Simultaneous detection of Neisseria meningitidis, Streptococcus pneumoniae and Haemophilus influenzae by quantitative PCR from CSF samples with negative culture in Morocco

2021 ◽  
Author(s):  
Youssef Ikken ◽  
Amina Benaouda ◽  
Latifa Ibn Yaich ◽  
Farida Hilali ◽  
Yassine Sekhsokh ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Bacterial Meningitis ◽  
Haemophilus Influenzae ◽  
Molecular Epidemiology ◽  
Neisseria Meningitidis ◽  
Quantitative Pcr ◽  
Sensitivity And Specificity ◽  
Simultaneous Detection ◽  
Rt Pcr ◽  
Negative Culture

AbstractBacteriological cultures from cerebrospinal fluids (CSF) have less sensitivity and specificity compared to quantitative PCR (RT-PCR), and multiple facts still conduct to the increase of negative culture. The aims of this study are to determine the molecular epidemiology and the simultaneous detection of bacterial meningitis in Morocco by using RT-PCR and compared this molecular approach with culture method to improve the etiological diagnosis of meningitis. The CSFs were collected over one-year period in 2018 in different hospitals covering all regions of the Kingdom of Morocco, from patients with suspected meningitis. The results showed the confirmation rate per culture recorded a rate of 33% and the RT-PCR of 70%. Molecular epidemiology is predominant of Neisseria meningitidis followed by Streptococcus pneumoniae and a dramatic reduction in meningitis due to Haemophilus influenzae following the introduction of conjugate vaccine in 2007. Also, the epidemiological profile shows a sex ratio M/F of 1.4 and a median age of 2 years. The national distribution showed a predominant of meningococcal disease followed by pneumococcal disease, especially a dominance of N. meningitidis over S. pneumoniae in two regions and a slight predominance of S. pneumoniae in the other two regions over N. meningitidis. Our research shows that culture in our country has less sensitivity and specificity than RT-PCR in diagnosis of bacterial meningitis and that molecular biology technique at bacteriology laboratories is desirable for diagnosis, early management of meningitis cases and in the context of the surveillance of meningitis in Morocco in parallel with culture.

scholarly journals A Narrative Review of the Molecular Epidemiology and Laboratory Surveillance of Vaccine Preventable Bacterial Meningitis Agents: Streptococcus pneumoniae, Neisseria meningitidis, Haemophilus influenzae and Streptococcus agalactiae

2021 ◽  
Vol 9 (2) ◽  
pp. 449
Author(s):  
Raymond S. W. Tsang
Keyword(s):  
Streptococcus Pneumoniae ◽  
Bacterial Meningitis ◽  
Haemophilus Influenzae ◽  
Molecular Epidemiology ◽  
Neisseria Meningitidis ◽  
Narrative Review ◽  
Vulnerable Groups ◽  
Human Pathogens ◽  
Public Health Importance ◽  
Laboratory Surveillance

This narrative review describes the public health importance of four most common bacterial meningitis agents, Streptococcus pneumoniae, Neisseria meningitidis, Haemophilus influenzae, and S. agalactiae (group B Streptococcus). Three of them are strict human pathogens that normally colonize the nasopharynx and may invade the blood stream to cause systemic infections and meningitis. S. agalactiae colonizes the genito-gastrointestinal tract and is an important meningitis agent in newborns, but also causes invasive infections in infants or adults. These four bacteria have polysaccharide capsules that protect them against the host complement defense. Currently licensed conjugate vaccines (against S. pneumoniae, H. influenza, and N. meningitidis only but not S. agalactiae) can induce protective serum antibodies in infants as young as two months old offering protection to the most vulnerable groups, and the ability to eliminate carriage of homologous serotype strains in vaccinated subjects lending further protection to those not vaccinated through herd immunity. However, the serotype-specific nature of these vaccines have driven the bacteria to adapt by mechanisms that affect the capsule antigens through either capsule switching or capsule replacement in addition to the possibility of unmasking of strains or serotypes not covered by the vaccines. The post-vaccine molecular epidemiology of vaccine-preventable bacterial meningitis is discussed based on findings obtained with newer genomic laboratory surveillance methods.

scholarly journals Detection of Neisseria meningitidis, Streptococcus pneumoniae and Haemophilus influenzae in Cerebrospinal Fluid by Multiplex PCR for Diagnosis of Acute Bacterial Meningitis in Paediatric Population

2017 ◽  
Vol 11 (2) ◽  
pp. 9-16
Author(s):  
Nilufar Yeasmin Nili
Keyword(s):  
Streptococcus Pneumoniae ◽  
16S Rrna ◽  
Bacterial Meningitis ◽  
Haemophilus Influenzae ◽  
Neisseria Meningitidis ◽  
Multiplex Pcr ◽  
Bacterial Culture ◽  
Acute Bacterial Meningitis ◽  
Pcr Assay ◽  
Etiologic Diagnosis

The present study was done to evaluate a multiplex PCR based method for simultaneous detection of Neisseria meningitidis, Streptococcus pneumoniae and Haemophilus influenzae in CSF. A cross sectional study was carried out with 140 children (2 months to 12 years of age) with clinical suspicion of acute meningitis during July 2010 to June 2011. Three species-specific primers were used along with universal primers of bacterial gene 16S rRNA, in a two-stage PCR assay for diagnosis of acute bacterial meningitis.Among 140 patients, 42 (30%) cases were diagnosed as bacterial meningitis and other 98 (70%) as viral meningitis by clinical and cytobiochemical criteria. Out of 42 bacterial meningitis cases, 9 (21.43%) were positive by Gram stain.These 9 cases were also positive by bacterial culture and PCR. Again, 15 (35.71%) were positive by bacterial culture which were also PCR positive. In 27 cases (out of 42), the etiologic diagnosis was not possible using routine bacteriological methods; in 11 of these patients, the etiologic agents were identified by PCR. In addition, PCR recognized 5 more cases whose etiologic diagnosis was not possible, as they were identified by universal primer of 16S rRNA. Hence, among 31 (73.81%) PCR positive cases, 12 (38.71%) were S. pneumoniae, 10 (32.26%) were H. influenzae, 4 (12.9%) were N. meningitidis and 5 (16.13%) were other bacteria.Among the antibiotic users, bacterial meningitis case detection by PCR was higher (65.52%) than that of culture (10.34%) and Gram staining (6.90%). The overall sensitivity and specificity of PCR assay was 100% and 66% respectively when bacterial culture was considered as gold standard. PCR can be used as a valuable supplementary diagnostic technique in routine clinical practice for diagnosis of acute bacterial meningitis in hospital setting. Bangladesh J Med Microbiol 2017; 11 (2): 9-16

scholarly journals Comparison of PCR-based methods for the simultaneous detection of Neisseria meningitidis, Haemophilus influenzae, and Streptococcus pneumoniae in clinical samples

2016 ◽  
Vol 20 (4) ◽  
pp. 335-341 ◽  
Author(s):  
Ivano de Filippis ◽  
Claudia Ferreira de Andrade ◽  
Nathalia Caldeira ◽  
Aline Carvalho de Azevedo ◽  
Antonio Eugenio de Almeida
Keyword(s):  
Streptococcus Pneumoniae ◽  
Haemophilus Influenzae ◽  
Neisseria Meningitidis ◽  
Simultaneous Detection ◽  
Clinical Samples

scholarly journals Molecular detection and characterization of bacteria from CSF samples of patients with suspected cerebrospinal meningitis in parts of northern Nigeria using metagenomic DNA extracts

2021 ◽  
Vol 22 (3) ◽  
pp. 365-376
Author(s):  
I.C. Peletiri ◽  
E.I. Ikeh ◽  
G.M. Ayanbimpe ◽  
E. Nna
Keyword(s):  
Streptococcus Pneumoniae ◽  
Haemophilus Influenzae ◽  
Neisseria Meningitidis ◽  
Real Time Pcr ◽  
Molecular Detection ◽  
Northern Nigeria ◽  
Rt Pcr ◽  
Metagenomic Dna ◽  
Pcr Multiplex

Background: The most commonly used approaches for detection and characterization of bacterial pathogens of meningitis in developing countries include culture, Gram stain, and latex agglutination. The positivity rate of culture is relatively low due to suboptimal storage and transportation conditions, culture practice, and/or antibiotic treatment administered before specimens are collected. Specimens that yield no growth in culture can still be analyzed using molecular methods, and metagenomic DNA (mDNA) extracted directly from clinical samples (CSF) can be used. We aimed to detect and characterize three major bacterial causes of cerebrospinal meningitis (CSM); Neisseria meningitidis, Haemophilus influenzae, and Streptococcus pneumoniae using mDNA extracted directly from CSF samples. Methodology: Metagenomic DNA templates were prepared directly from CSF specimens collected from 210 patients with suspected CSM. A multiplex Real Time PCR (mRT-PCR) using the ABI StepOne Plus Machine and Taqman Probe chemistry was used in the molecular detection, while serogroup/serotype-specific singleplex RT-PCR was used to characterize all positives samples. Results: Eighty-eight (41.9%) of the 210 samples were positive with the mRT-PCR assay for one or a combination of two of the three bacteria. Of these, 59 (67.1%) were N. meningitidis, 2 (2.3%) were H. influenzae, 3 (3.4%) were S. pneumoniae, 15 (17 %) had co-infections of N. meningitidis with H. influenzae, and 9 (10.2%) had co-infections of H. influenzae and S. pneumoniae. The serogroups of N. meningitidis encountered were A (13.5%), B (23%), C (8.1%), W135 (8.1%), X (5.4%), Y (32.4%), and non-groupable (9.5%). The serotypes of H. influenzae were Hia (3.8%), Hib (57.7%), Hic (3.85%), Hie (11.5%) and Hif (23.1%). The serotypes of S. pneumoniae were Wxy1 (8.3%), Wxy4 (33.3%), Wxy5 (50.0%), and Wxy9 (8.3%). Conclusion: Multiplex RT-PCR is a fast and accurate method for detecting and characterizing serogroups/serotypes of major bacteria implicated in CSM. Isolating DNA directly from CSF improves turnaround time, which will speed up patient care and management. Keywords: Cerebrospinal meningitis, metagenomic DNA, multiplex Real Time PCR, Northern Nigeria   French title: Détection moléculaire et caractérisation de bactéries à partir d'échantillons de LCR de patients suspectés de méningite cérébrospinale dans certaines parties du nord du Nigéria à l'aide d'extraits d'ADN métagénomique   Contexte: Les approches les plus couramment utilisées pour la détection et la caractérisation des agents pathogènes bactériens de la méningite dans les pays en développement comprennent la culture, la coloration de Gram et l'agglutination au latex. Le taux de positivité de la culture est relativement faible en raison des conditions de stockage et de transport sous-optimales, des pratiques de culture et/ou du traitement antibiotique administré avant le prélèvement des échantillons. Les échantillons qui ne donnent pas de croissance en culture peuvent toujours être analysés à l'aide de méthodes moléculaires, et l'ADN métagénomique (ADNm) extrait directement d'échantillons cliniques (LCR) peut être utilisé. Nous visions à détecter et à caractériser trois causes bactériennes majeures de la méningite cérébrospinale (CSM); Neisseria meningitidis, Haemophilus influenzae et Streptococcus pneumoniae à l'aide d'ADNm extrait directement d'échantillons de LCR. Méthodologie: Des matrices d'ADN métagénomique ont été préparées directement à partir d'échantillons de LCR prélevés sur 210 patients suspects de CSM. Une PCR multiplex en temps réel (mRT-PCR) utilisant la chimie de la machine ABI StepOne Plus et de la sonde Taqman a été utilisée pour la détection moléculaire, tandis que la RT-PCR monoplex spécifique au sérogroupe/sérotype a été utilisée pour caractériser tous les échantillons positifs. Résultats: Quatre-vingt-huit (41,9%) des 210 échantillons étaient positifs avec le test mRT-PCR pour une ou une combinaison de deux des trois bactéries. Parmi ceux-ci, 59 (67,1%) étaient N. meningitidis, 2 (2,3%) étaient H. influenzae, 3 (3,4%) étaient S. pneumoniae, 15 (17%) avaient des co-infections de N. meningitidis avec H. influenzae et 9 (10,2%) avaient des co-infections à H. influenzae et S. pneumoniae. Les sérogroupes de N. meningitidis rencontrés étaient A (13,5%), B (23%), C (8,1%), W135 (8,1%), X (5,4%), Y (32,4%) et non groupables (9,5%). Les sérotypes de H. influenzae étaient Hia (3,8%), Hib (57,7%), Hic (3,85%), Hie (11,5%) et Hif (23,1%). Les sérotypes de S. pneumoniae étaient Wxy1 (8,3%), Wxy4 (33,3%), Wxy5 (50,0%) et Wxy9 (8,3%). Conclusion: La RT-PCR multiplex est une méthode rapide et précise de détection et de caractérisation des sérogroupes/sérotypes des principales bactéries impliquées dans le CSM. Isoler l'ADN directement du LCR améliore le temps de traitement, ce qui accélérera les soins et la gestion des patients. Mots clés: méningite cérébro-spinale, ADN métagénomique, PCR multiplex en temps réel, nord du Nigéria

scholarly journals Multicenter Hospital-Based Prospective Surveillance Study of Bacterial Agents Causing Meningitis and Seroprevalence of Different Serogroups of Neisseria meningitidis, Haemophilus influenzae Type b, and Streptococcus pneumoniae during 2015 to 2018 in Turkey

mSphere ◽  
2020 ◽  
Vol 5 (2) ◽  
Author(s):  
Mehmet Ceyhan ◽  
Yasemin Ozsurekci ◽  
Sevgen Tanır Basaranoglu ◽  
Nezahat Gurler ◽  
Enes Sali ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Bacterial Meningitis ◽  
Haemophilus Influenzae ◽  
Neisseria Meningitidis ◽  
Surveillance Study ◽  
Haemophilus Influenzae Type ◽  
Acute Bacterial Meningitis ◽  
Close Monitoring ◽  
Type B ◽  
Content Type

ABSTRACT The etiology of bacterial meningitis in Turkey changed after the implementation of conjugated vaccines against Streptococcus pneumoniae and Haemophilus influenzae type b (Hib) in the Turkish National Immunization Program (NIP). Administration of Hib vaccine and PCV-7 (7-valent pneumococcal conjugate vaccine) was implemented in NIP in 2006 and 2009, respectively. In 2011, PCV-7 was replaced with PCV-13. Meningococcal vaccines have not yet been included in Turkish NIP. This prospective study comprised 27 hospitals located in seven regions of Turkey and represented 45% of the population. Children aged between 1 month and 18 years who were hospitalized with suspected meningitis were included. Cerebrospinal fluid (CSF) samples were collected, and bacterial identification was made according to the multiplex PCR assay results. During the study period, 994 children were hospitalized for suspected meningitis, and Hib (n = 3, 2.4%), S. pneumoniae (n = 33, 26.4%), and Neisseria meningitidis (n = 89, 71%) were detected in 125 samples. The most common meningococcal serogroup was MenB. Serogroup W comprised 13.9% (n = 5) and 7.5% (n = 4) of the meningococci in 2015 to 2016 and 2017 to 2018, respectively. Serogroup C was not detected. There were four deaths in the study; one was a pneumococcus case, and the others were serogroup B meningococcus cases. The epidemiology of meningococcal diseases has varied over time in Turkey. Differing from the previous surveillance periods, MenB was the most common serogroup in the 2015-to-2018 period. Meningococcal epidemiology is so dynamic that, for vaccination policies, close monitoring is crucial. IMPORTANCE Acute bacterial meningitis (ABM) is one of the most common life-threatening infections in children. The incidence and prevalence of ABM vary both geographically and temporally; therefore, surveillance systems are necessary to determine the accurate burden of ABM. The Turkish Meningitis Surveillance Group has been performing a hospital-based meningitis surveillance study since 2005 across several regions in Turkey. Meningococcus was the major ABM-causing agent during the 2015-to-2018 period, during which MenB was the dominant serogroup.

scholarly journals Identification of Common Bacterial Pathogens Causing Meningitis in Culture-Negative Cerebrospinal Fluid Samples Using Real-Time Polymerase Chain Reaction

2016 ◽  
Vol 2016 ◽  
pp. 1-5 ◽  
Author(s):  
Walaa Shawky Khater ◽  
Safia Hamed Elabd
Keyword(s):  
Streptococcus Pneumoniae ◽  
Bacterial Meningitis ◽  
Haemophilus Influenzae ◽  
Neisseria Meningitidis ◽  
Real Time ◽  
Real Time Pcr ◽  
Latex Agglutination ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Culture Negative

Background. Meningitis is a serious communicable disease with high morbidity and mortality rates. It is an endemic disease in Egypt caused mainly byStreptococcus pneumoniae,Neisseria meningitidis, andHaemophilus influenzae. In some settings, bacterial meningitis is documented depending mainly on positive cerebrospinal fluid (CSF) culture results or CSF positive latex agglutination test, missing the important role of prior antimicrobial intake which can yield negative culture and latex agglutination test results. This study aimed to utilize molecular technology in order to diagnose bacterial meningitis in culture-negative CSF samples.Materials and Methods. Forty culture-negative CSF samples from suspected cases of bacterial meningitis were examined by real-time polymerase chain reaction (real-time PCR) for the presence oflytA,bexA, andctrAgenes specific forStreptococcus pneumoniae,Haemophilus influenzae, andNeisseria meningitidis, respectively.Results. Positive real-time PCR results forStreptococcus pneumoniaewere detected in 36 (90%) of culture-negative CSF samples while no positive results forHaemophilus influenzaeorNeisseria meningitidiswere detected. Four (10%) samples were negative by real-time PCR for all tested organisms.Conclusion. The use of molecular techniques as real-time PCR can provide a valuable addition to the proportion of diagnosed cases of bacterial meningitis especially in settings with high rates of culture-negative results.

scholarly journals Detection of Streptococcus pneumoniae, Neisseria meningitidis and Haemophilus influenzae in Culture Negative Cerebrospinal Fluid Samples from Meningitis Patients Using a Multiplex Polymerase Chain Reaction in Nepal

2021 ◽  
Vol 13 (1) ◽  
pp. 173-180
Author(s):  
Supriya Sharma ◽  
Jyoti Acharya ◽  
Dominique A. Caugant ◽  
Megha Raj Banjara ◽  
Prakash Ghimire ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Cerebrospinal Fluid ◽  
Streptococcus Pneumoniae ◽  
Bacterial Meningitis ◽  
Haemophilus Influenzae ◽  
Neisseria Meningitidis ◽  
Multiplex Polymerase Chain Reaction ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Culture Negative

The rapid identification of bacteria causing meningitis is crucial as delays in the treatment increase mortality rate. Though considered as the gold standard for the laboratory diagnosis of bacterial meningitis, culture might give false negative results in a case of patients under antibiotics prior to lumbar puncture. This study aimed to detect Streptococcus pneumoniae, Neisseria meningitidis and Haemophilus influenzae by a multiplex polymerase chain reaction (PCR) in culture-negative cerebrospinal fluid samples collected from clinically suspected meningitis cases attending different hospitals in Kathmandu, Nepal from January 2017 to December 2019. S. pneumoniae, N. meningitidis and H. influenzae were detected in 8.59% (33/384) of the specimens by PCR and 7.55% (29/384) of the specimens by culture. Correlation between culture and PCR of the same sample was good (Spearman’s rho correlation coefficient = 0.932). However, the difference in positivity between culture and PCR was statistically not significant (p value > 0.05). In four specimens, culture could not detect any of the targeted bacteria whereas PCR could detect presence of H. influenzae. PCR increases the diagnostic yield for bacterial meningitis. PCR may be considered as an adjunctive test for establishing the cause of infection in culture negative clinically suspected meningitis cases.

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