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Background The World Health Organization (WHO) coordinates the Global Invasive Bacterial Vaccine-Preventable Diseases (IB-VPD) Surveillance Network to support vaccine introduction decisions and use. The network was established to strengthen surveillance and laboratory confirmation of meningitis caused by Streptococcus pneumoniae, Haemophilus influenzae, and Neisseria meningitidis. Methods Sentinel hospitals report cases of children Results From 2014 through 2019, >137 000 suspected meningitis cases were reported by 58 participating countries, with 44.6% (n = 61 386) reported from countries in the WHO African Region. More than half (56.6%, n = 77 873) were among children Conclusions This multitier, global surveillance network has supported countries in detecting and serotyping the 3 principal invasive bacterial pathogens that cause pediatric meningitis. Streptococcus pneumoniae was the most common bacterial pathogen detected globally despite the growing number of countries that have nationally introduced PCV. The large proportions of deaths due to S. pneumoniae reflect the high proportion of meningitis cases caused by this pathogen. This global network demonstrated a strong correlation between PCV introduction status and reduction in the proportion of pneumococcal meningitis infections caused by vaccine serotypes. Maintaining case-based, active surveillance with laboratory confirmation for prioritized vaccine-preventable diseases remains a critical component of the global agenda in public health. The World Health Organization (WHO)-coordinated Invasive Bacterial Vaccine-Preventable Disease (IB-VPD) Surveillance Network reported data from 2014 to 2019, contributing to the estimates of the disease burden and serotypes of pediatric meningitis caused by Streptococcus pneumoniae, Haemophilus influenzae and Neisseria meningitidis.

Meningitis confirmation in Burkina Faso uses PCR for detecting Streptococcus pneumoniae, Neisseria meningitidis, or Hemophilus influenzae. We identified 38 cases of meningitis among 590 that were PCR-positive for 3 nonpneumococcal streptococcal pathogens, including 21 cases of Streptococcus suis. Among the country's 13 regions, 10 had S. suis-positive cases.

Despite contributing to the large disease burden in West Africa, little is known about the genomic epidemiology of Streptococcus pneumoniae which cause meningitis among children under 5 years old in the region. We analysed whole-genome sequencing data from 185 S . pneumoniae isolates recovered from suspected paediatric meningitis cases as part of the World Health Organization (WHO) invasive bacterial diseases surveillance from 2010 to 2016. The phylogeny was reconstructed, accessory genome similarity was computed and antimicrobial-resistance patterns were inferred from the genome data and compared to phenotypic resistance from disc diffusion. We studied the changes in the distribution of serotypes pre- and post-pneumococcal conjugate vaccine (PCV) introduction in the Central and Western sub-regions separately. The overall distribution of non-vaccine, PCV7 (4, 6B, 9V, 14, 18C, 19F and 23F) and additional PCV13 serotypes (1, 3, 5, 6A, 19A and 7F) did not change significantly before and after PCV introduction in the Central region (Fisher's test P value 0.27) despite an increase in the proportion of non-vaccine serotypes to 40 % (n=6) in the post-PCV introduction period compared to 21.9 % (n=14). In the Western sub-region, PCV13 serotypes were more dominant among isolates from The Gambia following the introduction of PCV7, 81 % (n=17), compared to the pre-PCV period in neighbouring Senegal, 51 % (n=27). The phylogeny illustrated the diversity of strains associated with paediatric meningitis in West Africa and highlighted the existence of phylogeographical clustering, with isolates from the same sub-region clustering and sharing similar accessory genome content. Antibiotic-resistance genotypes known to confer resistance to penicillin, chloramphenicol, co-trimoxazole and tetracycline were detected across all sub-regions. However, there was no discernible trend linking the presence of resistance genotypes with the vaccine introduction period or whether the strain was a vaccine or non-vaccine serotype. Resistance genotypes appeared to be conserved within selected sub-clades of the phylogenetic tree, suggesting clonal inheritance. Our data underscore the need for continued surveillance on the emergence of non-vaccine serotypes as well as chloramphenicol and penicillin resistance, as these antibiotics are likely still being used for empirical treatment in low-resource settings. This article contains data hosted by Microreact.

This study presents a triplex real-time PCR assay that allows for the direct detection of Streptococcus pneumoniae, Neisseria meningitidis, and Haemophilus influenzae in one reaction without DNA extraction, with similar sensitivity and specificity to singleplex assays. This approach saves time, specimen volume and reagents while achieving a higher testing throughput.

Mycoplasma pneumoniae is an important respiratory pathogen, accounting for up to 25% of community-acquired pneumonia, and is a common cause of hospitalized pneumonia in otherwise healthy adults and children. Mycoplasma pneumoniae isolates can be classified into two main genomic groups (type 1 and type 2) based on sequence variation within the gene encoding the major adhesion molecule P1. Although numerous publications have described real-time PCR assays for the detection of M. pneumoniae, none has been able to discriminate the two genomic types. Here, a real-time PCR assay that can distinguish each type of M. pneumoniae utilizing high-resolution melt-curve analysis is reported. Using this method, 102 isolates obtained from patients from 1965 to the present, including those from recent outbreaks, were typed along with reference strains M129 (type 1) and FH (type 2). The results show that 55 isolates (54%) can be classified as type 1 and 47 isolates (46%) as type 2, and 100% correlation was demonstrated when compared with a standard PCR-restriction fragment length polymorphism typing procedure. Typing of isolates obtained from recent outbreaks in the USA has revealed the presence of both types. This assay provides a rapid, reliable and convenient method for typing M. pneumoniae isolates and may be useful for surveillance purposes and epidemiological investigations, and may provide insight into the biology of M. pneumoniae distribution within populations.

Background We investigated an outbreak of severe neurologic disease and pneumonia that occurred among students at 4 schools in Rhode Island. Methods We identified cases of encephalitis, encephalomyelitis, and pneumonia that occurred among schoolchildren from 1 September 2006 through 9 February 2007, and we performed serologic tests, polymerase chain reaction (PCR) analysis, and culture for the detection of multiple pathogens in oropharyngeal and nasopharyngeal specimens. Students with positive results of M. pneumoniae IgM serologic testing and no alternative diagnosis were considered to be infected with M. pneumoniae. At school A, we used questionnaires to identify students and their household contacts who made visits to physicians for pneumonia and cough. We compared observed and expected rates of pneumonia. Results Rates of pneumonia among elementary students (122 cases/1000 student-years) were > 5-fold higher than expected. Three students had encephalitis or encephalomyelitis, and 76 had pneumonia. Of these 2 groups of students, 2 (66%) and 57 students (75%), respectively, had M. pneumoniae infection. M. pneumoniae was detected by PCR in 10 students with pneumonia; 5 of these specimens were cultured, and M. pneumoniae was isolated in 4. Of 202 households of students attending school A, 20 (10%) accounted for 61% of visits to physicians for pneumonia or cough. Of 19 household contacts of students with pneumonia, 8 (42%) developed pneumonia and 6 (32%) reported visits for cough. Conclusions M. pneumoniae caused a community-wide outbreak of cough illness and pneumonia and was associated with the development of life-threatening neurologic disease. Although M. pneumoniae was detected in schools, its transmission in households amplified the outbreak. Interrupting household transmission should be a priority during future outbreaks.

Mycoplasma pneumoniae is a significant cause of community-acquired pneumonia, which is often empirically treated with macrolides or azalides such as erythromycin or azithromycin. Recent studies have discovered the existence of macrolide-resistant strains within the population that have been mapped to mutations within the domain V region of the 23S rRNA gene. Currently, identification of these resistant strains relies on time-consuming and labor-intensive procedures such as restriction fragment length polymorphism, MIC studies, and sequence analysis. The current study reports two distinct real-time PCR assays that can detect the A2063G or A2064G base mutation (A2058G or A2059G by Escherichia coli numbering) conferring macrolide resistance. By subjecting the amplicon of the targeted domain V region of the 23S rRNA gene to a high-resolution melt curve analysis, macrolide-resistant strains can quickly be separated from susceptible strains. Utilizing this method, we screened 100 clinical isolates and found 5 strains to possess mutations conferring resistance. These findings were concordant with both sequencing and MIC data. This procedure was also used successfully to identify both susceptible and resistant genotypes in 23 patient specimens. These patient specimens tested positive for the presence of M. pneumoniae by a separate real-time PCR assay, although the bacteria could not be isolated by culture. This is the first report of a real-time PCR assay capable of detecting the dominant mutations that confer macrolide resistance on M. pneumoniae , and these assays may have utility in detecting resistant strains of other infectious agents. These assays may also allow for clinicians to select appropriate treatment options more rapidly and may provide a convenient method to conduct surveillance for genetic mutations conferring antibiotic resistance.

Mycoplasma pneumoniae is an important etiologic agent of primary atypical pneumonia in children and adults. The diagnosis of M. pneumoniae infection is commonly confirmed through serologic testing. In this study, we used paired sera from 51 patients (all with confirmed M. pneumoniae infection and positive complement fixation [CF] titers) to compare the results of eight enzyme immunoassays (EIAs) available commercially in the United States. We compared two single-use EIAs and six plate-type EIAs. Results from acute-phase sera ranged from only 7 (14%) positive by ImmunoWELL (GenBio) immunoglobulin M (IgM) EIA to 23 (45%) positive by Zeus IgG EIA. When both the acute-phase and convalescent-phase serum samples were examined, positive results ranged from 20 (39%) by the ImmunoWELL (GenBio) IgM assay to 45 (88%) positive by the Remel IgG-IgM EIA. In this study, the single-use EIAs by Remel and Meridian were more reliable than were the plate-type EIAs. Among the plate-type EIAs, the Zeus and DiaSorin assays (which detect antibodies to protein antigens) were more sensitive than the ImmunoWELL assay (which detects antibodies to glycolipid antigens). In general, IgG EIAs on convalescent-phase sera were more concordant with one another than were IgM EIAs with one another. Scatter plot analysis of convalescent-phase sera showed that, as the CF titer dropped, the IgM assays identified fewer positive convalescent-phase sera. In contrast, the IgG assays provided fairly consistent positive results for convalescent-phase sera with CF titers of 64 and above. Results of individual tests and overall limitations of serodiagnostics for M. pneumoniae infections are discussed.

This chapter explores the fascinating biology and history of human mycoplasma infections with a focus on disease caused by Mycoplasma pneumoniae. It addresses the association of M. pneumoniae infections with asthma. An important question is whether mycoplasmas are a primary cause of asthma, or if mycoplasma infection is at least a cofactor in the development of asthma. The suggestion that M. pneumoniae may play a role beyond simple, acute exacerbation comes from multiple lines of evidence. First, M. pneumoniae can be recovered more often from the airways of patients with chronic, stable asthma than from matched control patients . Second, chronic reductions in lung function tests 1 year, 3 years, and up to 9 years or more after an episode of mycoplasma respiratory tract infection have been reported, establishing the ability of mycoplasmas to induce chronic to permanent lung damage long after resolution of respiratory tract symptoms. Third, M. pneumoniae is known to induce a number of inflammatory mediators implicated in the pathogenesis of asthma. A number of important research questions need renewed attention to provide a better understanding of pathogenesis. These include the role of autoimmunity induction in a variety of mycoplasma syndromes, the existence and importance of an asymptomatic carrier state, and a much better understanding of the role of direct invasion in extrapulmonary syndromes.

Outbreaks of Mycoplasma pneumoniae (MP) in closed communities can have a high attack rate and can last several months. Azithromycin chemoprophylaxis has not been evaluated as a means of limiting transmission. This randomized, double-blinded placebo-controlled trial of azithromycin was conducted among asymptomatic hospital employees during an MP outbreak. Oropharyngeal swabs were obtained for detection of MP by polymerase chain reaction, and questionnaires were administered to assess clinical illness. Of the 147 employees who were enrolled, 73 received azithromycin and 74 received placebo. Carriage was similar within and between groups at weeks 1 and 6 (9.6% vs. 6.7% and 10.3% vs. 13.2%, respectively). Four episodes of clinically significant respiratory illness occurred in the azithromycin group versus 16 episodes in the placebo group (protective efficacy, 75%; 95% confidence interval, 28%-91%). Use of azithromycin prophylaxis in asymptomatic persons during an MP outbreak in a closed setting may be of value in reducing clinical illness.

Mycoplasma pneumoniae is a leading cause of community-acquired pneumonia. Although two genetically distinct types of M. pneumoniae are known, variants of each also exist. We used a real-time PCR high-resolution melt genotyping assay to identify clinical variants which may provide greater insight into the genetic distribution of M. pneumoniae strains.

Methods: We analyzed 2 outbreaks of CAP due to M. pneumoniae. Nasopharyngeal and/or oropharyngeal swab specimens and serum samples were obtained from persons with clinically defined cases, household contacts, and asymptomatic individuals. Real-time polymerase chain reaction (PCR) for M. pneumoniae was performed on all swab specimens, and the diagnostic utility was compared with that of 2 commercially available serologic test kits. Results: For cases, 21% yielded positive results with real-time PCR, whereas 81% and 54% yielded positive results with the immunoglobulin M and immunoglobulin G/immunoglobulin M serologic tests, respectively. For noncases, 1.8% yielded positive results with real-time PCR, whereas 63% and 79% yielded serologically positive results with the immunoglobulin M and immunoglobulin G/immunoglobulin M kits, respectively. The sensitivity of real-time PCR decreased as the duration between symptom onset and sample collection increased, with a peak sensitivity of 48% at 0-21 days. Real time PCR may be most sensitive when collected within 21 days of symptoms. A specificity of 43% for the immunoglobulin M antibody detection assay was observed for persons aged 10-18 years, but the sensitivity increased to 82% for persons aged 19 years. The IgM antibody Meridian test was more sensitive than the Remel test. Discussion: No single test proved reliable for the identification of an outbreak of CAP due to M. pneumoniae. A combination of testing methodologies proved to be the most reliable approach for identification of outbreaks of CAP due to M. pneumoniae, especially in the absence of other suspected respiratory pathogens. Editorial Comment by Victor L Yu : Laboratory diagnosis for Mycoplasma pneumoniae in sporadic pneumonia (as opposed to outbreak) cases is unreliable. Serologic tests can be used but acute and convalescent samples need to be drawn to allow a 4-fold increase in antibody titer rise. Definitive diagnosis for M pneumoniae remains elusive.

The interaction between hosts and the viruses that infect them is a dynamic one, and a growing literature documents the fact that many viruses have developed mechanisms designed to avoid elimination by the host immune system. One of the immune strategies used by the host and targeted by virus proteins is apoptosis triggered by the cytokine tumor necrosis factor (TNF). Mouse fibroblast LM cells are spontaneously sensitive to TNF. When the wild-type E6 protein from the human papillomavirus type 16 (HPV 16) was expressed in LM cells, the cells became resistant to TNF. This resistance was examined by several means, including cell morphology, the dose- and time-independent response to TNF in a cell death ELISA, trypan blue exclusion, and cell proliferation. The level of p53 did not rise in TNF-treated cells prior to apoptosis, suggesting a p53-independent mechanism. Significant, though not complete, resistance to TNF was also observed following transfection of a plasmid expressing a mutant E6 protein, which is unable to mediate rapid degradation of the p53 tumor suppressor. These results indicate that the HPV 16 E6 protein can protect LM cells from TNF-triggered apoptosis and likely does so by a mechanism other than mediation of p53 degradation.

The bioterrorism-associated human anthrax epidemic in the fall of 2001 highlighted the need for a sensitive, reproducible, and specific laboratory test for the confirmatory diagnosis of human anthrax. The Centers for Disease Control and Prevention developed, optimized, and rapidly qualified an enzyme-linked immunosorbent assay (ELISA) for immunoglobulin G (IgG) antibodies to Bacillus anthracis protective antigen (PA) in human serum. The qualified ELISA had a minimum detection limit of 0.06 µg/mL, a reliable lower limit of detection of 0.09 µg/mL, and a lower limit of quantification in undiluted serum specimens of 3.0 µg/mL anti-PA IgG. The diagnostic sensitivity of the assay was 97.8%, and the diagnostic specificity was 97.6%. A competitive inhibition anti-PA IgG ELISA was also developed to enhance diagnostic specificity to 100%. The anti-PA ELISAs proved valuable for the confirmation of cases of cutaneous and inhalational anthrax and evaluation of patients in whom the diagnosis of anthrax was being considered. aturally occurring anthrax is a zoonotic disease of herbivores, with low-level sporadic infection of humans. Since 1950, human anthrax in the United States was confined to those occupationally at risk, with only 235 confirmed cases, mostly cutaneous, reported from 1955 to 2002 (1–3). The occurrence of human anthrax in the country and the public perception of the disease changed dramatically in the fall of 2001, with the first successful bioterrorist anthrax attack on the U.S. civilian population. This event necessitated the simultaneous development and application of qualified laboratory assays— including serologic assays—to evaluate patients suspected of having anthrax. The major obstacle to serologic analysis of human anthrax