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Purpose: Streptococcus pneumoniae is an important human respiratory tract pathogen causing pneumococcal diseases in majority of children and adults. The capsule is a significant virulence factor of Pneumococci which determines the bacterial serotype and is the component used for synthesis of pneumococcal vaccines. This cross-sectional study aimed to isolate Streptococcus pneumoniae from clinical samples and determine the occurrence of its circulating serotypes in Assam, North East India., Materials and Methods: A total of 80 clinical samples were collected from June 2019 to May 2020 from patients clinically suspected from pneumococcal infection and also included samples routinely sent to bacteriology laboratory. Isolation and identification of S. pneumoniae was performed using conventional culture and molecular methods. Antibiotic susceptibility patterns were monitored. Capsular serotyping was performed using PCR of cpsA gene followed by DNA sequencing., Results: Majority of the cases suspected of pneumococcal infection belong to the paediatric group aged less than 5 years. Out of 80 samples, 10 (12.50%) were found to be positive by PCR of recP gene. Culture was positive in 80% (8/10) of the total positives. Co-trimoxazole resistance was seen in 33.33% of the isolate from sputum. Serotypes 6A, 6B, 6C and 19F were detected in our region, out of which 6C is a non-vaccine serotype., Conclusion: Continued surveillance is needed to monitor trends in non-vaccine serotypes that may emerge as highly associated with antibiotic resistance. Also, the need to continuous monitoring of the antibiotic susceptibility of S. pneumoniae in North eastern parts of India is of outmost importance., Competing Interests: Conflict of interest None declared., (Copyright © 2023 Indian Association of Medical Microbiologists. Published by Elsevier B.V. All rights reserved.)

Background: Invasive bacterial disease (IBD; including pneumonia, meningitis, sepsis) is a major cause of morbidity and mortality in children in low-income countries., Methods: We analyzed data from a surveillance study of suspected community-acquired IBD in children <15 years of age in Kathmandu, Nepal, from 2005 to 2013 before introduction of pneumococcal conjugate vaccines (PCV). We detailed the serotype-specific distribution of invasive pneumococcal disease (IPD) and incorporated antigen and PCR testing of cerebrospinal fluid (CSF) from children with meningitis., Results: Enhanced surveillance of IBD was undertaken during 2005-2006 and 2010-2013. During enhanced surveillance, a total of 7956 children were recruited of whom 7754 had blood or CSF culture results available for analysis, and 342 (4%) had a pathogen isolated. From 2007 to 2009, all 376 positive culture results were available, with 259 pathogens isolated (and 117 contaminants). Salmonella enterica serovar Typhi was the most prevalent pathogen isolated (167 cases, 28% of pathogens), followed by Streptococcus pneumoniae (98 cases, 16% pathogens). Approximately, 73% and 78% of pneumococcal serotypes were contained in 10-valent and 13-valent PCV, respectively. Most cases of invasive pneumococcal disease (IPD) were among children ≥5 years of age from 2008 onward. Antigen and PCR testing of CSF for pneumococci, Haemophilus influenzae type b and meningococci increased the number of these pathogens identified from 33 (culture) to 68 (culture/antigen/PCR testing)., Conclusions: S. enterica serovar Typhi and S. pneumoniae accounted for 44% of pathogens isolated. Most pneumococcal isolates were of serotypes contained in PCVs. Antigen and PCR testing of CSF improves sensitivity for IBD pathogens., Competing Interests: The authors have no conflicts of interest to disclose., (Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.)

Study Objective: This study sought to describe the clinical presentation of normocellular community-acquired bacterial meningitis in adults., Methods: Using the prospective, nationwide, population-based database of the Danish Study Group of Infections of the Brain, the study identified all adults with normocellular community-acquired bacterial meningitis who were treated at departments of infectious diseases in Denmark from 2015 through 2018. Normocellular community-acquired bacterial meningitis was defined as a cerebrospinal fluid leukocyte count of up to 10×10 6 /L combined with detection of bacteria in the cerebrospinal fluid. Outcome was categorized according to the Glasgow Outcome Scale at discharge., Results: Normocellular cerebrospinal fluid was observed in 12 of 696 (2%) patients with community-acquired bacterial meningitis. The median age was 70 years (range 17 to 92 years), and 8 of 12 (67%) patients were male. All patients had symptoms suggestive of community-acquired bacterial meningitis and pathogens identified by culture (Streptococcus pneumoniae, n=10; Staphylococcus aureus, n=1) or polymerase chain reaction (Neisseria meningitidis; n=1) of the cerebrospinal fluid. Bacteremia was found in 9 of 12 (75%) patients, and 1 of 12 (8%) presented with septic shock. None of the patients had serious underlying immunocompromising conditions. The median times from admission to lumbar puncture and meningitis treatment were 2.5 hours (interquartile range 1.1 to 3.9 hours) and 2.6 hours (interquartile range 0.9 to 22.8 hours). In 3 of 11 (27%) patients, empiric treatment for community-acquired bacterial meningitis was interrupted by a normal cerebrospinal fluid cell count. The overall case-fatality rate was 3 of 12 (25%); meningitis treatment was interrupted in 1 of these patients, and 8 of 12 (67%) had a Glasgow Outcome Scale score of 1 to 4 at discharge., Conclusion: Normocellular community-acquired bacterial meningitis is not very common, but it is important to consider and may be associated with a pneumococcal cause., (Copyright © 2020 American College of Emergency Physicians. Published by Elsevier Inc. All rights reserved.)

Background: Patients with humoral immune deficiency are susceptible to invasive pneumococcal disease (IPD). This study estimates the prevalence of underlying hypogammaglobulinemia in admitted IPD cases and examines whether IPD cases had received preventative treatment., Methods: All adult IPD cases (Streptococcus pneumoniae in blood or cerebrospinal fluid) admitted to The Ottawa Hospital (TOH) from January 2013 to December 2015 were identified through the Eastern Ontario Regional Laboratory. Documented clinical demographics, S. pneumoniae serotype, serum immunoglobulins measured previously or in convalescence, and vaccination status of the cases were collected retrospectively for descriptive analyses., Results: There were 134 IPD in 133 patients (47.4% male; mean age 63, standard deviation [SD] = 15.6 years) during a 3-year observation period. All-cause mortality rate was 22.6% over a mean follow-up time of 362, SD = 345 days. Fifty-seven patients (42.9%) had serum immunoglobulin levels measured. Eighteen were either found to have hypogammaglobulinemia in convalescence (8/18) or previously known to have hypogammaglobulinemia (10/18). None of the known hypogammaglobulinemic patients had received antibiotic prophylaxis and/or immunoglobulin replacement therapy within 4 months prior to IPD. The high and low estimates of prevalence of hypogammaglobulinemia were 31.6% (of all measured) and 13.5% (of all cases). Among 18 patients with hematological malignancies in our cohort, 13 had hypogammaglobulinemia. Many isolates were vaccine serotypes; however, only 8 had documented previous pneumococcal vaccination., Conclusions: IPD has high mortality, and hypogammaglobulinemia was present in at least 13.5% of IPD cases. Secondary hypogammaglobulinemia is especially common in cases with hematological malignancy and IPD., (© The Author(s) 2017. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

Cerebrospinal fluid/serum albumin ratio is one of the most informative parameters for blood-brain barrier (BBB) integrity in cases of central nervous system (CNS) infectious diseases. Normally, CNS albumin concentration is a function of diffusion processes along with CSF drainage and resorption. In pathological processes CSF albumin levels are dependent only on the rate of CSF drainage resulting in non-linear reciprocal changes of albumin quotient (Qalb). IgG, IgA and IgM concentrations both in CSF and serum can be compared to Qalb, thus determining the intrathecal immune response. The aim of the study was to detect BBB permeability impairment and the intrathecal immune response in patients with CNS infections with various etiologies. CSF/serum ratios were calculated and related to IgG IgA and IgM concentrations in CSF and blood serum. The results were integrated and presented by Reibergrams. The results demonstrated typical patterns which prove albumin to be the main modulator of protein dynamics and at the same time explicates the complex pathophysiological mechanisms involved in BBB disruption and intrathecal immune response in CNS infections. The diagnostic model presented in our study seeks to explain the observations of meningitis and meningoencephalitis pathophysiology and points out the mandatory cooperation between clinicians and laboratory for accurate diagnosis and proper treatment.

Neisseria meningitidis (N. meningitidis), Streptococcus pneumoniae (S. pneumoniae), and Haemophilus influenzae type b (Hib) are three most common pathogens accounting for most bacterial meningitis, a serious global infectious disease with high fatality, especially in developing nations. Because the treatment and antibiotics differ among each type, the identification of the exact bacteria causing the disease is vital. Herein, we report a polymer/paper hybrid microfluidic biochip integrated with loop-mediated isothermal amplification (LAMP) for multiplexed instrument-free diagnosis of these three major types of bacterial meningitis, with high sensitivity and specificity. Results can be visually observed by the naked eye or imaged by a smartphone camera under a portable UV light source. Without using any specialized laboratory instrument, the limits of detection of a few DNA copies per LAMP zone for N. meningitidis, S. pneumoniae and Hib were achieved within 1h. In addition, these three types of microorganisms spiked in artificial cerebrospinal fluid (ACSF) were directly detected simultaneously, avoiding cumbersome sample preparation procedures in conventional methods. Compared with the paper-free non-hybrid microfluidic biochip over a period of three months, the hybrid microfluidic biochip was found to have a much longer shelf life. Hence, this rapid, instrument-free and highly sensitive microfluidic approach has great potential for point-of-care (POC) diagnosis of multiple infectious diseases simultaneously, especially in resource-limited settings., (Copyright © 2016 Elsevier B.V. All rights reserved.)

For accurate and earlier detection of invasive pneumococcal serogroup/serotypes from India, we have rearranged the African sequence of multiplex PCR provided by the Centers for Disease Control and Prevention, USA. This modified approach can successfully be adapted for earlier serotype detection of 95% of the pneumococcal strains prevalent in India., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Background The aim of this study was to evaluate the clinical outcome of patients with bacterial meningitis following the introduction of dexamethasone treatment in Denmark. Methods Adult patients with bacterial meningitis, admitted from 2003-2010 to two different university hospitals, were included retrospectively. Data at clinical presentation, Glasgow outcome scale (GOS), cerebrospinal fluid and blood biochemistry were collected. Relative risk (RR) with 95% confidence interval (CI) was computed by Cox proportional hazard regression analysis. Results One hundred and forty-seven patients were included in the study. The population had a median age of 62 years and 31% had an immunosuppressive co-morbidity. Eighty-nine patients had an unfavourable outcome (GOS score = 1-4). Adjuvant treatment with corticosteroids (RR = 0.48; 95% CI = 0.30-0.76) was associated with a favourable outcome (GOS score = 5), while altered mental status (RR = 2.36; 95% CI = 1.17-4.78) and age (RR = 1.03; 95% CI = 1.01-1.04) per year increment was associated with an unfavourable outcome. Adjuvant corticosteroid treatment did not affect short- or long-term survival. Short-term mortality was influenced by age (RR = 1.06; 95% CI = 1.04-1.09). Long-term mortality was influenced by age (RR = 1.06; 95% CI = 1.03-1.08) and female sex (RR = 1.81; 95% CI = 1.05-3.14). Conclusion This study indicated that adjuvant corticosteroid treatment in acute bacterial meningitis improves the outcome and can safely be administered in an elderly population with high levels of immunosuppressive co-morbidity.

Objectives/hypothesis: The objectives of this study were to examine the presence of β-2 transferrin (β2TRNSF) in cerebrospinal fluid (CSF) contaminated in vitro by various bacteria and explore the mechanism (passive or active) responsible for β2TRNSF elimination. Early diagnosis of CSF leakage may change treatment decisions and minimize the risk of meningitis and encephalitis. β2TRNSF is a protein present exclusively in CSF. Its detection is highly useful in cases of CSF leakage, although it has never been examined in the presence of central nervous system infection., Study Design: Prospective patient analysis., Methods: Sterile CSF drawn from patients was contaminated in vitro with several microorganisms chosen for their ability to cause neurosurgical-related infections: Streptococcus pneumoniae, methicillin-sensitive Staphylococcus aureus, Staphylococcus epidermidis, and Pseudomonas aeruginosa. β2TRNSF was examined at two time points: following immediate inoculation (t0) and following an overnight incubation (t18) over various bacterial concentrations. Samples of CSF inoculated with S pneumoniae were also examined in the presence of ciprofloxacin. For β2TRNSF analysis we used immunoblotting electrophoresis and enzyme-linked immunosorbent assay (ELISA)., Results: CSF samples collected from nine patients were analyzed. β2TRNSF was not detected following S pneumoniae inoculation at both time points when immunoblotting electrophoresis was used. Quantitative analysis using ELISA demonstrated significant β2TRNSF concentration decrease. The addition of ciprofloxacin led to the same results., Conclusions: CSF leak detection using β2TRNSF may be deceiving in the presence of a S pneumoniae cerebral nervous system infection. A passive process is suggested, as β2TRNSF disappeared either immediately or following incubation with inactive bacteria., (© 2014 The American Laryngological, Rhinological and Otological Society, Inc.)

Invasive disease caused by Streptococcus pneumoniae is a major cause of morbidity and mortality in high-risk individuals with severe comorbidities, including asplenia, chronic alcoholism, and altered immune status. The risk of invasive pneumococcal disease has been significantly higher in transplant patients compared with the general population. Here, we report an unusual case of a disseminated pneumococcal infection with meningitis, endocarditis, spondylodiscitis, and muscle abscess in an asplenic patient on chronic immunosuppressive therapy for liver transplantation performed 17 years before., (© 2013 John Wiley & Sons A/S.)

We examined associations of serotypes with multilocus sequence typing (MLST) data for 7 housekeeping genes and the genotype concerning penicillin resistance based on penicillin-binding protein (PBP) alterations in Streptococcus pneumoniae isolates from children with meningitis. From throughout Japan, we collected 115 pneumococcal isolates from the cerebrospinal fluid of patients 15 years old or younger from January 2007 to December 2009. We then carried out serotyping, MLST, and genotypic classification. Isolates included 24 serotypes and 52 sequence types (STs) according to MLST, of which 18 were novel. The 4 predominant serotypes included a variety of STs: 14 STs in serotype 6B (n = 24), 2 STs in 19F (n = 17), 6 STs in 23F (n = 14), and 5 STs in 14 (n = 11). Resistance genotypes included 6 types: 44.3% for gPRSP (pbp1a + 2x + 2b), 13.9% for gPISP (pbp1a + 2x), 9.6% for gPISP (pbp2x + 2b), 19.1% for gPISP (pbp2x), 3.5% for gPISP (pbp2b), and 9.6% for gPSSP. Interestingly, the most prevalent serotype of 6B included 7 newly identified STs and a variety of genotypes for resistance. STs in serotypes 23F and 14 were highly diverse, but not in 19F. These results suggest that various genetic elements in S. pneumoniae might be intrinsically susceptible to genetic mutations and recombination, with acceleration of emergence reflecting selection pressures such as antibiotic overuse.

Background: Highly active antiretroviral treatment (HAART) programs have been associated with declines in the burden of invasive pneumococcal disease (IPD) in industrialized countries. The aim of this study was to evaluate trends in IPD hospitalizations in HIV-infected adults in Soweto, South Africa, associated with up-scaling of the HAART program from 2003 to 2008., Methods: Laboratory-confirmed IPD cases were identified from 2003 through 2008 through an existing surveillance program. The period 2003-04 was designated as the early-HAART era, 2005-06 as the intermediate-HAART era and 2007-08 as the established-HAART era. The incidence of IPD was compared between the early-HAART and established-HAART eras in HIV-infected and-uninfected individuals., Results: A total of 2,567 IPD cases among individuals older than 18 years were reported from 2003 through 2008. Overall incidence of IPD (per 100,000) did not change during the study period in HIV-infected adults (207.4 cases in the early-HAART and 214.0 cases in the established-HAART era; p = 0.55). IPD incidence, actually increased 1.16-fold (95% CI: 1.01; 1.62) in HIV-infected females between the early-and established-HAART eras (212.1 cases and 246.2 cases, respectively; p = 0.03). The incidence of IPD remained unchanged in HIV-uninfected adults across the three time periods., Conclusion: Despite a stable prevalence of HIV and the increased roll-out of HAART for treatment of AIDS patients in our setting, the burden of IPD has not decreased among HIV-infected adults. The study indicates a need for ongoing monitoring of disease and HAART program effectiveness to reduce opportunistic infections in African adults with HIV/AIDS, as well as the need to consider alternate strategies including pneumococcal conjugate vaccine immunization for the prevention of IPD in HIV-infected adults.

The intercellular adhesion molecule is a transmembrane glycoprotein belonging to the immunoglobulin superfamily. Serum and cerebrospinal fluid (CSF) soluble intercellular adhesion molecule 1 (sICAM-1) from normal control children as well as from children with Guillain-Barré syndrome (GBS), with Coxsackie A9 virus meningoencephalitis and with Streptococcus pneumoniae meningoencephalitis were studied. sICAM-1 was quantified using an immunoenzimatic assay and albumin using the immunodiffusion technique in both biological fluids. Increased sICAM-1 values in CSF in patients with GBS correspond to an increase of the albumin CSF/serum quotient. In contrast, in inflammatory diseases like S. pneumoniae and Coxsackie A9 virus meningoencephalitis an increased brain-derived fraction was observed. In particular cases these values are 60-65% and 70-75% respectively. The results indicate an additional synthesis of sICAM-1 in subarachnoidal space during central nervous system (CNS) inflammatory process. An important role of sICAM-1 in the transmigration of different cell types into CSF during CNS inflammation in children with S. pneumoniae and Coxsackie A9 meningoencephalitis may be suggested.

Background: Acute bacterial meningitis (ABM) is a serious disease that needs rapid diagnosis for an accurate treatment. The most important etiological agents are: Streptococcus pneumoniae, Neisseria meningitidis and Haemophilus influenzae type b. Overall pathogen detection rate in patients with ABM in Chile is 83%., Aim: To evaluate a Polymerase Chain Reaction (PCR) protocol for simultaneous detection of several pathogens in patients with ABM., Material and Methods: We designed and evaluated a multiplex PCR protocol for simultaneous specific genes identifications of S pneumoniae ( inverted exclamation markytA and ply genes), N meningitidis (ctrA, crgA) and H influenzae (bexA) in cerebrospinal fluid (CSF) samples from pediatric patients with suspected diagnosis of ABM. Sensitivity, specificity and minimum detection levels of DNA were determined. Amplifications ofrDNA 16S gene was done to confirm extraction of bacterial DNA., Results: Ninety nine CSF samples were studied, 90 from children with fever and negative CSF culture, and 9 from ABM and positive culture patients. The PCR protocol had a sensitivity of 89%, specificity of 100%, positive predictive value 100% and negative predictive value 99%., Conclusions: We observed a high concordance (89%) between bacteriological cultures and the PCR protocol results. This diagnostic tool could increase identification of agents in specific settings such as patients previously treated with antibiotics.

Background: Chemokines play an important role in the recruitment and regulation of leukocyte traffic during bacterial infection. The aims of this study were to investigate the chemokine response to invasive pneumococcal disease (IPD) and to examine the influence of HIV infection on the chemokine response, pneumococcal bacterial loads, and outcome., Methods: We prospectively studied 95 children with IPD, and blood and cerebrospinal fluid (CSF) samples were taken at admission for the determination of chemokines, interferon-gamma (IFNgamma), and pneumococcal bacterial loads., Results: Plasma CXCL8 and CCL2, CSF CXCL8 and CCL4, and IFNgamma were significantly higher in HIV-infected children than in HIV-uninfected children. Blood and CSF pneumococcal bacterial loads correlated with plasma and CSF chemokines, respectively, and were higher in HIV-infected children compared with HIV-uninfected children. Among HIV-infected children, plasma concentrations of CXCL8 and CCL2 were significantly higher in nonsurvivors than in survivors, but CCL5 was significantly lower. HIV-infected and HIV-uninfected children with IPD had higher concentrations of chemokines (except CCL5) than acutely ill HIV-infected and HIV-uninfected children with no detectable bacterial infection. Male gender and low plasma CCL2 concentrations were shown to be independently associated with survival., Conclusions: Chemokines, in particular CCL2, are associated with survival in IPD and correlate with pneumococcal bacterial loads, disease presentation, and outcome.

Introduction: Outcomes following bacterial meningitis are significantly improved by adjunctive treatment with corticosteroids. However, little is known about the levels and significance of intrathecal endogenous cortisol. The aim of this study was to assess cortisol as a biological and diagnostic marker in patients with bacterial meningitis., Methods: Forty-seven consecutive patients with bacterial meningitis and no prior treatment were evaluated. For comparison, a group of 37 patients with aseptic meningitis and a group of 13 healthy control individuals were included., Results: The mean age of the bacterial meningitis patients was 42 years, and the mean Glasgow Coma Scale, Acute Physiology and Chronic Health Evaluation II, and Sequential Organ Failure Assessment scores on admission were 12, 13 and 4, respectively. Altogether, 40 patients (85%) were admitted to the intensive care unit, with a median (interquartile range) length of stay of 8 (4 to 15) days. A bacterial etiology was confirmed in 35 patients (74%). The median (interquartile range) cortisol concentration in cerebrospinal fluid (CSF) was 133 (59 to 278) nmol/l. CSF cortisol concentrations were positively correlated with serum cortisol levels (r = 0.587, P < 0.001). Furthermore, CSF cortisol levels correlated with Acute Physiology and Chronic Health Evaluation II score (r = 0.763, P < 0.001), Sequential Organ Failure Assessment score (r = 0.650, P < 0.001), Glasgow Coma Scale score (r = -0.547, P < 0.001) and CSF lactate levels (r = 0.734, P < 0.001). CSF cortisol was only weakly associated with intrathecal levels of IL-6 (r = 0.331, P = 0.02) and IL-8 (r = 0.296, P < 0.05). CSF cortisol levels in bacterial and aseptic meningitis significantly differed (P < 0.001). The CSF cortisol concentration of 46.1 nmol/l was found to be the optimal cutoff value for diagnosis of bacterial meningitis., Conclusion: CSF cortisol levels in patients with bacterial meningitis are highly elevated and correlate with disease severity. Moreover, our findings also suggest that intrathecal cortisol may serve as a valuable marker in discriminating between bacterial and aseptic meningitis.

The aim of our study was to evaluate a frequency of isolation and susceptibility to antibiotics of Streptococcus pneumoniae penicillin resistant among 154 strains S. pneumoniae isolated between 2003 and 2006 in University Hospital of Dr. A. Jurasza in Bydgoszcz. Antimicrobial susceptibility was assessed by disc-diffusion method according to the guidelines of Clinical and laboratory Standards Institute and The national Reference Centre for Antimicrobial Susceptibility. Minimal inhibitory concentrations for penicillin and cefotaxime were assessed by E-test method. Study shows increasing isolation of SPPR strains from 8,2% in 2003 to 32,0% in 2006. Strains were mostly isolated from patients ofNeurosurgery and Neurotraumatology Clinic and Rehabilitation Clinic. SPPR strains were mainly isolated from respiratory tract. Over 68% of SPPR showed intermediate resistance to penicillin and 73,3% of strains were susceptible to cefotaxime. Between 2003 and 2006 increased percentage of resistance strains to erythromycin, tetracycline and sulphometoxasol.

Pneumococci isolated from blood and cerebrospinal fluid from 1998 to 2001 in 2 counties in south-west Sweden were serotyped with the capsular reaction test. Of the 836 strains, 353 (42%), 598 (72%) and 789 (94%) belonged to serotypes included in the 7- and 11-valent pneumococcal conjugate vaccines and in the 23-valent polysaccharide vaccine, respectively. The most common serotype was type 1 (119 isolates) followed in descending frequency by serotypes 7F, 9V, 14, 4 and 12F (90-49 isolates per serotype). The coverage rates of the 7- and 11-valent conjugate vaccines among 58 strains isolated from children and adolescents 0-19 y of age were 46% and 93%, respectively. A comparison of clinical characteristics of infections caused by different serotypes showed that types 1 and 7F were less commonly associated with severe underlying diseases, that patients infected with these serotypes were younger than the average and, thus, had a lower case-fatality rate.

We monitored the susceptibility to penicillin of invasive strains of Streptococcus pneumoniae in two reference laboratories; 502 positive cultures from patients with an active invasive infectious process were analyzed. Streptococcus pneumoniae was identified through conventional procedures, and the oxacillin disc diffusion method was used to check for penicillin susceptibility. Statistical analysis included calculations of the frequency distribution, with 95% confidence intervals (CI), as well as chi-square tests and chi-square for linear trend for temporal analysis of susceptibility. The bacterium was isolated from patients less than a year old (40.7% of the isolates), from infants (55.9%), and from individuals less than 15 years old (64.4%). The majority (88.2%, 95% CI = 85.5%-91.1%) of the 502 isolates were susceptible to penicillin. There was no significant temporal trend of elevation of resistance rate during the study period (p=0.56). We conclude that resistance of S. pneumoniae to penicillin is not yet an important clinical-epidemiological concern in the State of Minas Gerais. To provide necessary support for the adoption of therapeutic and prophylactic measures, epidemiological surveillance should be implemented at a national level to monitor the profile of susceptibility/resistance of S. pneumoniae to penicillin and other antimicrobials.

Objective: To evaluate the precision and clinical applicability of the Bayer ADVIA 120 cytometer (Bayer Healthcare, Fernwald, Germany) for cerebrospinal fluid (CSF) cell count and differentiation., Study Design: One hundred six analyses of CSF from 98 patients by the ADVIA 120 were compared with routine cell count and microscopic differentiation. Correlation coefficients were calculated., Results: In general, the total cell counts of both methods correlated well. The best correlations were seen at higher cell counts, > or = 100 cells per microliter with < 100 erythrocytes per microliter. The best correlations of cell differentiation were seen for lymphocytes and neutrophils, while the results for monocytes and eosinophils were less precise. In some cases, considerable differences between automated and microscopic cell counts and differentiation were seen that were relevant to clinical decision making. The detection of pathologic cell types, such as hemosiderophages, mitoses and neoplastic cells, was not provided by automated cytometry., Conclusion: When experienced personnel are not available, a preliminary cell count and differentiation between neutrophilic and lymphocytic reactions by automated cytometry may be valuable in allowing initial therapeutic decision making. Since the detection of pathologic cell types is not provided and the precision at low cell counts is only moderate, a personal microscopic evaluation of each sample is still indispensable to avoid misdiagnoses.

Streptococcus pneumoniae is a leading cause of invasive bacterial disease. This is the first study to examine the expression of S. pneumoniae genes in vivo by using whole-genome microarrays available from The Institute for Genomic Research. Total RNA was collected from pneumococci isolated from infected blood, infected cerebrospinal fluid, and bacteria attached to a pharyngeal epithelial cell line in vitro. Microarray analysis of pneumococcal genes expressed in these models identified body site-specific patterns of expression for virulence factors, transporters, transcription factors, translation-associated proteins, metabolism, and genes with unknown function. Contributions to virulence predicted for several unknown genes with enhanced expression in vivo were confirmed by insertion duplication mutagenesis and challenge of mice with the mutants. Finally, we cross-referenced our results with previous studies that used signature-tagged mutagenesis and differential fluorescence induction to identify genes that are potentially required by a broad range of pneumococcal strains for invasive disease.

Objective: Evaluation of elevated cerebrospinal fluid levels of glutamate in children with bacterial meningitis as a predictor of seizures or other adverse outcomes., Design: Prospective cohort study with controls., Setting: A 36-bed pediatric intensive care unit and primary pediatric referral center., Patients: From 1999 to 2001, a total of 55 patients, between the ages of 0 and 18 yrs, with lumbar punctures performed for suspected meningitis., Measurements and Main Results: A total of 23 patients had bacterial meningitis confirmed by cerebrospinal fluid/blood culture and elevated cerebrospinal fluid white blood cell counts, and 32 patients, who tested negative, were included as controls. The median age for the patients with meningitis was 1.0 yr (range, 0.0-15.2 yrs), and in the culture-negative group (control group), the median age was 0.3 yrs (range, 0.0-17.0 yrs). The average cerebrospinal fluid white blood cell count was 2707 +/- 3897 in the group with bacterial infection, whereas in the control group, the average was 148 +/- 259 (p < .01). Patients with bacterial meningitis had a mean cerebrospinal fluid glutamate level of 60.5 +/- 88.4 mol/L, whereas the mean cerebrospinal fluid glutamate level in the control group was 4.9 +/- 11.0 mol/L (p < .01). However, only 10 of 23 children with bacterial meningitis had a second lumbar puncture performed during the study. There was no correlation between the cerebrospinal fluid white blood cell count and cerebrospinal fluid glutamate levels in either the study or control patients. None of the control patients developed seizures or neurologic deficits, despite some patients having elevated glutamate levels. However, four patients with bacterial meningitis developed seizures after admission to the hospital, and ten were discharged with at least some neurologic sequelae attributable to their infection. Two out of the three who developed seizures and had a repeat lumbar puncture demonstrated persistent elevation of cerebrospinal fluid glutamate levels. In addition, 70% of patients (7 of 10) with Streptococcus pneumoniae meningitis developed neurologic complications (p = .04)., Conclusions: Bacterial meningitis in children causes an increase in cerebrospinal fluid glutamate that in many cases persists over time. However, in this limited study, neither higher nor persistent elevation of cerebrospinal fluid glutamate levels is predictive of which patients might develop seizures or other apparent immediate adverse outcomes after invasive infection. The responsible organism seems to have far more significance in predicting the development of adverse sequelae.

The variability of the course of infection by Streptococcus pneumoniae is well known but poorly understood. Most animal models of pneumonia, sepsis or meningitis have been forced to use site-specific bacterial inoculation to mimic localized human infection. This study examined the differences in the progression of disease-causing strains D39 (serotype 2), A66.1 (serotype 3) and TIGR4 (serotype 4) using isolates transformed with the Gram-positive lux transposon cassette, Tn4001 luxABCDE Km(r). Expression of the lux operon results in bioluminescence, permitting the detection of the bacteria within a living animal while using a CCD camera. Mice infected intranasally with A66.1 developed only pneumonia, those challenged with D39 experienced high-grade sepsis, while TIGR4 infection resulted in low-grade pneumonia and bacteremia ultimately progressing to meningitis. Quantitative analysis of bacterial titers confirmed these patterns, which were consistent across different lineages of mice. Mice anesthetized with ketamine and xylazine developed more severe forms of the disease compared with isoflurane. These studies unambiguously characterize 3 distinct models of the natural course of pneumococcal infection. Mapping these models provides a framework for detailed molecular modeling of pneumococcal virulence determinants at specific stages of disease.

Using two different animal models of Streptococcus pneumoniae infection, we have demonstrated that this organism is able to spread to the central nervous system and cause meningitis by bypassing the bloodstream. Following respiratory tract infection induced via intranasal inoculation, bacteria were rapidly found in the bloodstream and brains in the majority of infected mice. A similar pattern of dissemination occurred following otitis media infection via transbullar injection of gerbils. However, a small percentage of animals infected by either route showed no bacteria in the blood and yet did have significant numbers of bacteria in brain tissue. Subsequent experiments using a galU mutant of S. pneumoniae, which is impaired in its ability to disseminate to the bloodstream following infection, showed that this organism is able to spread to the brain and cerebrospinal fluid. These results demonstrate that, unlike many bacterial pathogens that cause meningitis, S. pneumoniae is able to do so independent of bloodstream involvement upon different routes of infection. This may address the difficulty in treating human infections caused by this organism.

Objectives: Interleukin-6 (IL-6) is elevated in the cerebrospinal fluid (CSF) of humans and animals with bacterial meningitis. This study's hypothesis was that anti-IL-6 antibodies will attenuate meningeal inflammation in a rat model of bacterial meningitis., Methods: 14 male Sprague-Dawley rats were inoculated intracisternally (IC) with 0.1 mL of heat-killed pneumococci. At one hour post-inoculation, the rats received intraperitoneal doses of either 1.0 mL phosphate-buffered saline (PBS treatment group, n = 7) or 70 microg anti-IL-6 antibodies in 1.0 mL PBS (anti-IL-6 antibody treatment group, n = 7). Nine rats (normal group, n = 9) had no inoculation, and four rats (surgical sham group, n = 4) had IC inoculations of saline. At six hours post-inoculation, all the animals had CSF removed via IC tap. The CSF protein and white blood cell (WBC) count measures were compared using a t-test., Results: Mean CSF WBC for the anti-IL-6 treatment group was 2,458/microL, versus the PBS controls' mean of 9,697/microL (p = 0.007). Mean CSF protein for the anti-IL-6 group was 180 mg/dL, versus 296 mg/dL for the controls (p = 0.032). The surgical sham and normal animals had normal CSF WBC and protein values., Conclusions: In this rat meningitis model, systemic treatment with anti-IL-6 antibodies after the induction of meningitis suppressed both CSF WBC count and CSF protein level, two important indices of meningeal inflammation.

Effective laboratory animal models of bacterial meningitis are needed to unravel the pathophysiology of this disease. Previous models have failed to simulate human meningitis by using a directly intracerebral route of infection. Hyaluronidase is a virulence factor of Streptococcus pneumoniae. In this study, a novel model of murine meningitis is described. Intranasal administration of S. pneumoniae with hyaluronidase induced meningitis in 50% of inoculated mice, as defined by a positive cerebrospinal fluid (CSF) culture and an inflammatory infiltrate in the meninges. None of the mice inoculated without hyaluronidase developed meningitis. Hyaluronidase was found to facilitate pneumococcal invasion of the bloodstream after colonization of the upper respiratory tract. Meningitis was characterized by pleocytosis of CSF and the induction of proinflammatory cytokines and CXC chemokines in brain tissue. These results indicate that this murine model mimics important features of human disease and allow for the use of this model for studying issues related to the pathophysiology and the treatment of pneumococcal meningitis.

From January 1991 through December 1998, a total of 1046 pneumococcal isolates were received from 23 laboratories participating in the statewide surveillance system. Of these, 1037 were recovered from normally sterile sites (blood and cerebrospinal and pleural fluid) and were available for serotyping and susceptibility testing. Ninety-two percent of these isolates were serotypes represented in the 23-valent pneumococcal polysaccharide vaccine. Serotypes in the 7-valent pneumococcal conjugate vaccine (4, 6B, 9V, 14, 18C, 19F, and 23F) were recovered from 72% of Alaska Natives and 84% of non-Native children <5 years old with invasive disease. Statewide, 7.3% and 3.2% of isolates had intermediate and high levels of resistance to penicillin, respectively; 9.2% were resistant to erythromycin (minimal inhibitory concentration, >/=1 microg/mL) and 19% to trimethoprim/sulfamethoxazole (minimal inhibitory concentration, >/=4/76 microg/mL). Twelve percent of invasive isolates were resistant to >/=2 classes of antibiotics; of these, serotype 6B accounted for 33%, and 63% were recovered from children <5 years old.

The bactericidal activities of monotherapy with trovafloxacin (-0.37 +/- 0.15 Delta log(10) CFU/ml. h), vancomycin (-0.32 +/- 0.12 Delta log(10) CFU/ml. h), and ceftriaxone (-0.36 +/- 0.19 Delta log(10) CFU/ml. h) for the treatment of experimental meningitis in rabbits due to a clinical penicillin-resistant pneumococcal strain (MIC, 4 mg/liter) were similar. The combination of ceftriaxone with trovafloxacin considerably improved the killing rates (-0.67 +/- 0.16 Delta log(10) CFU/ml. h) and was slightly superior to ceftriaxone with vancomycin (killing rate, -0.53 +/- 0. 22 Delta log(10) CFU/ml. h), the regimen most commonly used in clinical practice. In vitro, synergy was demonstrated between ceftriaxone and trovafloxacin by the checkerboard method (fractional inhibitory concentration index, 0.5) and by time-killing assays over 8 h.

The main object of this study was to describe the features of antibiotic resistance in pneumococci from children in four regions of France in 1995. Despite the high prevalence (40%) of pneumococci with diminished susceptibilty to penicillin (PDSP), resistance to amoxycillin (0.8%) and cefotaxime (0.4%) was rare; 16% of pneumococci were resistant to penicillin G (PRP, MIC > 1 mg/l). PDSP showed the expected resistance to macrolides (67%) and cotrimoxazole (57%) and were predominantly serotypes 23F, 14, 9 and 6. This study by the Regional Pneumococcal Observatories confirms the high prevalence and the main characteristics of antibiotic resistance among pneumococci isolated from children. Nevertheless, the resistance to all antibiotics was lower than that found in French multicentre, nationwide surveys, possibly because of differences in the mode of strain collection and geographic origin.

Murine experimental meningitis models induced by either Escherichia coli LPS, live Streptococcus pneumoniae, or Listeria monocytogenes were used to study the origin and potential function of soluble CD14 (sCD14) in the brain during bacterial meningitis. Whereas intracerebral infection caused only a minor and/or transient increase of sCD14 levels in the serum, dramatically elevated concentrations of sCD14 were detected in the cerebrospinal fluid. Reverse-transcriptase PCR and FACS analysis of the leukocytes invading the subarachnoid compartment revealed an active amplification of CD14 transcription and concomitant surface expression. These findings were confirmed by in situ hybridization and immunohistochemical analysis. In contrast, parenchymal astrocytes and microglial cells were shown not to significantly contribute to the elevated levels of sCD14. Simultaneous intracerebral inoculation of rsCD14 and S. pneumoniae resulted in a markedly increased local cytokine response. Taken together, these data provide the first evidence that sCD14 can act as an inflammatory co-ligand in vivo. Thus, during bacterial meningitis, sCD14 is massively released by intrathecal leukocytes, and the sCD14 found in the cerebrospinal fluid can play an important role in the pathogenesis of this disease.

Basic fibroblast growth factor (bFGF), a neurotrophic factor in the CNS, is expressed at high levels in response to seizures or strokes. We examined the expression of bFGF during experimental bacterial meningitis and the levels of bFGF in the cerebrospinal fluid (CSF) of children with bacterial meningitis. For the experimental study, a mouse model of meningitis was established by intracranial injection of Streptococcus pneumoniae. Twenty-four hours after induced meningitis, the brains were sectioned and stained immunohistochemically for bFGF. Neutrophils and macrophages infiltrating the leptomeninges and the ventricles exhibited strong bFGF immunoreactivity. The neurons in the areas adjacent to the inflamed ventricles also showed enhanced bFGF expression. For the clinical study, we used an enzyme immunoassay to measure bFGF in CSF in 18 children with bacterial meningitis, 12 with aseptic meningitis, and 18 controls. The CSF levels of bFGF were twice as high in children with bacterial meningitis (medians 6.75-7.21 pg/mL) compared with those who had aseptic meningitis (2.9 pg/mL) or in control subjects (2.65 pg/mL, p < 0.0001, respectively). In patients with bacterial meningitis who survived, CSF bFGF decreased significantly after 24-50 h of antibiotic therapy (p < 0.0005). Patients who developed major sequelae or died had much higher levels of CSF bFGF than those without (134.9 pg/mL versus 7.38 pg/mL, p < 0.05). These findings of enhanced immunoreactivity of bFGF in experimental bacterial meningitis and an association of CSF levels of bFGF with disease severity in childhood bacterial meningitis suggest a biologic role for this neurotrophic factor in the pathophysiology of bacterial meningitis.

The activity of moxifloxacin (BAY 12-8039) against a Streptococcus pneumoniae type 3 strain (MIC and minimum bactericidal concentration [MBC] of moxifloxacin, 0.06 and 0.25 microgram/ml, respectively; MIC and MBC of ceftriaxone, 0.03 and 0.06 microgram/ml, respectively) was determined in vitro and in a rabbit model of meningitis. Despite comparable bactericidal activity, 10 micrograms of moxifloxacin per ml released lipoteichoic and teichoic acids less rapidly than 10 micrograms of ceftriaxone per ml in vitro. Against experimental meningitis, 10 mg of moxifloxacin per kg of body weight per ml reduced the bacterial titers in cerebrospinal fluid (CSF) almost as rapidly as ceftriaxone did (mean +/- standard deviation, -0.32 +/- 0.14 versus -0.39 +/- 0.11 delta log CFU/ml/h). The activity of moxifloxacin could be described by a sigmoid dose-response curve with a maximum effect of -0.33 delta log CFU/ml/h and with a dosage of 1.4 mg/kg/h producing a half-maximal effect. Maximum tumor necrosis factor activity in CSF was observed later with moxifloxacin than with ceftriaxone (5 versus 2 h after the initiation of treatment). At 10 mg/kg/h, the concentrations of moxifloxacin in CSF were 3.8 +/- 1.2 micrograms/ml. Adjunctive treatment with dexamethasone at 1 mg/kg prior to the initiation of antibiotic treatment only marginally reduced the concentrations of moxifloxacin in CSF (3.3 +/- 0.6 micrograms/ml). In conclusion, moxifloxacin may qualify for use in the treatment of S. pneumoniae meningitis.

A seminested-PCR assay, based on the amplification of the pneumococcal penicillin-binding protein 2B gene (pbp2B), was developed for the detection of penicillin-resistant and -susceptible pneumococci in cerebrospinal fluid (CSF) specimens. Species-specific primers (P5 and P6) which amplified a 682-bp conserved region of the transpeptidase-encoding region of the pbp2B gene were used. Four "resistance" primers were designed to bind to altered areas of the pbp2B gene identified in penicillin-resistant South African wild-type strains. Together with the downstream primer P6, the upstream resistance primers amplified fragments which were used to detect the presence of penicillin resistance. This system identified all 35 of the S. pneumoniae isolates evaluated, including strains of 11 different serotypes and a range of penicillin-resistant and -susceptible strains. The specificity of the assay was demonstrated by its inability to amplify DNA from other bacterial species which commonly cause meningitis. It was possible to detect pneumococcal DNA from culture-negative CSF inoculated with 2.5 pg of purified DNA or 18 CFU. Analysis of 285 CSF specimens showed that PCR detected the pneumococcus in 18 samples positive by culture, including the identification of four penicillin-resistant isolates. The positive predictive value and the negative predictive value of the assay were each 100%.

N-acetylmuramyl-L-alanine amidase (NAMLAA) specifically degrades peptidoglycan, a major component of bacterial cell walls. Lysozyme degrades peptidoglycan differently by hydrolyzing the aminosugar backbone of peptidoglycan. In another study, it was shown that the two enzymes act synergistically to inactivate the inflammatory properties of peptidoglycan. The presence of lysozyme and NAMLAA was determined in serum and cerebrospinal fluid (CSF) of patients with bacterial meningitis. High concentrations of lysozyme were found in CSF while, surprisingly, NAMLAA was not present. To explain this phenomenon, the degranulation pattern of neutrophils in CSF was compared with that of neutrophils from blood. Specific granules contain lysozyme and the azurophil granules contain both lysozyme and NAMLAA. CD66b expression on the cell surface, indicative for fusion of the specific granules with the cell membrane, was higher in CSF than in blood, while the marker for the azurophil granules was lower.

A laboratory-based study of diagnosed bacterial meningitis in the national paediatric hospital identified 852 cases of meningitis in the 8-year period 1987-1994. There were 587 culture-positive cases, of which Haemophilus influenzae (47%) and Streptococcus pneumoniae (32%) accounted for 80%. H. influenzae showed a remarkable increase of 700% during the study period. Most of the H. influenzae cases (90%) occurred in the 1st 2 years of life. Analysis of culture-negative specimens by antigen detection in the last 2 years also revealed the predominance of H. influenzae (71.4%) and S. pneumoniae (22.4%). Typing of H. influenzae isolates during this time showed that 98% of the strains were type b. This retrospective hospital-based study indicated a pronounced increase in the incidence of H. influenzae meningitis and strongly supports the need for large-scale Hib vaccination for young children. However, such nationwide intervention will probably need to be based on a prospective on a prospective and population-based surveillance of H. influenzae infections.

The appearance of polymorphonuclear and mononuclear leukocytes in the cerebrospinal fluid (csf) is an important hallmark of bacterial meningitis. Chemokines are candidate mediators of cell migration from blood into the subarachnoid space. Therefore, concentrations of C-X-C and C-C chemokines in the csf of patients with pyogenic meningitis were measured by ELISA. Highly significant elevations of chemokine levels in comparison with noninflammatory csf controls were found for IL-8 (median, 21.6 ng/ml; range, < 0.1 to 191.3), growth-related gene product alpha (median, 5.6 ng/ml; range, < 0.1 to 48.2), monocyte chemotactic protein-1 (median, 26.4 ng/ml; range, < 0.2 to 193.8), macrophage inflammatory protein-1 alpha (MIP-1 alpha; median, 1.8 ng/ml; range, < 0.5 to 18.0), MIP-1 beta (median, 10.6 ng/ml; range, < 0.3 to 84.4), but not for RANTES (regulated upon activation, normal T cell expressed and secreted). The csf of bacterial meningitis were chemotactic for neutrophils and mononuclear leukocytes. Correlation analysis demonstrated a strong association between individual chemokine levels and chemotactic activity mediated by csf. A significant reduction of neutrophil chemotaxis was obtained by anti-IL-8 and anti-growth-related gene product alpha Abs, and a reduction of mononuclear cell migration was achieved by a combination of anti-monocyte chemotactic protein-1, anti-MIP-1 alpha, and anti-MIP-1 beta Abs. Since no significant correlation was found between csf leukocyte counts and chemokine concentrations or chemotactic activity mediated by csf, additional factors influence the extent of pleocytosis in vivo.

Streptococcus pneumoniae is one of the important human pathogens in clinical microbiology. A polymerase chain reaction assay was designed to detect and identify S. pneumoniae through amplification of the ribosomal DNA spacer regions between the pneumococcal 16S-23S ribosomal RNA genes. Thirty-two Streptococcus and non-Streptococcus strains were tested to verify the specificity of the assay, and only S. pneumoniae strains gave a positive reaction. This method is a powerful technique for the rapid identification of S. pneumoniae.

The serological properties of six new pneumococcal capsular types are described. A table listing all 90 pneumococcal types and their cross-reactions is included.

Capsular types of pneumococci from normally sterile body sites of 1,622 patients in Brazil were analyzed. Of 1,477 isolates from cerebrospinal fluid, 76.1% were of types represented in the currently available pneumococcal vaccine. The importance of age, time, and place in determining the optimal formulation of pneumococcal vaccine is considered.

Neuron-specific enolase (NSE) is one of the glycolytic enzymes distributed exclusively in neurons. It was measured serially in the cerebrospinal fluid (CSF) of 10 children with bacterial meningitis during the illness using radio-immunoassay. The relationship between CSF-NSE levels and neurological complications or outcome was examined. CSF-NSE levels were significantly higher in the patients with bacterial meningitis than in the patients with the other central nervous system (CNS) infectious diseases, suggesting that CNS damage in those patients with bacterial meningitis was exacerbated. As CSF-NSE levels increased to above 25 ng/mL in the acute phase, all patients except one had subdural effusion. In those patients whose CSF-NSE level rose again during the illness, CNS complications or sequelae occurred. CSF-NSE may be a useful prognostic factor for predicting CNS damage in childhood bacterial meningitis.

Lysozyme had no effect on the rate of multiplication of growing cultures of Streptococcus pneumoniae, but it greatly reduced the lag period that precedes autolysis of these bacteria in stationary phase. Several experiments were done to understand the mechanism of this effect. Lysozyme had no hydrolytic activity on intact cell walls, and cell walls of pneumococci grown with or without lysozyme had identical composition and susceptibility to the pneumococcal autolysin. The acceleration of stationary-phase autolysis by lysozyme involved triggering of the pneumococcal autolytic enzyme since lysozyme had no detectable effect on nonautolysing (LytA-) mutants and heat-inactivated lysozyme completely lacking enzymatic activity was as effective as the nondenatured enzyme in facilitating stationary-phase autolysis. The role of lysozyme in host defense against pneumococcal infection remains elusive.