Cerebrospinal Fluid and Plasma Lipopolysaccharide Levels in Human Immunodeficiency Virus Type 1 Infection and Associations With Inflammation, Blood-Brain Barrier Permeability, and Neuronal Injury.

Human immunodeficiency virus (HIV) infection is associated with increased systemic microbial translocation, neuroinflammation, and occasionally, neuronal injury. Whether systemic lipopolysaccharide (LPS) penetrates into the brain and contributes to neuroinflammation remain unknown in HIV. Here, we measured plasma and cerebrospinal fluid (CSF) LPS levels along with biomarkers of neuroinflammation (white blood cell counts and 40 soluble markers) and neurofilament light chain (NfL). Notably, CSF LPS was undetectable in all samples, including 3 HIV-infected individuals with dementia. Increased plasma LPS, neuroinflammation, and blood-brain barrier (BBB) dysfunction were found in untreated HIV-infected individuals, but not in healthy or treated HIV-infected individuals. Plasma LPS levels were directly correlated with various markers of inflammation in both plasma and CSF, as well as with degree of BBB permeability but not with CSF NfL in HIV-infected subjects. These results suggest that the magnitude of microbial translocation associates with neuroinflammation and BBB permeability in HIV without direct penetration into the central nervous system.

Prospective Cohort Study on Performance of Cerebrospinal Fluid (CSF) Xpert MTB/RIF, CSF Lipoarabinomannan (LAM) Lateral Flow Assay (LFA), and Urine LAM LFA for Diagnosis of Tuberculous Meningitis in Zambia.

Tuberculous meningitis (TBM) is a devastating infection of the central nervous system lacking an adequate point-of-care diagnostic test. We conducted a prospective cohort study of 550 Zambian adults with suspected TBM to determine the diagnostic accuracy of cerebrospinal fluid (CSF) Xpert MTB/RIF, CSF lipoarabinomannan (LAM), urine LAM, CSF total protein, and CSF glucose compared with the gold standard of CSF culture. We categorized patients with a positive CSF tuberculosis (TB) culture as definite TBM. We also assessed inpatient and 1-year mortality on definite TBM patients when CSF Xpert MTB/RIF results were available in real time to treating physicians relative to a historical comparison cohort in whom Xpert results were not available in real time. Of the 550 patients, 474 (86.2%) were HIV-infected and 105/550 (19.1%) had definite TBM based on a positive CSF culture. The sensitivity/specificity of the diagnostic tests were CSF Xpert MTB/RIF, 52.9%/94.2%; CSF LAM, 21.9%/94.2%; urine LAM, 24.1%/76.1%; and CSF glucose <40 mg/dl, and total protein, >100 mg/dl, 66.3%/90%. A model including CSF Xpert MTB/RIF, CSF LAM, CSF glucose, and CSF total protein demonstrated an area under the receiver operating curve of 0.90. The inpatient and 1-year mortality for definite TBM was 43% and 57%, respectively. There was low sensitivity for the diagnosis of TBM across all diagnostics tests. CSF Xpert MTB/RIF and CSF LAM are highly specific for the diagnosis of TBM. Despite the use of Xpert MTB/RIF for diagnostic purpose in real time, TBM was still associated with a high mortality in Zambian patients.

Diagnostic value of the cerebrospinal fluid lipoarabinomannan assay for tuberculous meningitis: a systematic review and meta-analysis.

Objective: This systematic review aims to evaluate the diagnostic accuracy of cerebrospinal fluid (CSF) lipoarabinomannan (LAM) assays in detecting tuberculous meningitis (TBM). Methods: A systematic review search was conducted in PubMed and five other databases up to April 2023. Studies that evaluated the diagnostic accuracy of CSF LAM assays were included with either definitive or composite reference standard used as the preferred reference standard. The quality of the included studies was assessed using the QUADAS-2 tool. We performed a bivariate random-effects meta-analysis and calculated the summary diagnostic statistics. Results: A total of six studies, including a sample size of 999, were included in the final analysis. The pooled sensitivity, specificity, and area under the receiver operating characteristic curve (AUC) of CSF LAM for diagnosing TBM were determined to be 0.44 (95% CI: 0.31-0.58), 0.89 (95% CI: 0.81-0.93), and 0.76 (95% CI: 0.73-0.80), respectively. Significant heterogeneity was observed in both sensitivity (Q = 73.82, p < 0.01; I2 = 86.45, 95%CI: 79.64-93.27) and specificity (Q = 95.34, p < 0.01; I2 = 89.51, 95% CI: 84.61-94.42). Regression analysis indicated that the study design (retrospective vs. prospective) was associated with the heterogeneity of pooled sensitivity and specificity (all p < 0.05). Conclusion: Although more prospective studies are required to validate the role of the CSF LAM assay, current evidence supports that the performance of the CSF LAM assay is unsatisfactory for the TBM diagnosis. Additionally, the optimization of the CSF LAM assay (e.g., improvements in CSF collection and preparation methods) should be considered to improve its performance.

Contribution of capsular and clonal types and beta-lactam resistance to the severity of experimental pneumococcal meningitis.

We used a rabbit model to assess the effects of capsular serotype, genetic background and beta-lactam resistance on the course and severity of experimental meningitis. Meningitis was induced by five pneumococcal strains belonging to five different clones with known invasive potential: two serotype 3 strains (ST260(3) and Netherlands(3)-31 clones) and three serotype 23F strains with different beta-lactam susceptibility patterns (Spain(23F)-1 clone, Tennessee(23F)-4 clone and a double locus variant of the Tennessee(23F)-4 clone). Major differences in secondary bacteremia and mortality rates were observed between serotypes 3 and 23F, as were divergences in the CSF lactate, protein and lipoteichoic-teichoic acid concentrations. Minor differences in the CSF-induced inflammatory response were found among strains belonging to the same serotype. Our results suggest that capsular serotype might be the main factor determining the course and severity of pneumococcal meningitis and genetic background contributes to a lesser extent. The acquisition of beta-lactam resistance does not reduce the virulence of the invasive clones. Since five strains belonging to two serotypes were studied, our findings have to be confirmed with other pneumococcal serotypes.

Clinical outcome in pneumococcal meningitis correlates with CSF lipoteichoic acid concentrations.

Lipoteichoic and teichoic acids are components of the cell wall of Streptococcus pneumoniae. A recently developed enzyme immunoassay was used in patients with pneumococcal meningitis to investigate lipoteichoic and teichoic acid concentrations in CSF at the first lumbar puncture in relation to the clinical outcome determined by the Glasgow Outcome Scale. Lipoteichoic and teichoic acid concentrations in CSF were significantly associated with neurologic sequelae and mortality in S. pneumoniae meningitis.

Specific detection of Haemophilus influenzae type b lipooligosaccharide by a polymyxin B monoclonal antibody assay.

A monoclonal antibody (MAb)-based enzyme immunoassay was developed for detection of Haemophilus influenzae type b (Hib) lipooligosaccharides (LOS). The high affinity of polymyxin B for lipid A was used to bind the Hib LOS to microtiter wells. The immobilized LOS was detected with MAbs directed against the oligosaccharide component of Hib endotoxin. Hib LOS concentrations were measured in in vitro samples and in cerebrospinal fluid (CSF) sample obtained from rabbits with experimental Hib meningitis. The sensitivity of the assay was 1 ng LOS/ml sample and the results obtained with this assay correlated significantly with those obtained with the standard Limulus amebocyte lysate assay. This new assay provides a method for specific detection of Hib LOS in CSF samples and in aqueous laboratory fluids. This general methodology should also be useful for experimental research involving specific LPS/LOS molecules.

Inducible nitric oxide synthase expression elicited in the mouse brain by inflammatory mediators circulating in the cerebrospinal fluid.

Expression of inducible nitric oxide synthase (iNOS) protein was studied in the brain after intracerebroventricular injections of interferon (IFN)-gamma, and IFN-gamma combined with lipopolysaccharide (LPS) or tumor necrosis factor (TNF)-alpha, compared to ovalbumin as control. Wild-type mice and mice with targeted deletion of the IFN-gamma receptor gene were used. Findings based on iNOS immunoreactivity were evaluated at 1, 2, 4 and 7 days post-injection, using also quantitative image analysis and double labeling with glial cell markers. IFN-gamma administration induced iNOS immmunostaining in activated microglia and macrophages in the parenchyma surrounding the ventricular system, several cortical fields and fiber tracts. IFN-gamma-elicited iNOS immunoreactivity was down-regulated after 1 day. The number of iNOS-immunopositive cells was significantly enhanced by co-administration of LPS or TNF-alpha; IFN-gamma+TNF-alpha injections also resulted in longer persistence of iNOS immunoreactivity. No immunopositive cells were seen in the brain of IFN-gamma receptor knockout mice after IFN-gamma administration; very few immunostained macrophages were detected in these cases, mostly around the injection needle track, after co-administration of LPS or TNF-alpha. Western blot analysis confirmed a marked iNOS induction in the brain of wild-type mice 24 h after IFN-gamma+LPS injections. The findings show that inflammatory mediators circulating in the cerebrospinal fluid induce in vivo iNOS in the brain with topographical selectivity and temporal regulation. The data also demonstrate that the signaling cascade activated by IFN-gamma binding to its receptor is critical for iNOS induction, and the synergistic action of LPS and TNF-alpha as iNOS inducers in brain cells is largely mediated by the receptor-regulated action of IFN-gamma.

The lipooligosaccharide (LOS) of Neisseria meningitidis serogroup B strain NMB contains L2, L3, and novel oligosaccharides, and lacks the lipid-A 4'-phosphate substituent.

The complete structure of the lipooligosaccharide (LOS) from Neisseria meningitidis strain NMB (serotype 2b:P1.2,5), a serogroup B cerebrospinal fluid isolate, was determined. Two oligosaccharide (OS) fractions and lipid-A were obtained following mild acid hydrolysis of the LOS. The structures in these fractions were determined using glycosyl composition and linkage analyses, N spectroscopy and mass spectrometry. One oligosaccharide fraction (OS1) consists of a molecule having a glycosyl sequence identical to that previously reported for the LOS from immunotype L2 N. meningitidis [A. Gamain, M. Beurret, F. Michon, J.-R. Brisson, and H.J. Jennings, J. Biol. Chem.,267,(112) 922-925] i.e., a lacto-N-neotetraose is attached to heptose I (Hep I), with terminally linked N-acetylglucosaminosyl and glucosyl residues attached to Hep II of the inner core. Approximately 70% of this structure is acetylated at O-6 of the terminally linked alpha-N-acetyl-glucosaminosyl residue. As with the L2 structure, the NMB LOS contained phosphoethanolamine (PEA) at O-6 or O-7 of the Hep II residue. The second oligosaccharide fraction (OS2) contains a a mixture of three different molecules, all of which vary from one another in their glycosyl substitution patterns of the Hep II residue. The most abundant molecule in OS2 has a structure identical to that of OSI, i.e., it has the L2 glycosyl sequence. A second molecule (OS2a) lacks the terminal glucosyl residue at O-3 of Hep II; i.e., it has a glycosyl sequence identical to that of the mild acid released oligosaccharide of N. meningitidis immunotype L3, L4, or L7 LOSs. The third molecule (OS2b) is a novel structure that lacks the terminal N-acetylglucosaminosyl residue linked to O-2 of Hep II. Overall, 76% of OS released from NMB LOS has the L2 structure, 15% is OS2a (L3), and 9% is OS2b. A portion (20%) of the molecules in the NMB LOS preparation also contained terminally linked sialic acid attached to O-3 of the lacto-N-neotetraose galactosyl residue, which is also consistent with the L3, or L4 LOS structures. In contrast to the previously reported structure of N. meningitidis lipid-A [V. A. Kulshin, U. Zähringer, B. Linder, C.E. Frasch, C-M. Tsai, B.A. Dmitriev, and E.T Rietschel, J. Bacteriol., 174, (1992)1793-1800], only 30% of the lipid-A from NMB LOS possesses 4'-phosphate. Comparison with the lipid-A of LOS purified from an isogenic acapsulate mutant, M7, revealed that the 4'-position was almost completely occupied with phosphate. These data emphasize the structural heterogeneity of the OS and phosphate substituents of Hep II, and 4'-phosphorylation of lipid-A of meningococcal LOS.

Comparison of a clinical prediction rule and a LAM antigen-detection assay for the rapid diagnosis of TBM in a high HIV prevalence setting.

BACKGROUND/OBJECTIVE: The diagnosis of tuberculous meningitis (TBM) in resource poor TB endemic environments is challenging. The accuracy of current tools for the rapid diagnosis of TBM is suboptimal. We sought to develop a clinical-prediction rule for the diagnosis of TBM in a high HIV prevalence setting, and to compare performance outcomes to conventional diagnostic modalities and a novel lipoarabinomannan (LAM) antigen detection test (Clearview-TB®) using cerebrospinal fluid (CSF). METHODS: Patients with suspected TBM were classified as definite-TBM (CSF culture or PCR positive), probable-TBM and non-TBM. RESULTS: Of the 150 patients, 84% were HIV-infected (median [IQR] CD4 count = 132 [54; 241] cells/µl). There were 39, 55 and 54 patients in the definite, probable and non-TBM groups, respectively. The LAM sensitivity and specificity (95%CI) was 31% (17;48) and 94% (85;99), respectively (cut-point ≥ 0.18). By contrast, smear-microscopy was 100% specific but detected none of the definite-TBM cases. LAM positivity was associated with HIV co-infection and low CD4 T cell count (CD4<200 vs. >200 cells/µl; p = 0.03). The sensitivity and specificity in those with a CD4<100 cells/µl was 50% (27;73) and 95% (74;99), respectively. A clinical-prediction rule ≥ 6 derived from multivariate analysis had a sensitivity and specificity (95%CI) of 47% (31;64) and 98% (90;100), respectively. When LAM was combined with the clinical-prediction-rule, the sensitivity increased significantly (p<0.001) to 63% (47;68) and specificity remained high at 93% (82;98). CONCLUSIONS: Despite its modest sensitivity the LAM ELISA is an accurate rapid rule-in test for TBM that has incremental value over smear-microscopy. The rule-in value of LAM can be further increased by combination with a clinical-prediction rule, thus enhancing the rapid diagnosis of TBM in HIV-infected persons with advanced immunosuppression.

Increased circulating interleukin-1 and interleukin-6 after intracerebroventricular injection of lipopolysaccharide.

Recently, it was demonstrated that intracerebroventricular (icv) injection of interleukin-1 (IL-1) stimulated circulating interleukin-6 (IL-6) to a greater degree than intravenous (i.v.) injection of IL-1. The goal of this study was to compare the efficacy of lipopolysaccharide (LPS), injected both icv and i.v., on circulating concentrations of IL-1 and IL-6. Both i.v. and icv injection of LPS stimulated plasma levels of IL-1 to a similar degree. However, i.v. injection of LPS was significantly more efficacious than icv injection of LPS in elevating circulating IL-6. These results demonstrate that like i.v. injection of LPS, icv injection of LPS stimulates plasma levels of IL-1 and IL-6. Increases in circulating cytokines during infectious diseases which are limited to the central nervous system may serve to activate peripheral functions of an acute phase response.

Dry up method as a revised Limulus test with a new technique for gelatin inhibitor removing.

To detect endotoxins, Limulus test, especially its tube method, is recently used most widely. But this method has shortcomings considerably, for example, the lack of the objectivity on the judgement, the necessity of long handling time and the requirement of relatively large amount of Limulus lysate. To revise these shortcomings we established a new method. In our method, sample and Limulus lysate are mixed on a silicone coated slide glass and incubated at 37 degrees C for 30 min, then heated to dry up for the judgement. In samples which contain protein, the pretreatments with (NH4)2SO4, dilution and boiling are performed to remove gelation inhibitor. It was proved that this method could be applied to such samples as physiological saline, plasma, urine, transudate, exudate, cerebrospinal fluid (CSF) and suspension of Escherichia coli (E. coli). This method has advantages in its (1) objectivity of the judgement because of the clear difference of the dry up patterns between positive reaction and negative, (2) shortness of the handling time (results can be obtained within 2 hr from sampling), (3) requirement of little amount of sample and of Limulus lysate (a fifth volume of sample and a tenth volume of lysate are needed compared with the conventional method) and (4) sensitivity (0.1 or 0.5 ng/ml of lipopolysaccharide (LPS)).

Rifampin followed by ceftriaxone for experimental meningitis decreases lipoteichoic acid concentrations in cerebrospinal fluid and reduces neuronal damage in comparison to ceftriaxone alone.

Rifampin (RIF) releases smaller quantities of lipoteichoic acids (LTAs) from Streptococcus pneumoniae than ceftriaxone (CRO). Due to the rapid development of resistance, RIF cannot be used as a single agent for therapy of bacterial meningitis. For this reason, we compared the effect of treatment with RIF followed by treatment with CRO (RIF-CRO) or the effect of treatment with clindamycin (CLI) followed by treatment with CRO (CLI-CRO) to that of CRO alone on the concentrations of LTAs and teichoic acids in vitro. The effects of RIF-CRO on LTA concentrations in cerebrospinal fluid (CSF) and on neuronal injury were investigated in a rabbit model of S. pneumoniae meningitis. In vitro, bacterial titers were effectively reduced by CRO, RIF-CRO, and CLI-CRO when each drug was used at 10 micro g/ml. The levels of release of LTAs after the initiation of therapy were lower in RIF-CRO- and CLI-CRO-treated cultures than in cultures treated with CRO alone (P < 0.05 from 3 to 12 h after initiation of treatment). Similarly, in rabbits, the increase in the amount of LTAs in CSF was lower in RIF-CRO-treated animals than in CRO-treated animals (P = 0.02). The density of dentate apoptotic granular cells was lower after RIF-CRO therapy than after CRO therapy (medians, 58.4 and 145.6/mm(2), respectively; 25th quartiles, 36.3 and 81.7/mm(2), respectively; 75th quartiles, 100.7 and 152.3/mm(2), respectively; P = 0.03). Therefore, initiation of therapy with a protein synthesis-inhibiting antibacterial and continuation of therapy with a combination that includes a beta-lactam may be a strategy to decrease neuronal injury in bacterial meningitis.

Use of robotized DNA isolation and real-time PCR to quantify and identify close correlation between levels of Neisseria meningitidis DNA and lipopolysaccharides in plasma and cerebrospinal fluid from patients with systemic meningococcal disease.

The present study, using robotized DNA isolation and quantitative PCR based on the Neisseria meningitidis-specific capsular transport A gene, demonstrates the ease, rapidity, specificity, and sensitivity of quantifying neisserial DNA in plasma (n = 65) and cerebrospinal fluid (CSF) (n = 12) from patients with systemic meningococcal disease. We found a close correlation between the levels of neisserial DNA and lipopolysaccharides in plasma (r = 0.905) and in CSF (r = 0.964). The median concentration of neisserial DNA in plasma in 23 patients with persistent shock was 2 x 10(7) copies/ml, versus <10(3) copies/ml in 42 nonshock patients. Furthermore, quantitative PCR made possible estimates of the total number of meningococci in plasma, as opposed to conventional blood cultures, suggesting about 1,000 dead meningococci for every viable bacterium. Finally, with logistic regression analyses, neisserial DNA may predict a patient's disease severity and outcome at hospital admission. The number of meningococci in plasma and CSF appears to be the main determinant of the lipopolysaccharide levels, clinical presentation, and outcome.

Haemophilus influenzae type b lipooligosaccharide induces meningeal inflammation.

We evaluated the ability of two Haemophilus influenzae type b (Hib) components, lipooligosaccharide (LOS) and capsular polysaccharide, to provoke meningeal inflammation in rabbits. Intracisternal inoculation of 2 fg-200 ng of LOS produced a dose-dependent increase in concentrations of white blood cells and protein in cerebrospinal fluid, whereas 4 micrograms of Hib capsular polysaccharide did not provoke meningeal inflammation. Preincubation of LOS with a murine monoclonal antibody to Hib LOS did not reduce the potency of the LOS. Incubation of LOS with polymyxin B (which neutralizes LOS by binding to its lipid A region) and deacylation of the LOS with acyloxyacyl hydrolase (a neutrophil enzyme that removes nonhydroxylated fatty acyl chains from lipid A) reduced meningeal inflammation. We demonstrated that purified Hib LOS induced meningeal inflammation in this model and suggest that the lipid A moiety of Hib LOS is principally responsible for this host response.

Cerebrospinal fluid endotoxin levels in children with H. influenzae meningitis before and after administration of intravenous ceftriaxone.

Total, cell-free, and cell-bound endotoxin and bacterial density were measured in cerebrospinal fluid (CSF) of 22 children with Hemophilus influenzae meningitis. Also the effect of ceftriaxone on CSF endotoxin levels was investigated in eight patients by reexamining their CSF 2-6 h after the initial dose. Initial CSF bacterial density correlated with initial CSF endotoxin levels (P less than .001). Ceftriaxone induced a marked increase of free endotoxin in CSF, from an initial (mean +/- SE) 0.75 +/- 0.21 to 1.29 +/- 0.23 log10 ng/ml (P less than .01). This increase correlated positively with the number of bacteria killed in the CSF (P less than .01). The increase in free endotoxin was associated with an increase in mean CSF lactate levels from 8.5 to 9.7 units/l (P less than .05) and mean lactate dehydrogenase levels from 102 to 180 mmol/l (P less than .02) and a decrease in mean CSF glucose from 1.17 to 0.46 mmol/l (P less than .05). Initial CSF total endotoxin concentrations correlated both with the Herson-Todd clinical severity score (P less than .001) and with the number of febrile hospital days (P less than .001). These findings suggest that highly bactericidal agents initially lead to release of free endotoxin from gram-negative organisms into CSF, with associated enhanced inflammatory response by the host.

Specific detection of Haemophilus influenzae type b lipooligosaccharide by immunoassay.

Three monoclonal antibody (MAb)-based immunoassays were developed for specific detection of Haemophilus influenzae type b (Hib) lipooligosaccharide (LOS). (i) Hib LOS was captured onto microtiter plates by polyclonal Hib-directed antibodies and detected with MAbs to the oligosaccharide component of Hib LOS in an enzyme-linked immunosorbent assay, (ii) The high affinity of polymyxin B for lipid A was used to bind Hib LOS to microtiter wells, and the oligosaccharide-specific MAbs were used as the detection system in the polymyxin B-MAb assay. (iii) Hib LOS solubilized in detergent was captured by MAbs, and the immobilized LOS was detected with a chromogenic Limulus amebocyte lysate method in the immunolimulus assay. Endotoxin concentrations were measured in in vitro samples and cerebrospinal fluid samples from rabbits with experimental Hib meningitis. The results were compared with those obtained with the standard chromogenic Limulus amebocyte lysate assay. There were significant correlations between the results of all four assays. These new immunoassays provide methods for specific detection of Hib LOS in laboratory fluids and in research involving quantification of Hib endotoxin in experimental animal models.

Bacterial components and the pathophysiology of injury to the blood-brain barrier: does cell wall add to the effects of endotoxin in gram-negative meningitis?

In animal models, the lipopolysaccharide (LPS) from Haemophilus influenzae contributes to all the signs of meningitis associated with living bacteria. However, when tested in vitro, the amount of LPS in cerebrospinal fluid (CSF) in natural disease shows much greater effects on leukocytes than on endothelial permeability. To investigate whether other bacterial components act with LPS to incite meningeal inflammation, animals were challenged intracisternally with H. influenzae lysates. Upon neutralization of endotoxin, leukocytosis was greatly attenuated, but protein accumulation in CSF persisted. Cell wall from H. influenzae induced meningeal inflammation in a pattern opposite to that of LPS. Its ability to induce blood-brain barrier permeability greatly exceeded its ability to induce leukocytosis in vivo. Thus, cell wall, by acting on endothelia, and LPS, by inducing leukocytosis, may cooperate to induce inflammation in H. influenzae meningitis. Optimal reduction of inflammation and tissue damage in meningitis may require agents directed against cell wall as well as LPS.

Compartmentalization of lipopolysaccharide production correlates with clinical presentation in meningococcal disease.

Lipopolysaccharide (LPS, endotoxin) was quantified in plasma and cerebrospinal fluid (CSF) collected simultaneously from patients with systemic meningococcal disease. High levels (median, 3800 ng/L; range, 750-14,000) were present in plasma and low levels (median, 40 ng/L; range, less than 25-165) in CSF of patients with fulminant septicemia. Conversely, high levels (median, 2500 ng/L; range, less than 25-500,000) in CSF and low or undetectable levels (median, less than 25 ng/L; range, less than 25-210) in plasma were associated with meningitis without septic shock. Levels of LPS were significantly correlated with protein levels in CSF (r = .50, P = .01) and inversely correlated with the ratio of glucose in CSF to that in blood (r = -.62, P = .0005). LPS level in CSF greater than 800 ng/L was significantly associated with greater than or equal to 10(9) leukocytes/L, protein levels greater than 0.5 g/L, and a glucose ratio less than 0.5. Thus, quantification of LPS levels in the plasma and CSF in systemic meningococcal disease is a better predictor of pathophysiologic events than is demonstrating the presence of live bacteria as in conventional culture.

In vivo and in vitro expression of Haemophilus influenzae type b lipooligosaccharide epitopes.

An indirect immunofluorescence system involving monoclonal antibodies (MAbs) directed against surface epitopes of Haemophilus influenzae type b (Hib) lipooligosaccharide (LOS) was used to examine individual Hib cells in cerebrospinal fluid (CSF) from infants with Hib meningitis. In four of five CSF samples studied, 100% of the bacteria bound at least one LOS-directed MAb. When the bacteria from these CSF samples were grown in broth, most of these cells lost some or all of their ability to bind the MAb(s) that were bound by the same organisms present in human CSF. When in vitro-grown cells were used for intracisternal injection of rabbits, the populations of Hib cells observed in rabbit CSF after the development of meningitis generally resembled those of the respective broth-grown inocula in terms of their LOS antigenic characteristics. Hib cell populations recovered in infant rat CSF after intranasal challenge again had LOS antigenic characteristics similar to those of the in vitro-grown challenge inocula. These results indicate that a population of Hib cells growing in the infected human host may be quite different, with regard to its LOS antigenic characteristics, from the same Hib strain growing in vitro or in vivo in animal models.

Enzyme immunoassay detecting teichoic and lipoteichoic acids versus cerebrospinal fluid culture and latex agglutination for diagnosis of Streptococcus pneumoniae meningitis.

A newly developed enzyme immunoassay (EIA) was used to detect the presence of pneumococcal teichoic and lipoteichoic acids in cerebrospinal fluid (CSF) from patients with Streptococcus pneumoniae meningitis who were being treated with antibiotics. All initial CSF samples, which on culture grew S. pneumoniae, were positive in the EIA. A total of 14 subsequent culture-negative samples gave clear signals in the EIA up to day 15 after the onset of antibiotic treatment. For 11 CSF specimens, culture, microscopy, and latex agglutination were negative while the EIA detected pneumococcal antigens. The EIA did not react either with CSF of patients with meningitis caused by bacteria other than S. pneumoniae or by viral pathogens. In conclusion, this EIA can be a valuable tool for the diagnosis of S. pneumoniae meningitis from CSF samples in cases in which prior antimicrobial therapy minimizes the usefulness of culture or other antigen detection tests.