Cerebrospinal fluid inflammatory markers to differentiate between neonatal bacterial meningitis and sepsis: A prospective study of diagnostic accuracy.

OBJECTIVES: We evaluated the diagnostic accuracy of cerebrospinal fluid (CSF) inflammatory markers for diagnosing bacterial meningitis in neonates with sepsis and/or meningitis. METHODS: Cases were identified from a prospective multicenter study including patients aged 0-3 months with Group B Streptococcal (GBS) or Escherichia coli culture positive sepsis/meningitis. CSF CXCL10, MDC, IL-6, IL-8, IL-10, TNF- α, MIF, IL-1RA, CXCL13, IL-1ß, CRP and procalcitonin concentrations were measured with Luminex technology. RESULTS: In 61/373 patients (17%) residual CSF from the lumbar puncture was available, of whom 16 (26%) had definitive meningitis, 15 (25%) probable meningitis and 30 (49%) had sepsis. All biomarkers were detectable in CSF and showed significantly higher concentrations in definitive meningitis versus sepsis patients and six biomarkers in probable meningitis versus sepsis patients. Discrimination between definitive meningitis and sepsis was excellent for IL-1RA (area under the receiver operating characteristic curve [AUC] 0.93), TNF-α (AUC 0.92), CXCL10 (AUC 0.90), IL-1ß (AUC 0.92), IL-6 (AUC 0.94), IL-10 (AUC 0.93) and a combination of IL-1RA, TNF-α, CXCL-10 and CSF leukocyte count (AUC 0.95). CSF leukocyte count remained the predictor with the highest diagnostic accuracy (AUC 0.96). CONCLUSION: CSF inflammatory markers can be used to differentiate between neonatal sepsis and meningitis.

Diagnostic Utility of Cerebrospinal Fluid White Blood Cell Components for the Identification of Bacterial Meningitis in Infants.

BACKGROUND: To evaluate the diagnostic and predictive utility of cerebrospinal fluid (CSF) white blood cell (WBC) components in the diagnosis of bacterial meningitis in infants discharged from the neonatal intensive care unit (NICU). METHODS: We identified a cohort of infants discharged from a Pediatrix NICU between 1997 and 2020 who did not have an immunodeficiency, had at least 1 CSF culture collected within the first 120 days of life, and at least 1 CSF laboratory specimen obtained on the day of culture collection. We only included an infant's first CSF culture and excluded cultures from CSF reservoirs and those growing contaminants or nonbacterial organisms. We examined the utility of CSF WBC components to diagnose or predict bacterial meningitis by calculating sensitivity, specificity, positive and negative predictive values, likelihood ratios, and area under the receiver operating curve (AUC) at different cutoff values for each parameter. We performed subgroup analysis excluding infants treated with antibiotics the day before CSF culture collection. RESULTS: Of the 20 756 infants that met the study inclusion criteria, 320 (2%) were diagnosed with bacterial meningitis. We found (AUC [95% CI]) CSF WBC count (0.76 [0.73-0.79]), CSF neutrophil count (0.74 [0.70-0.78]), and CSF neutrophil percent (0.71 [0.67-0.75]) had the highest predictive values for bacterial meningitis, even when excluding infants with early antibiotic administration. CONCLUSIONS: No single clinical prediction rule had the optimal discriminatory power for predicting culture-proven bacterial meningitis, and clinicians should be cautious when interpreting CSF WBC parameters in infants with suspected meningitis.

Comparison of bacterial culture with BioFire® FilmArray® multiplex PCR screening of archived cerebrospinal fluid specimens from children with suspected bacterial meningitis in Nigeria.

BACKGROUND: Diagnosis of bacterial meningitis remains a challenge in most developing countries due to low yield from bacterial culture, widespread use of non-prescription antibiotics, and weak microbiology laboratories. The objective of this study was to compare the yield from standard bacterial culture with the multiplex nested PCR platform, the BioFire® FilmArray® Meningitis/Encephalitis Panel (BioFire ME Panel), for cases with suspected acute bacterial meningitis. METHODS: Following Gram stain and bacterial culture on cerebrospinal fluid (CSF) collected from children aged less than 5 years with a clinical suspicion of acute bacterial meningitis (ABM) as defined by the WHO guidelines, residual CSF specimens were frozen and later tested by BioFire ME Panel. RESULTS: A total of 400 samples were analyzed. Thirty-two [32/400 (8%)] of the specimens were culture positive, consisting of; three Salmonella spp. (2 Typhi and 1 non-typhi), three alpha hemolytic Streptococcus, one Staphylococcus aureus, six Neisseria meningitidis, seven Hemophilus influenzae, 11 Streptococcus pneumoniae and 368 were culture negative. Of the 368 culture-negative specimens, the BioFire ME Panel detected at least one bacterial pathogen in 90 (24.5%) samples, consisting of S. pneumoniae, N. meningitidis and H. influenzae, predominantly. All culture positive specimens for H. influenzae, N. meningitidis and S. pneumoniae also tested positive with the BioFire ME Panel. In addition, 12 specimens had mixed bacterial pathogens identified. For the first time in this setting, we have data on the viral agents associated with meningitis. Single viral agents were detected in 11 (2.8%) samples while co-detections with bacterial agents or other viruses occurred in 23 (5.8%) of the samples. CONCLUSIONS: The BioFire® ME Panel was more sensitive and rapid than culture for detecting bacterial pathogens in CSF. The BioFire® ME Panel also provided for the first time, the diagnosis of viral etiologic agents that are associated with meningoencephalitis in this setting. Institution of PCR diagnostics is recommended as a routine test for suspected cases of ABM to enhance early diagnosis and optimal treatment.

[A case of tuberculous meningitis diagnosed early by direct Loop-Mediated Isothermal Amplification (LAMP) method using centrifuged medium of cerebrospinal fluid culture].

Tuberculous meningitis (TBM) is a central nervous system infection with a high mortality rate and requires early diagnosis and treatment. Identification of Mycobacterium tuberculosis in the cerebrospinal fluid is of primary importance in the diagnosis of TBM, however, conventional methods have some disadvantages: Rapid results tests such as smear and regular PCR method do not have sufficient diagnostic sensitivity; Nested PCR, which is one of the most sensitive tests, is not available in all facilities; Culture tests require a long period of 4-8 weeks for results. Here we report a case of TBM, diagnosed 14 days earlier than culture test by direct Loop-Mediated Isothermal Amplification (LAMP) method using centrifuged medium of cerebrospinal fluid (day 18) culture. The method we used here is simple, widely available, and considered to be useful for early detection of TBM.

Are cerebrospinal fluid biochemical parameters valid to predict positive results in microbiological molecular diagnostic platforms? A 4-year experience with the FilmArray® Panel Meningitis/Encephalitis for detection of community-acquired bacterial meningitis.

OBJECTIVE: To demonstrate whether the use of the FilmArray® Meningitis/Encephalitis panel (M/E) in the diagnosis of bacterial meningitis can be optimized based on the screening of cerebrospinal fluid biochemical parameters and whether there is a correlation between biochemical data and positive results of the technique. MATERIAL AND METHODS: We used data from La Paz University Hospital between September 5, 2017 and December 1, 2021, from patients who had had the FilmArray® panel M/E performed on cerebrospinal fluid samples. RESULTS: Bacterial meningitis was suspected in 63.9% of the patients; 38.15% had a positive result on the FilmArray® panel M/E, of which 68.97% were isolated in culture. Of the biochemical parameters studied in cerebrospinal fluid, white blood cell count, lactate, and protein were increased in PCR-positive patients, but glucose was decreased. DISCUSSION: Only lactate showed a significant contribution to the model, with a cut-off point of 4.65 mmol/L with a sensitivity of 81.5% and a specificity of 96.4%.

A Refined, Controlled 16S rRNA Gene Sequencing Approach Reveals Limited Detection of Cerebrospinal Fluid Microbiota in Children with Bacterial Meningitis.

Advances in both laboratory and computational components of high-throughput 16S amplicon sequencing (16S HTS) have markedly increased its sensitivity and specificity. Additionally, these refinements have better delineated the limits of sensitivity, and contributions of contamination to these limits, for 16S HTS that are particularly relevant for samples with low bacterial loads, such as human cerebrospinal fluid (CSF). The objectives of this work were to (i) optimize the performance of 16S HTS in CSF samples with low bacterial loads by defining and addressing potential sources of error, and (ii) perform refined 16S HTS on CSF samples from children diagnosed with bacterial meningitis and compare results with those from microbiological cultures. Several bench and computational approaches were taken to address potential sources of error for low bacterial load samples. We compared DNA yields and sequencing results after applying three different DNA extraction approaches to an artificially constructed mock-bacterial community. We also compared two postsequencing computational contaminant removal strategies, decontam R and full contaminant sequence removal. All three extraction techniques followed by decontam R yielded similar results for the mock community. We then applied these methods to 22 CSF samples from children diagnosed with meningitis, which has low bacterial loads relative to other clinical infection samples. The refined 16S HTS pipelines identified the cultured bacterial genus as the dominant organism for only 3 of these samples. We found that all three DNA extraction techniques followed by decontam R generated similar DNA yields for mock communities at the low bacterial loads representative of CSF samples. However, the limits of detection imposed by reagent contaminants and methodologic bias precluded the accurate detection of bacteria in CSF from children with culture-confirmed meningitis using these approaches, despite rigorous controls and sophisticated computational approaches. Although we did not find current DNA-based diagnostics to be useful for pediatric meningitis samples, the utility of these methods for CSF shunt infection remains undefined. Future advances in sample processing methods to minimize or eliminate contamination will be required to improve the sensitivity and specificity of these methods for pediatric meningitis. IMPORTANCE Advances in both laboratory and computational components of high-throughput 16S amplicon sequencing (16S HTS) have markedly increased its sensitivity and specificity. These refinements have better delineated the limits of sensitivity, and contributions of contamination to these limits, for 16S HTS that are particularly relevant for samples with low bacterial loads such as human cerebrospinal fluid (CSF). The objectives of this work were to (i) optimize the performance of 16S HTS in CSF samples by defining and addressing potential sources of error, and (ii) perform refined 16S HTS on CSF samples from children diagnosed with bacterial meningitis and compare results with those from microbiological cultures. We found that the limits of detection imposed by reagent contaminants and methodologic bias precluded the accurate detection of bacteria in CSF from children with culture-confirmed meningitis using these approaches, despite rigorous controls and sophisticated computational approaches.

Bacterial ribosomal RNA detection in cerebrospinal fluid using a viromics approach.

BACKGROUND: In patients with central nervous system (CNS) infections identification of the causative pathogen is important for treatment. Metagenomic next-generation sequencing techniques are increasingly being applied to identify causes of CNS infections, as they can detect any pathogen nucleic acid sequences present. Viromic techniques that enrich samples for virus particles prior to sequencing may simultaneously enrich ribosomes from bacterial pathogens, which are similar in size to small viruses. METHODS: We studied the performance of a viromic library preparation technique (VIDISCA) combined with low-depth IonTorrent sequencing (median ~ 25,000 reads per sample) for detection of ribosomal RNA from common pathogens, analyzing 89 cerebrospinal fluid samples from patients with culture proven bacterial meningitis. RESULTS: Sensitivity and specificity to Streptococcus pneumoniae (n = 24) before and after optimizing threshold parameters were 79% and 52%, then 88% and 90%. Corresponding values for Neisseria meningitidis (n = 22) were 73% and 93%, then 67% and 100%, Listeria monocytogenes (n = 24) 21% and 100%, then 27% and 100%, and Haemophilus influenzae (n = 18) 56% and 100%, then 71% and 100%. A higher total sequencing depth, no antibiotic treatment prior to lumbar puncture, increased disease severity, and higher c-reactive protein levels were associated with pathogen detection. CONCLUSION: We provide proof of principle that a viromic approach can be used to correctly identify bacterial ribosomal RNA in patients with bacterial meningitis. Further work should focus on increasing assay sensitivity, especially for problematic species (e.g. L. monocytogenes), as well as profiling additional pathogens. The technique is most suited to research settings and examination of idiopathic cases, rather than an acute clinical setting.

Cerebrospinal fluid and plasma procalcitonin for the diagnosis of neonatal bacterial meningitis.

AIM: There is a paucity of data on cerebrospinal fluid (CSF) procalcitonin (PCT) to diagnose neonatal meningitis. We evaluated CSF PCT to diagnose bacterial meningitis among neonates with suspected sepsis. METHODS: Neonates undergoing lumbar puncture (LP) as part of sepsis workup were included. INDEX TESTS: CSF PCT, plasma PCT, CSF:plasma PCT ratio and CSF cytochemistry. Reference Standards: 'Definite meningitis' defined by positive CSF culture and/or gram stain and/or broad-based primer 16S rDNA polymerase chain reaction. 'Definite or probable' meningitis is defined as definite meningitis or abnormal cytochemistry. RESULTS: Of 216 eligible neonates, 18 had 'definite meningitis' and 37 'definite or probable meningitis'. Median (Q1 , Q3 ) CSF PCT level was significantly higher in 'definite meningitis' compared to 'no definite meningitis' (0.429 (0.123, 1.300) vs. 0.181 (0.119, 0.286) ng/mL respectively, P = 0.028). Likewise, it was significantly higher in 'definite or probable meningitis' compared to no meningitis (0.245 (0.136, 0.675) vs. 0.170 (0.116, 0.28), P = 0.01). The area under the receiver operator characteristics curve of CSF PCT level for definite meningitis was 0.656 and for 'definite or probable meningitis' 0.635. Paired comparisons of area under the receiver operator characteristics curve of CSF PCT with the other index tests showed no significant differences. Based on a priori cut-off of 0.2 ng/mL, CSF PCT level had a sensitivity (95% confidence interval) of 67% (50, 80), specificity 58% (54, 61), LR+ 1.6 (1.1, 2.0) and LR- 0.6 (0.3, 0.9). CONCLUSIONS: Higher values of CSF PCT are associated with neonatal bacterial meningitis. However, the diagnostic performance of CSF PCT is modest and not significantly different from standard tests.

Bacterial meningitis presenting with a normal cerebrospinal fluid leukocyte count.

OBJECTIVES: We describe clinical characteristics and outcome of adults with bacterial meningitis presenting with a normal CSF leukocyte count. METHODS: We studied community-acquired bacterial meningitis with a normal CSF leukocyte count (≤ 5 per mm3) in adults from a prospective nationwide cohort study. RESULTS: From 2006 through 2020, 39 of 2,357 (2%) episodes presented with a normal CSF leukocyte count. Immunocompromising conditions were present in 19 of 39 patients (49%), compared to 690 of 2303 (30%) in patients with elevated leukocytes (P = 0.02). The triad of fever, neck stiffness, and altered consciousness was present in 6 of 34 patients (18%). CSF protein was abnormal in 25 of 37 patients (68%). We identified 3 clinical subgroups: those with severe pneumococcal meningitis (20 patients [51%]), with mainly sepsis (8 [21%]), and a miscellaneous group (11 [28%]). All patients with severe pneumococcal meningitis presented with high CSF protein levels and 18 of 19 (95%) had bacteria in the CSF Gram stain. Outcome was unfavorable in 23 of 39 (59%) patients and 12 (31%) died. CONCLUSION: Patients with bacterial meningitis may present with normal CSF leukocyte counts. In these patients, CSF protein levels and Gram staining are important diagnostic parameters.

Optimization of cerebrospinal fluid microbial DNA metagenomic sequencing diagnostics.

Infection in the central nervous system is a severe condition associated with high morbidity and mortality. Despite ample testing, the majority of encephalitis and meningitis cases remain undiagnosed. Metagenomic sequencing of cerebrospinal fluid has emerged as an unbiased approach to identify rare microbes and novel pathogens. However, several major hurdles remain, including establishment of individual limits of detection, removal of false positives and implementation of universal controls. Twenty-one cerebrospinal fluid samples, in which a known pathogen had been positively identified by available clinical techniques, were subjected to metagenomic DNA sequencing. Fourteen samples contained minute levels of Epstein-Barr virus. The detection threshold for each sample was calculated by using the total leukocyte content in the sample and environmental contaminants found in the bioinformatic classifiers. Virus sequences were detected in all ten samples, in which more than one read was expected according to the calculations. Conversely, no viral reads were detected in seven out of eight samples, in which less than one read was expected according to the calculations. False positive pathogens of computational or environmental origin were readily identified, by using a commonly available cell control. For bacteria, additional filters including a comparison between classifiers removed the remaining false positives and alleviated pathogen identification. Here we show a generalizable method for identification of pathogen species using DNA metagenomic sequencing. The choice of bioinformatic method mainly affected the efficiency of pathogen identification, but not the sensitivity of detection. Identification of pathogens requires multiple filtering steps including read distribution, sequence diversity and complementary verification of pathogen reads.

Trends of Antimicrobial Susceptibility in Clinically Significant Coagulase-Negative Staphylococci Isolated from Cerebrospinal Fluid Cultures in Neurosurgical Adults: a Nine-Year Analysis.

Coagulase-negative staphylococci (CoNS) are the main pathogens in health care-associated ventriculitis and meningitis (HCAVM). This study aimed to assess antimicrobial susceptibility. Moreover, the treatment and clinical outcome were described. All neurosurgical adults admitted to one of the largest neurosurgical centers in China with clinically significant CoNS isolated from cerebrospinal fluid cultures in 2012 to 2020 were recruited. One episode was defined as one patient with one bacterial strain. Interpretive categories were applied according to the MICs. The clinical outcomes were dichotomized into poor (Glasgow Outcome Scale 1 to 3) and acceptable (Glasgow Outcome Scale 4 to 5). In total, 534 episodes involving 519 patients and 16 bacteria were analyzed. Over the 9 years, eight antimicrobial agents were used in antimicrobial susceptibility tests, including six in over 80% of CoNS. The range of resistance rates was 0.8% to 84.6%. The vancomycin resistance rate was the lowest, whereas the penicillin resistance rate was the highest. The linezolid (a vancomycin replacement) resistance rate was 3.1%. The rate of oxacillin resistance, representing methicillin-resistant staphylococci, was 70.2%. There were no significant trends of antimicrobial susceptibility over the 9 years for any agents analyzed. However, there were some apparent changes. Notably, vancomycin-resistant CoNS appeared in recent years, while linezolid-resistant CoNS appeared early and disappeared in recent years. Vancomycin (or norvancomycin), the most common treatment agent, was used in 528 (98.9%) episodes. Finally, 527 (98.7%) episodes had acceptable outcomes. It will be safe to use vancomycin to treat CoNS-related HCAVM in the immediate future, although continuous monitoring will be needed. IMPORTANCE Coagulase-negative staphylococci are the main pathogens in health care-associated ventriculitis and meningitis. There are three conclusions from the results of this study. First, according to antimicrobial susceptibility, the rates of resistance to primary antimicrobial agents are high and those to high-level agents, including vancomycin, are low. Second, the trends of resistance rates are acceptable, especially for high-level agents, although long-term and continuous monitoring is necessary. Finally, the clinical outcomes of neurosurgical adults with coagulase-negative staphylococci-related health care-associated ventriculitis and meningitis are acceptable after treatment with vancomycin. Therefore, according to the antimicrobial susceptibility and clinical practice, vancomycin will be safe to treat coagulase-negative staphylococci-related health care-associated ventriculitis and meningitis.

Comparative diagnostic utility of metagenomic next-generation sequencing, GeneXpert, modified Ziehl-Neelsen staining, and culture using cerebrospinal fluid for tuberculous meningitis: A multi-center, retrospective study in China.

BACKGROUND: Early diagnosis of tuberculosis meningitis (TBM) remains a great challenge during clinical practice. The diagnostic efficacies of cerebrospinal fluid (CSF)-based mycobacterial growth indicator tube (MGIT) culture, modified Ziehl-Neelsen (ZN) staining, Xpert MTB/RIF, and metagenomic next-generation sequencing (mNGS) for TBM remained elusive. METHODS: A total of 216 adult patients with suspicious TBM were retrospectively enrolled in this multi-cohort study. The diagnostic performances for MGIT, modified ZN staining, Xpert MTB/RIF, and mNGS using CSF samples were evaluated. RESULTS: Uniform clinical case definition classified 88 (40.7%) out of 216 patients as the definite TBM, 5 (2.3%) patients as probable TBM cases, and 24 (11.1%) patients as possible TBM cases. The sensitivities of MGIT, modified ZN staining, Xpert MTB/RIF, and mNGS for TBM diagnosis against consensus uniform case definition for definite TBM were 25.0%, 76.1%, 73.9%, and 84.1%, respectively. Negative predictive values (NPVs) were 66.0%, 85.9%, 84.8%, and 90.1%, respectively. The sensitivities of MGIT, modified ZN staining, Xpert MTB/RIF, and mNGS for TBM diagnosis against consensus uniform case definition for definite, probable, and possible TBM were 18.8%, 57.3%, 55.5%, and 63.2%, respectively. Negative predictive values (NPVs) were 51.0%, 66.4%, 65.6%, and 69.7%, respectively. mNGS combined with modified ZN stain and Xpert could cover TBM cases against a composite microbiological reference standard, yielding 100% specificity and 100% NPV. CONCLUSION: Metagenomic next-generation sequencing detected TBM with higher sensitivity than Xpert, ZN staining and MGIT culture, but mNGS cannot be used as a rule-out test. mNGS combined with Xpert or modified ZN staining could enhance the sensitivity of diagnostic tests for TBM.

An uncommon case of brucellosis with ocular presentation / Um caso incomum de brucelose com apresentação ocular

ABSTRACT We report an unusual case of brucellosis presented with headache, diminished vision, papillitis and multiple peripapillary hemorrhages accompanied by subretinal fluid extending up to macula. Diagnosis of brucellosis was made based on positive polymerase chain reaction of cerebrospinal fluid sample for Brucella species DNA, accompanied by a raised titer of anti-brucella antibodies. Patient showed remarkable improvement on triple drug therapy in form of doxycycline, rifampicin and ceftriaxone.

RESUMO Relatamos um caso incomum de brucelose apresentada com cefaleia, visão diminuída, papilite e múltiplas hemorragias peripapilares acompanhadas por fluido sub-retinal, estendendo-se até a mácula. O diagnóstico de brucelose foi feito com base na reação em cadeia da polimerase positiva de amostra de líquido cefalorraquidiano para DNA de espécies de Brucella, acompanhada por um título elevado de anticorpos antibrucela. O paciente apresentou melhora notável com a terapia tripla com drogas na forma de doxiciclina, rifampicina e ceftriaxona.

Capsule-dependent impact of MAPK signalling on host cell invasion and immune response during infection of the choroid plexus epithelium by Neisseria meningitidis.

BACKGROUND: The Gram-negative bacterium Neisseria meningitidis (Nm) can cause meningitis in humans, but the host signalling pathways manipulated by Nm during central nervous system (CNS) entry are not completely understood. METHODS: We investigate the role of the mitogen-activated protein kinases (MAPK) Erk1/2 and p38 in an in vitro model of the blood-cerebrospinal fluid barrier (BCSFB) based on human epithelial choroid plexus (CP) papilloma (HIBCPP) cells during infection with Nm serogroup B (NmB) and serogroup C (NmC) strains. A transcriptome analysis of HIBCPP cells following infection with Nm by massive analysis of cDNA ends (MACE) was done to further characterize the cellular response to infection of the barrier. RESULTS: Interestingly, whereas NmB and NmC wild type strains required active Erk1/2 and p38 pathways for infection, invasion by capsule-deficient mutants was independent of Erk1/2 and, in case of the NmB strain, of p38 activity. The transcriptome analysis of HIBCPP cells following infection with Nm demonstrated specific regulation of genes involved in the immune response dependent on Erk1/2 signalling. Gene ontology (GO) analysis confirmed loss of MAPK signalling after Erk1/2 inhibition and revealed an additional reduction of cellular responses including NFκB and JAK-STAT signalling. Interestingly, GO terms related to TNF signalling and production of IL6 were lost specifically following Erk1/2 inhibition during infection with wild type Nm, which correlated with the reduced infection rates by the wild type in absence of Erk1/2 signalling. CONCLUSION: Our data point towards a role of MAPK signalling during infection of the CP epithelium by Nm, which is strongly influenced by capsule expression, and affects infection rates as well as the host cell response.

Cerebrospinal Fluid from Healthy Pregnant Women Does Not Harbor a Detectable Microbial Community.

Cerebrospinal fluid (CSF) circulating in the human central nervous system has long been considered aseptic in healthy individuals, because normally, the blood-brain barrier can protect against microbial invasions. However, this dogma has been called into question by several reports that microbes were identified in human brains, raising the question of whether there is a microbial community in the CSF of healthy individuals without neurological diseases. Here, we collected CSF samples and other samples, including one-to-one matched oral and skin swab samples (positive controls), from 23 pregnant women aged between 23 and 40 years. Normal saline samples (negative controls), sterile swabs, and extraction buffer samples (contamination controls) were also collected. Twelve of the CSF specimens were also used to evaluate the physiological activities of detected microbes. Metagenomic and metatranscriptomic sequencing was performed in these 116 specimens. A total of 620 nonredundant microbes were detected, which were dominated by bacteria (74.6%) and viruses (24.2%), while in CSF samples, metagenomic sequencing found only 26 nonredundant microbes, including one eukaryote, four bacteria, and 21 viruses (mostly bacteriophages). The beta diversity of microbes compared between CSF metagenomic samples and other types of samples (except negative controls) was significantly different from that of the CSF self-comparison. In addition, there was no active or viable microbe in the matched metagenomic and metatranscriptomic sequencing of CSF specimens after subtracting those also found in normal saline, DNA extraction buffer, and skin swab specimens. In conclusion, our results showed no strong evidence of a colonized microbial community present in the CSF of healthy individuals. IMPORTANCE The microbiome is prevalent throughout human bodies, with profound health implications. However, it remains unclear whether it is present and active in human CSF, which has been long considered aseptic due to the blood-brain barrier. Here, we applied unbiased metagenomic and metatranscriptomic sequencing to detect the presence of a microbiome in CSF collected from 23 pregnant women with matched controls. Analysis of 116 specimens found no strong evidence to support the presence of a colonized microbiome in CSF. Our findings will strengthen our understanding of the internal environment of the CSF in healthy people, which has strong implications for human health, especially for neurological infections and disorders, and will help further disease diagnostics, prevention, and therapeutics in clinical settings.

Diagnostic Model for Discrimination Between Tuberculous Meningitis and Bacterial Meningitis.

Background: The differential diagnosis between tuberculous meningitis (TBM) and bacterial meningitis (BM) remains challenging in clinical practice. This study aimed to establish a diagnostic model that could accurately distinguish TBM from BM. Methods: Patients with TBM or BM were recruited between January 2017 and January 2021 at Tongji Hospital (Qiaokou cohort) and Sino-French New City Hospital (Caidian cohort). The detection for indicators involved in cerebrospinal fluid (CSF) and T-SPOT assay were performed simultaneously. Multivariate logistic regression was used to create a diagnostic model. Results: A total of 174 patients (76 TBM and 98 BM) and another 105 cases (39 TBM and 66 BM) were enrolled from Qiaokou cohort and Caidian cohort, respectively. Significantly higher level of CSF lymphocyte proportion while significantly lower levels of CSF chlorine, nucleated cell count, and neutrophil proportion were observed in TBM group when comparing with those in BM group. However, receiver operating characteristic (ROC) curve analysis showed that the areas under the ROC curve (AUCs) produced by these indicators were all under 0.8. Meanwhile, tuberculosis-specific antigen/phytohemagglutinin (TBAg/PHA) ratio yielded an AUC of 0.889 (95% CI, 0.840-0.938) in distinguishing TBM from BM, with a sensitivity of 68.42% (95% CI, 57.30%-77.77%) and a specificity of 92.86% (95% CI, 85.98%-96.50%) when a cutoff value of 0.163 was used. Consequently, we successfully established a diagnostic model based on the combination of TBAg/PHA ratio, CSF chlorine, CSF nucleated cell count, and CSF lymphocyte proportion for discrimination between TBM and BM. The established model showed good performance in differentiating TBM from BM (AUC: 0.949; 95% CI, 0.921-0.978), with 81.58% (95% CI, 71.42%-88.70%) sensitivity and 91.84% (95% CI, 84.71%-95.81%) specificity. The performance of the diagnostic model obtained in Qiaokou cohort was further validated in Caidian cohort. The diagnostic model in Caidian cohort produced an AUC of 0.923 (95% CI, 0.867-0.980) with 79.49% (95% CI, 64.47%-89.22%) sensitivity and 90.91% (95% CI, 81.55%-95.77%) specificity. Conclusions: The diagnostic model established based on the combination of four indicators had excellent utility in the discrimination between TBM and BM.

A clinical, aetiological, and public health perspective on central nervous system infections in Bolivia, 2017-2018.

Central nervous system (CNS) infections are important causes of morbidity and mortality worldwide. In Bolivia, aetiologies, case fatality, and determinants of outcome are poorly characterised. We attempted to investigate such parameters to guide diagnosis, treatment, prevention, and health policy. From Nov-2017 to Oct-2018, we prospectively enrolled 257 inpatients (20.2% HIV-positive patients) of all ages from healthcare centers of Cochabamba and Santa Cruz, Bolivia with a suspected CNS infection and a lumbar puncture performed. Biological diagnosis included classical microbiology, molecular, serological and immunohistochemical tests. An infectious aetiology was confirmed in 128/257 (49.8%) inpatients, including, notably among confirmed single and co-infections, Cryptococcus spp. (41.7%) and Mycobacterium tuberculosis (27.8%) in HIV-positive patients, and Mycobacterium tuberculosis (26.1%) and Streptococcus pneumoniae (18.5%) in HIV-negative patients. The total mortality rate was high (94/223, 42.1%), including six rabies cases. In multivariate logistic regression analysis, mortality was associated with thrombocytopenia (Odds ratio (OR) 5.40, 95%-CI 2.40-11.83) and hydrocephalus (OR 4.07, 95%-CI 1.35-12.23). The proportion of untreated HIV patients, late presentations of neurotuberculosis, the rate of pneumococcal cases, and rabies patients who did not benefit from a post-exposure prophylaxis, suggest that decreasing the burden of CNS infections requires reinforcing health policy regarding tuberculosis, rabies, S. pneumoniae vaccination, and HIV-infections.

Endogenous α7 nAChR Agonist SLURP1 Facilitates Escherichia coli K1 Crossing the Blood-Brain Barrier.

Alpha 7 nicotinic acetylcholine receptor (α7 nAChR) is critical for the pathogenesis of Escherichia coli (E. coli) K1 meningitis, a severe central nervous system infection of the neonates. However, little is known about how E. coli K1 manipulates α7 nAChR signaling. Here, through employing immortalized cell lines, animal models, and human transcriptional analysis, we showed that E. coli K1 infection triggers releasing of secreted Ly6/Plaur domain containing 1 (SLURP1), an endogenous α7 nAChR ligand. Exogenous supplement of SLURP1, combined with SLURP1 knockdown or overexpression cell lines, showed that SLURP1 is required for E. coli K1 invasion and neutrophils migrating across the blood-brain barrier (BBB). Furthermore, we found that SLURP1 is required for E. coli K1-induced α7 nAChR activation. Finally, the promoting effects of SLURP1 on the pathogenesis of E. coli K1 meningitis was significantly abolished in the α7 nAChR knockout mice. These results reveal that E. coli K1 exploits SLURP1 to activate α7 nAChR and facilitate its pathogenesis, and blocking SLURP1-α7 nAChR interaction might represent a novel therapeutic strategy for E. coli K1 meningitis.

Cerebrospinal fluid culture-positive bacterial meningitis increases the risk for neurologic damage among neonates.

OBJECTIVE: This study aimed to compare the clinical features and outcomes of neonatal bacterial meningitis (NBM) between patients with positive and negative cerebrospinal fluid (CSF) cultures and determine the risk factors for CSF culture-positive NBM. METHODS: We retrospectively reviewed the medical records of all patients with NBM. Perinatal clinical data, laboratory results, and cranial radiographs were obtained. RESULTS: Among the 186 neonates who met the inclusion criteria. The risk factors for positive CSF culture results were analysed using multiple logistic regression. The multivariable logistic regression analysis showed that the possible risk factors of NBM with positive CSF culture in this study were: Length of fever [OR = 1.126; 95% CI (0.999-1.268)], Neurologic symptoms [OR = 3.043; 95% CI (1.164-7.959)], Cerebrospinal fluid protein [OR = 1.001; 95% CI (1.000-1.001)]. Cases of NBM with a longer duration of fever, more neurologic symptoms, and higher levels of CSF protein were more likely to demonstrate positive results on CSF culture. CONCLUSION: Cases of NBM with CSF culture-positive results were more likely to have severe clinical manifestations and develop more serious neurologic damage. Patients with NBM who have longer durations of fever, more neurologic symptoms, and higher levels of CSF protein were more likely to have CSF culture-positive results, who should be followed up more closely.Key MessageBacterial meningitis is clinically defined as a serious inflammation of meningitis, usually caused by a variety of bacterial infections that may leave sequelae and long-term complications and high mortality rates. Early diagnosis is often difficult, particularly when the patient has been treated with antimicrobials.