = .03, respectively) compared with controls; frequency of antibodies to MOG, but not to myelin basic protein, was higher in patients with MS and patients with ONDs (P = .01 and P = .003, respectively, for the frequency of anti-MOG antibody, and P = .65 and P = .41, respectively, for the frequency of anti-myelin basic protein antibody). Plasma levels of antibodies to MOG and to myelin basic protein were higher in patients with MS compared with patients with ONDs (P = .003 for both comparisons) and with controls (P = .03 and P = .04, respectively); however, the frequency of antibodies to MOG and myelin basic protein was similar in patients with MS, patients with ONDs (P=.54 and P = .82, respectively), and controls (P = .50 and P = .14, respectively). The elevated presence of anti-MOG antibody is not specific for MS because a similar appearance was also demonstrated in patients with ONDs. Therefore, it is not clear whether this antibody is pathogenic in MS or, on the contrary, has a defensive role against further immune-mediated damage after myelin breakdown.

Diagnosis of viral infections of the central nervous system: clinical interpretation of PCR results

Standard laboratory techniques, such as viral culture and serology, provide only circumstantial or retrospective evidence of viral infections of the central nervous system (CNS). We assessed the diagnostic accuracy of PCR of cerebrospinal fluid (CSF) in the diagnosis of viral infections of the CNS. We examined all the CSF samples that were received at our diagnostic virology laboratory between May, 1994, and May, 1996, by nested PCR for viruses associated with CNS infections in the UK. We collected clinical and laboratory data for 410 patients from Oxford city hospitals (the Oxford cohort) whose CSF was examined between May, 1994, and May, 1995. These patients were classified according to the likelihood of a viral infection of the CNS. We used stratified logistic regression analysis to identify the clinical factors independently associated with a positive PCR result. We calculated likelihood ratios to estimate the clinical usefulness of PCR amplification of CSF. We tested 2233 consecutive CSF samples from 2162 patients. A positive PCR result was obtained in 143 patients, including 22 from the Oxford cohort. Logistic regression analysis of the Oxford cohort showed that fever, a virus-specific rash, and a CSF white-cell count of 5/microL or more were independent predictors of a positive PCR result. The likelihood ratio for a definite diagnosis of viral infection of the CNS in a patient with a positive PCR result, relative to a negative PCR result, was 88.2 (95% CI 20.6-378). The likelihood ratio for a possible diagnosis of viral infection of the CNS in a patient with a negative PCR result, relative to a positive PCR result, was 0.10 (0.03-0.39). A patient with a positive PCR result was 88 times as likely to have a definite diagnosis of viral infection of the CNS as a patient with a negative PCR result. A negative PCR result can be used with moderate confidence to rule out a diagnosis of viral infection of the CNS. We believe that PCR will become the first-line diagnostic test for viral meningitis and encephalitis.

Detection of cerebrospinal fluid leakage by isoelectric focusing on polyacrylamide gels with silver staining using the PhastSystem.

Cerebrospinal fluid (CSF) leakage, which sometimes occurs after skull trauma, is a life-threatening condition. A prompt start with antibiotics and/or prompt surgical treatment of fistulas is essential to avoid severe complications. This requires a fast and reliable method for detecting CSF leakage. This paper describes a fast (< 2 h) method based on the identification of the tau protein (beta 2-transferrin) band(s). Tau protein is a brain-specific variant of transferrin that is characteristic of CSF. The method includes iso-electric focusing (IEF) on pre-cast polyacrylamide gels and silver staining using the PhastSystem, an automated instrument for electrophoresis and staining. In the present study, this technology was applied on 200 consecutive CSF samples, 32 of which were from healthy volunteers. Tau protein was detected in all CSF samples but 5 (2.5%), all of which were from patients with blood-brain barrier (BBB) damage. In these cases, the tau protein band was indistinct when direct silver staining was used. Therefore, immunofixation with an antitransferrin antibody was performed, and after that the tau protein band was easy to detect. The specificity of the method was high, since no brain-specific tau protein band was detected in serum, tears, saliva, or nasal secretion. As IEF of CSF using the PhastSystem is increasingly used as the routine method for detection of oligoclonal bands of IgG in neurological disorders, it could readily be used in the clinical (neuro) chemical laboratory also for the less frequent cases of suspected CSF leakage.

Terminal Deoxynucleotidyl Transferase (TdT)-Positive Cells in Cerebrospinal Fluid and Development of Overt CNS Leukemia: A 5-Year Follow-up Study in 113 Children With a TdT-Positive Leukemia or Non-Hodgkin’s Lymphoma

AbstractWe investigated whether an indirect nuclear terminal deoxynucleotidyl transferase (TdT) immunofluorescence (IF) assay on single cells present in the cerebrospinal fluid (CSF) is more effective than conventional cytomorphology for early detection or exclusion of (minimal) meningeal leukemic infiltration in patients with a TdT+ malignancy. During a 5-year follow-up study, 1,661 consecutive CSF samples from 113 children with a TdT+ acute lymphoblastic leukemia (ALL) (n = 100), a TdT+ acute nonlymphoblas- tic leukemia (ANLL) (n = 8), or a TdT+ non-Hodgkin's lymphoma (NHL) (n = 5) were analyzed. In 1,511 (91.9%) of 1,643 evaluable CSF samples, the positive and negative findings of both cytomorphology and the TdT-IF assay were concordant. In 47 (2.9%) samples from 28 patients, the cytomorphology was suspect while the TdT-IF assay was negative; follow-up as long as 58 months revealed no CNS leukemia in any patient. In 85 (5.2%) samples, cytomorphology was negative (n = 70) or suspect (n = 15) but TdT+ cells were detected. RBC contamination seriously hampered evaluation in 31 of these 85 samples. From the remaining 54 TdT+ samples from 29 patients, 40 samples preceded overt CNS leukemia in 20 patients. Two consecutive findings of TdT + cells in the CSF were always followed by overt CNS leukemia. At initial diagnosis, 11 children had TdT+ cells in their RBC-free CSF. In one of these children, morphology was suspect; a repeated lumbar puncture was positive on both assays. Thus, initial CNS leukemia was diagnosed. In the other ten children, morphology was negative. In six of them, CNS leukemia was diagnosed 2 to 20 months later. In 32 other children examined at initial diagnosis, neither TdT* cells nor blasts were observed in the CSF. In none of these patients was a CNS leukemia diagnosed after a follow-up of 2.5 to 57 months (median 24 months). In 207 control CSF samples from 58 children with TdT“ oncologic, hematologic, or infectious diseases, no TdT+ cells could be detected. The TdT-IF assay is easy to perform and is a more reliable diagnostic tool for detection of CNS leukemia at an early stage than is cytomorphology. At initial diagnosis, the finding of TdT+ cells in a RBC-free CSF sample with a negative cytomorphology is highly predictive for development of overt CNS leukemia.

Terminal deoxynucleotidyl transferase (TdT)-positive cells in cerebrospinal fluid and development of overt CNS leukemia: a 5-year follow-up study in 113 children with a TdT-positive leukemia or non- Hodgkin's lymphoma

We investigated whether an indirect nuclear terminal deoxynucleotidyl transferase (TdT) immunofluorescence (IF) assay on single cells present in the cerebrospinal fluid (CSF) is more effective than conventional cytomorphology for early detection or exclusion of (minimal) meningeal leukemic infiltration in patients with a TdT+ malignancy. During a 5- year follow-up study, 1,661 consecutive CSF samples from 113 children with a TdT+ acute lymphoblastic leukemia (ALL) (n = 100), a TdT+ acute nonlymphoblastic leukemia (ANLL) (n = 8), or a TdT+ non-Hodgkin's lymphoma (NHL) (n = 5) were analyzed. In 1,511 (91.9%) of 1,643 evaluable CSF samples, the positive and negative findings of both cytomorphology and the TdT-IF assay were concordant. In 47 (2.9%) samples from 28 patients, the cytomorphology was suspect while the TdT- IF assay was negative; follow-up as long as 58 months revealed no CNS leukemia in any patient. In 85 (5.2%) samples, cytomorphology was negative (n = 70) or suspect (n = 15) but TdT+ cells were detected. RBC contamination seriously hampered evaluation in 31 of these 85 samples. From the remaining 54 TdT+ samples from 29 patients, 40 samples preceded overt CNS leukemia in 20 patients. Two consecutive findings of TdT+ cells in the CSF were always followed by overt CNS leukemia. At initial diagnosis, 11 children had TdT+ cells in their RBC-free CSF. In one of these children, morphology was suspect; a repeated lumbar puncture was positive on both assays. Thus, initial CNS leukemia was diagnosed. In the other ten children, morphology was negative. In six of them, CNS leukemia was diagnosed 2 to 20 months later. In 32 other children examined at initial diagnosis, neither TdT+ cells nor blasts were observed in the CSF. In none of these patients was a CNS leukemia diagnosed after a follow-up of 2.5 to 57 months (median 24 months). In 207 control CSF samples from 58 children with TdT- oncologic, hematologic, or infectious diseases, no TdT+ cells could be detected. The TdT-IF assay is easy to perform and is a more reliable diagnostic tool for detection of CNS leukemia at an early stage than is cytomorphology. At initial diagnosis, the finding of Tdt+ cells in a RBC-free CSF sample with a negative cytomorphology is highly predictive for development of overt CNS leukemia.

Neuronotrophic activities in cerebrospinal fluid of head trauma patients

Neuronotrophic factors (NTFs) are agents required for neurons to survive in tissue culture. In this study, we investigated the presence of NTFs in cerebrospinal fluid (CSF) of patients with central nervous system (CNS) injury. Cerebrospinal fluid was collected from 15 patients with acute CNS lesions in whom ventricular catheters had been placed to monitor and to facilitate the control of intracranial pressure. Neuronotrophic activity within the CSF was assayed using cultures of neurons derived from fetal rat hippocampus and embryonic chick cerebral cortex. Cerebrospinal fluid from all 15 patients contained NTFs which supported the survival of rat hippocampal neurons. Survival of chick cortex neurons was supported by eight of nine CSF samples. In the 11 patients from whom consecutive CSF samples were available, NTF activity assayed in rat hippocampal cultures tended to decrease during the first several days after CNS injury. In CSF collected from three patients by lumbar puncture for diagnosis of “nontraumatic” conditions, no NTFs were detectable. NTFs supporting hippocampal neurons were also detected in extracts of blood clot obtained from normal volunteers. Neuronotrophic activity in the CSF was heat sensitive, nondialyzable, and macromolecular, suggesting its association with a protein(s). These observations suggested that (i) NTFs are detectable in human CSF after CNS injury, (ii) NTFs appear in response to the injury itself, and (iii) at least some human NTFs can support the survival in culture of nonhuman CNS neurons.