No abstract

Over the past four decades, the incidence of meningitis caused by Haemophilus influenzae in children has decreased due to widespread vaccination against H. influenzae type b (Hib). The incidence of invasive diseases due to H. influenzae types not included in the vaccines, however, has increased. At present, there are a limited number of diagnostics available to detect non-type b H. influenzae. To address this issue, we developed a rapid, simple, and cost-effective method for detecting serotypes of H. influenzae. We designed LAMP primer sets based on published sequences for H. influenzae capsular types a, c, d, e, and f. The assay was evaluated to determine test reactivity, specificity, and sensitivity. To support its use in patients with suspected meningitis, we evaluated the detection limit of the non-Hib serotype specific LAMP assay using bacterial genomic DNA-spiked cerebrospinal fluid (CSF) specimens. The reactivity and specificity of the LAMP assays were confirmed using six serotypes and non-typeable H. influenzae strains, plus eight strains of other Haemophilus species and non-Haemophilus genera. The detection limits of the LAMP assay for capsular types a, c, d, e, and f were 102, 102, 102, 103, and 10 copies per reaction, while those of the PCR assay were 104, 104, 103, 103, and 104 genome copies per reaction, respectively. Using DNA-spiked CSF specimens, the detection limit of the LAMP assay was equivalent to that using purified DNA as the template. However, the detection limit of the PCR was reduced from 103 to 104 genome copies per reaction for serotype d and from 103 to 105 genome copies per reaction for serotype e. To the best of our knowledge, this is the first report of a serotype-specific identification assay for H. influenzae using the LAMP method. Our results suggest the potential of LAMP methods for patients with suspected meningitis in resource-limited laboratories or public health surveillance systems.

No abstract

In children, the incidence of pneumococcal meningitis has decreased since the introduction of pneumococcal conjugate vaccine (PCV7 and PCV13). However, since the introduction of the vaccine, developed countries have seen the emergence of non-PCV13 serotypes. However, invasive pneumococcal disease (IPD) caused by PCV13-targeted serotypes still represents an important public health problem in resource-limited countries. To develop a rapid, simple, and cost-effective assay to detect serotypes of Streptococcus pneumoniae, we developed a novel loop-mediated isothermal amplification (LAMP) assay based on the sequences available for the 13 capsular types that are included in PCV13: 1, 3, 4, 5, 6 A, 6B, 7 F, 9 V, 14, 18 C, 19 A, 19 F, and 23 F. We evaluated test reactivity, specificity, sensitivity and performance, and compared the results between established LAMP and conventional PCR assays. To support its clinical use, the detection limits of the LAMP assay were evaluated using bacterial genomic DNA-spiked cerebrospinal fluid (CSF) and blood specimens. We confirmed the specificity of the LAMP assay using 41 serotypes of pneumococcal strains. The sensitivity of the LAMP assay was 10 to 100 copies per reaction, compared to 10 to 104 copies per reaction for PCR assays. The detection limits of the LAMP assay were comparable when using DNA-spiked CSF and blood specimens, as compared to using purified DNA as the template. In conclusion, a rapid and simple LAMP-based pneumococcal serotyping method has been developed. This is the first report of a LAMP method for a PCV13 serotype-specific identification assay, which could be a promising step to facilitate epidemiological studies of pneumococcal serotyping.

Zika virus infection (ZVI) is a great concern for human health because it frequently causes fetal anomalies. Little is known about pathophysiology of ZVI because it has been regarded as a mild, no life-threatening infection. However, the latest endemic in