Experimental Meningitis Research Articles

Effect of Experimental Escherichia coli Meningitis on Concentrations of Excitatory and Inhibitory Amino Acids in the Rabbit Brain: In Vivo Microdialysis Study

Excessive extracellular fluid concentrations of the amino acids glutamate and aspartate play an important role in the pathogenesis of neuronal cell damage during hypoxia, hypoglycemia, and seizure. The purpose of these investigations was to test the hypothesis that bacterial meningitis causes progressive increase in excessive extracellular fluid concentrations of excitatory and inhibitory neurotransmitters. To test this hypothesis, Escherichia coli was injected intracisternally in juvenile rabbits after which neurotransmitter concentrations were measured with in vivo microdialysis. The data showed significant elevation of the excitatory amino acids aspartate and glutamate, as well as of the inhibitory neurotransmitters gamma-amino butyric acid and taurine in the excessive extracellular fluid of animals injected with E. coli compared with control animals injected with saline. However, concentrations of these excitatory and inhibitory amino acids rose late in the course of meningitis, at a time when the animals were hypotensive (mean blood pressure < or = 40 mm Hg). These data show that the major increase in excitatory neurotransmitters during experimental meningitis occurs in association with the cerebral ischemia produced by septic shock rather than being produced by the meningitis itself.

Immune-Mediated Injury in Bacterial Meningitis

Cytokines are involved in the host response to bacterial infections. In bacterial meningitis, intrathecal synthesis of TNF-alpha and IL-1 is likely to contribute to CNS injury by recruitment and activation of inflammatory cells with subsequent release of toxic factors, such as reactive oxygen intermediates and excitatory amino acids (glutamate), leading to neuronal cell death with neurologic sequelae. In rats with experimental meningitis, pretreatment with TGF-beta inhibits cerebrovascular changes and brain edema formation in the early, TNF-alpha-independent phase. Provided its local production in bacterial infection, TGF-beta may comprise a host factor interfering with immune pathologic events altering the integrity of the endothelial barrier.

EFA Commentary: A collaborative approach to epilepsy education in the school

Despite advances in antimicrobial therapy and advanced critical care neonatal bacterial meningitis has a mortality rate of over 10% and induces neurological sequelae in 20–50% of cases. Escherichia coli K1 (E. coli K1) is the most common gram-negative organism causing neonatal meningitis and is the second most common cause behind group B streptococcus. We previously reported that an E. coli K1 experimental meningitis infection in neonatal rats resulted in habituation and aversive memory impairment and a significant increase in cytokine levels in adulthood. In this present study, we investigated the oxidative stress profile including malondialdehyde (MDA) levels, carbonyl protein formation, myeloperoxidase activity (MPO) activity, superoxide dismutase (SOD) activity and catalase (CAT) activity 6, 12, 24, 48, 72 and 96 h after E. coli K1 experimental meningitis infection. In addition, sulfhydryl groups, nitrite and nitrate levels and activity of the mitochondrial respiratory chain enzymes were also measured in the frontal cortex and hippocampus of neonatal rats. The results from this study demonstrated a significant increase in MDA, protein carbonyls and MPO activity and a simultaneous decrease in SOD activity in the hippocampus of the neonatal meningitis survivors but the same was not observed in frontal cortex. In addition, we also observed a significant increase in complex IV activity in the hippocampus and frontal cortex of meningitis survivor rats. Thus, the results from this study reaffirmed the possible role of oxidative stress, nitric oxide and its related compounds in the complex pathophysiology of E. coli K1-induced bacterial meningitis.

Elevated body temperature restricts growth of Haemophilus influenzae type b during experimental meningitis.

Elevation of the environmental temperature appeared to counteract the temperature-depressing effects of urethane anesthetic and allowed rabbits intracisternally infected with Haemophilus influenzae type b to mimic the development of a fever following infection. Elevated core body temperature (greater than 39 degrees C) was associated with an inhibition of the growth of H. influenzae in cerebrospinal fluid (CSF) during the first 12 h postinfection, whereas bacterial growth was essentially unrestricted in rabbits with reduced (approximately 37 degrees C) body temperature. Bacterial densities 24 h postinfection were different, hyperthermic animals having log 6.0 +/- 0.4 CFU/ml of CSF and hypothermic rabbits having log 8.2 +/- 0.8 CFU/ml of CSF (P less than 0.05, Wilcoxon rank sum test). However, the growth of this bacterium in vitro, in either pooled rabbit CSF or brain heart infusion broth, was not inhibited at 39 degrees C. These results suggest that elevated body temperature associated with the development of fever during meningitis may be associated with restriction of the growth of H. influenzae in vivo but that this effect is apparently not due to an innate inability of the bacterium to grow at elevated temperatures.

Transforming growth factor beta 2 inhibits cerebrovascular changes and brain edema formation in the tumor necrosis factor alpha-independent early phase of experimental pneumococcal meningitis.

Macrophages and granulocytes seem to play a key role in the pathogenesis of bacterial meningitis. Transforming growth factor beta (TGF-beta) leads to macrophage deactivation, as well as to inhibition of cytokine production and of endothelial granulocyte adhesion. We have investigated the influence of TGF-beta on regional cerebral blood flow (rCBF), intracranial pressure (ICP), and brain edema formation during the early phase of experimental meningitis. Rats which were inoculated intracisternally with live pneumococci or with pneumococcal cell wall hydrolyzed by the M1 muramidase (PCW-M) developed an increase of rCBF and ICP within 4 h postintracisternal challenge. A single intraperitoneal injection of TGF-beta 2 but not of TGF-beta 2 vehicle-control prevented the changes of rCBF. Furthermore, TGF-beta 2 significantly reduced the increase of ICP in rats inoculated with PCW-M. Likewise, the elevation of brain water content after intracisternal injection of pneumococci or PCW-M was blocked by pretreatment of rats with TGF-beta 2. TGF-beta 1 exhibited similar inhibitory effects in PCW-M-injected rats. The beneficial effects of TGF-beta 2 on the initial phase after pneumococcal inoculation seem to be tumor necrosis factor alpha- (TNF-alpha) independent since (a) intracisternal or intraperitoneal injection of neutralizing anti-TNF-alpha antibodies did not significantly influence rCBF, ICP, and brain water content in PCW-M-induced meningitis; and (b) TNF-alpha was only occasionally detected at low levels in cerebrospinal fluid at 4 h after PCW-M application.

Pathophysiology of blood-brain barrier alterations during experimental Haemophilus influenzae meningitis.

These data demonstrate that haemocin production and type b encapsulation, although closely associated, are encoded by independent genetic elements. Studies with the infant rat model of infection suggest that haemocin may play a role in the pathogenesis of invasive H. influenzae type b infection. The availability of the cloned hmc locus and mutants deficient in the production of haemocin will allow further analysis of the genetic basis of haemocin production.

Effect of hydration status on cerebral blood flow and cerebrospinal fluid lactic acidosis in rabbits with experimental meningitis.

The effects of hydration status on cerebral blood flow (CBF) and development of cerebrospinal fluid (CSF) lactic acidosis were evaluated in rabbits with experimental pneumococcal meningitis. As loss of cerebrovascular autoregulation has been previously demonstrated in this model, we reasoned that compromise of intravascular volume might severely affect cerebral perfusion. Furthermore, as acute exacerbation of the inflammatory response in the subarachnoid space has been observed after antibiotic therapy, animals were studied not only while meningitis evolved, but also 4-6 h after treatment with antibiotics to determine whether there would also be an effect on CBF. To produce different levels of hydration, animals were given either 50 ml/kg per 24 h of normal saline ("low fluid") or 150 ml/kg 24 h ("high fluid"). After 16 h of infection, rabbits that were given the lower fluid regimen had lower mean arterial blood pressure (MABP), lower CBF, and higher CSF lactate compared with animals that received the higher fluid regimen. In the first 4-6 h after antibiotic administration, low fluid rabbits had a significant decrease in MABP and CBF compared with, and a significantly greater increase in CSF lactate concentration than, high fluid rabbits. This study suggests that intravascular volume status may be a critical variable in determining CBF and therefore the degree of cerebral ischemia in meningitis.

Concentrations of interleukin-1 beta, interleukin-6, interleukin-8 and TNF-alpha in cerebrospinal fluid from children with septic or aseptic meningitis.

Cytokines at an inflammatory site may be a better indicator of the clinical severity of an infectious disease than the serum levels of the cytokines. Concentrations of interleukin-1 beta (IL-1 beta) in paired samples of cerebrospinal fluid (CSF) from 10 rabbits with experimental bacterial meningitis caused by H. influenzae type b, were measured, and compared to the concentrations of four cytokines; IL-1 beta, interleukin-6 (IL-6), interleukin-8 (IL-8) and tumor necrosis factor-alpha (TNF-alpha) in CSF samples from 45 children with or without meningitis. The IL-1 beta concentrations in the CSF from rabbits with experimental meningitis were significantly higher than the concentrations in control animals without meningitis (p < 0.001). The mean CSF concentrations of IL-8 from meningitic children were significantly higher than in the control group without meningitis (p < 0.005). TNF-alpha was only detected in septic meningitis. Assays of IL-6, however, were not significantly different in the septic meningitis group, the aseptic meningitis group and the non-meningitis group. These data indicate a possible role of IL-1 beta, IL-8 and TNF-alpha as mediators in the meningeal inflammatory process in patients with meningitis and TNF-alpha, in particular, may play a role in the pathogenesis of septic meningitis.

Transferability of meropenem to cerebrospinal fluid in rabbits with meningitis caused by Staphylococcus aureus

The transferability of meropenem (MEPM) to cerebrospinal fluid (CSF) was studied employing rabbits with experimental meningitis caused by Staphylococcus aureus. The mean serum concentration was 93.1 +/- 13.5 micrograms/ml at 15 minutes after intravenous administration of MEPM at a dose level of 100 mg/kg. The mean concentration in CSF was maximum at 15 minutes after administration at 4.42 +/- 2.24 micrograms/ml. Pharmacokinetic parameters calculated from these values were as follows: Cmax (CSF/serum) 4.75%, AUC (CSF/serum) 10.4% between 15 and 60 minutes, 13.9% between 15 and 120 minutes and 15.7% between 15 and 180 minutes, T 1/2 for MEPM in CSF: 50.9 minutes, T 1/2 (CSF/serum): 2.19. In comparison to those of imipenem which were obtained in the same way, the transferability of MEPM was similar and in consideration of the antimicrobial potency against the main pathogens of meningitis, it appears worth-while of running clinical trials for this drug.

Animal models as predictors of outcome of therapy with broad spectrum cephalosporins.

Broad spectrum cephalosporins have been studied extensively in animal models of experimental infections. There is generally good correlation between the results of therapy of experimental infections and clinical trials in humans. However, the results of animal model studies are better predictors of the failure than of the success of a chemotherapeutic regimen. Cefotaxime and the new 'fourth' generation agent, cefpirome, were comparable in the treatment of experimental meningitis caused by Streptococcus pneumoniae. Cefpirome was the most effective cephalosporin as therapy for methicillin-susceptible Staphylococcus aureus experimental endocarditis. The most effective broad spectrum cephalosporins for the treatment of Gram-negative experimental pneumonia were cefpirome, cefotaxime and cefodizime. Cefpirome was equivalent to ceftazidime or cefazolin as treatment for Pseudomonas aeruginosa or methicillin-susceptible S. aureus experimental osteomyelitis. Because of its potent activity in vitro and in animal models of experimental infections caused by methicillin-susceptible S. aureus and Gram-negative bacilli, cefpirome may offer a therapeutic advantage over currently available broad spectrum cephalosporins.

Adjunctive therapy of experimental meningitis: agents other than steroids.

Adjunctive therapy of experimental meningitis: agents other than steroids.

Enhanced attenuation of meningeal inflammation and brain edema by concomitant administration of anti-CD18 monoclonal antibodies and dexamethasone in experimental Haemophilus meningitis.

Antiinflammatory therapy has been shown to reduce the adverse pathophysiological consequences that occur in bacterial meningitis and to improve outcome from disease. In the present study, modulation of two principal steps of the meningeal inflammatory cascade was accomplished by concomitant administration of dexamethasone to diminish overproduction of cytokines in response to a bacterial stimulus and of a monoclonal antibody directed against adhesion-promoting receptors on leukocytes to inhibit recruitment of white blood cells into the subarachnoid space. Dexamethasone and antibody therapy produced a marked attenuation of all indices of meningeal inflammation and reduction of brain water accumulation after H. influenzae-induced meningitis in rabbits compared with results of each agent given alone and of untreated animals. In addition, the enhanced host's meningeal inflammatory reaction that follows antibiotic-induced bacterial lysis was profoundly ameliorated when dual therapy was administered without affecting clearance rates of bacteria from cerebrospinal fluid and vascular compartments. The combination of both therapeutic approaches may offer a promising mode of treatment to improve further the outcome from bacterial meningitis.

The site of the lesion causing hearing loss in bacterial meningitis: a study of experimental streptococcal meningitis in guinea‐pigs

The pathological basis of hearing loss in bacterial meningitis was investigated using an animal model of Streptococcus suis meningitis. Forty guinea-pigs were infected after their hearing had been assessed by brain stem auditory evoked potentials. In 17 animals, it was possible to repeat the procedure at the onset of meningitis; this included one animal with subclinical disease. Fifteen animals showed evidence of hearing loss, which on subsequent histological examination was found to be associated invariably with suppurative labyrinthitis. The remaining two animals without hearing loss had normal cochleas. It is suggested that cochlear sepsis rather than eighth cranial nerve involvement by meningeal sepsis is primarily responsible for hearing loss in bacterial meningitis, and that bacteria enter the cochlea via the cochlear aqueduct and not the internal auditory canal. The tissue within the lumen of the cochlear aqueduct may act as a barrier against invasion by micro-organisms, and haemolytic streptococci could cause lysis of this barrier by the exotoxins they produce.

Use of ampicillin-sulbactam for treatment of experimental meningitis caused by a beta-lactamase-producing strain of Escherichia coli K-1

We evaluated the pharmacokinetics and therapeutic efficacy of ampicillin combined with sulbactam in a rabbit model of meningitis due to a beta-lactamase-producing strain of Escherichia coli K-1. Ceftriaxone was used as a comparison drug. The MIC and MBC were 32 and greater than 64 micrograms/ml (ampicillin), greater than 256 and greater than 256 micrograms/ml (sulbactam), 2.0 and 4.0 micrograms/ml (ampicillin-sulbactam [2:1 ratio, ampicillin concentration]) and 0.125 and 0.25 micrograms/ml (ceftriaxone). All antibiotics were given by intravenous bolus injection in a number of dosing regimens. Ampicillin and sulbactam achieved high concentrations in cerebrospinal fluid (CSF) with higher dose regimens, but only moderate bactericidal activity compared with that of ceftriaxone was obtained. CSF bacterial titers were reduced by 0.6 +/- 0.3 log10 CFU/ml/h with the highest ampicillin-sulbactam dose used (500 and 500 mg/kg of body weight, two doses). This was similar to the bactericidal activity achieved by low-dose ceftriaxone (10 mg/kg), while a higher ceftriaxone dose (100 mg/kg) produced a significant increase in bactericidal activity (1.1 +/- 0.4 log10 CFU/ml/h). It appears that ampicillin-sulbactam, despite favorable CSF pharmacokinetics in animals with meningitis, may be of limited value in the treatment of difficult-to-treat beta-lactamase-producing bacteria, against which the combination shows only moderate in vitro activity.

Ultrastructural localization of albumin transport across the cerebral microvasculature during experimental meningitis in the rat.

Injury to the blood brain barrier (BBB) is a fundamental sequela of bacterial meningitis, yet the precise mechanism facilitating exudation of albumin across the endothelium of the cerebral microvasculature remains conjectural. After intracisternal inoculation of Escherichia coli (0111:B4) lipopolysaccharide in rats to elicit a reversible meningitis and BBB injury, we utilized in situ tracer perfusion and immunolabeling procedures to identify by transmission electron microscopy the precise topography and microvascular exit pathway(s) of bovine serum albumin (BSA). Results revealed that during meningitis there was: (a) an inducible increase in immunodetectable monomeric BSA binding to the luminal membrane of all microvascular segments in the pia-arachnoid and superficial brain cortex; (b) similar uptake of both colloidal Au-BSA (as well as monomeric BSA) by plasmalemmal vesicles but no detectable transcytosis to the abluminal side; and (c) predominant exit of both perfused Au-BSA and immunodetectable monomeric BSA through open intercellular junctions of venules in the pia-arachnoid. This was corroborated in separate experiments documenting focal pial venular leaks of in situ perfused 0.01% colloidal carbon black during experimental meningitis. These results provide precise localization of BBB injury in meningitis to meningeal venules, confirm a paracellular exit pathway of albumin via open intercellular junctions, and suggest an injury mechanism amenable to specific therapeutic intervention.

Differences of Pathophysiology in Experimental Meningitis Caused by Three Strains of Streptococcus Pneumoniae

Differences in cytochemical and pathophysiologic abnormalities in experimental meningitis caused by pneumococcal strains A, B, and C were determined. Strain C produced the most severe abnormalities of cerebrospinal fluid (CSF) concentrations of lactate (P less than .01), protein (P less than .02), and glucose (P less than .01), CSF white blood cell count (P less than .04), cerebral blood flow (P less than .02), and clinical signs (P less than .05). Brain edema occurred only with strains A anc C, with no association with disease severity; intracranial hypertension was also independent of disease severity. Strain B, not C, achieved the highest bacterial titers in the CSF (P less than .005). The widely different abilities of strains of Streptococcus pneumoniae to induce intracranial abnormalities suggest that virulence determinants affect not only evasion of defense during colonization and invasion, as shown in other models, but also determine the course of disease once infection has been established. Differences of cell-wall metabolism among pneumococcal strains may play a role in this latter phase of the development of meningitis.

Effect of Indomethacin on the Pathophysiology of Experimental Meningitis in Rabbits

The effects of indomethacin on central nervous system abnormalities in rabbits with experimental pneumococcal meningitis were studied. As expected, prostaglandin E2 levels in cerebrospinal fluid were significantly lower in the indomethacin-treated group, indicating that the drug effectively reduced prostaglandin synthesis. Brain edema was markedly attenuated in the indomethacin-treated group; however, cerebrospinal fluid white blood cell counts, lactate and protein concentrations, and intracisternal pressure were not significantly different between groups. It seems that indomethacin, while effective in reducing brain edema, does not significantly affect other important pathophysiologic alterations in experimental pneumococcal meningitis.

Extremely High Incidence of Antibiotic Resistance in Clinical Isolates of Streptococcus pneumoniae in Hungary

An epidemiologic survey of antibiotic resistance among pneumococcal isolates collected during 1988 and 1989 in Hungary indicated that as many as 58% of all isolates and 70% of isolates from children were resistant to penicillin. These figures surpass even the highest values reported thus far for Spain and South Africa for the same period. Almost or more than 70% of the penicillin-resistant isolates were also resistant to tetracycline, erythromycin, and cotrimoxazole and approximately 30% to chloramphenicol. Intravenous administration of ampicillin (30 mg/kg) did not interfere with the growth in the cerebrospinal fluid of three resistant strains introduced into the rabbit model of experimental meningitis. No resistant strain showed beta-lactamase activity. A representative highly resistant strain contained altered penicillin-binding proteins (low penicillin affinities and abnormal molecular sizes) and was also resistant to the lytic and killing effects of penicillin.

Pathogenesis and Pathophysiology of Meningitis

Advances in the understanding of the pathogenesis and pathophysiology of meningitis have occurred primarily through the use of experimental animal models. These models have proven to be particularly valuable in experimental bacterial meningitis, focusing on the bacterial virulence factors responsible for the initiation of infections, CNS invasion, and induction of SAS inflammation. Recent studies have examined the formation of host inflammatory cytokines in response to these virulence factors. These cytokines may be responsible for many of the pathophysiologic consequences of bacterial meningitis (eg. increased BBB permeability, cerebral edema, and increased intracranial pressure). Meningitis due to C. neoformans occurs most commonly in patients with defects in cell-mediated immunity (eg, AIDS), and the depletion of T helper cells in AIDS patients may allow unrestricted cryptococcal growth. Viral meningitis is an illness of low prevalence when compared with the overall occurrence of viral infections at other sites. CNS infection usually occurs by means of traversal across barriers that normally exclude viral invasion of the CNS, primarily through hematogenous dissemination from initial sites of infection. These advances in the pathogenesis and pathophysiology of bacterial, fungal, and viral meningitis may lead to the development of innovative treatment strategies for these disorders.

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.