Development of a droplet digital PCR method for detection of Streptococcus agalactiae

Zeng, Y.; Chen, Chu-Mao; Li, Xiaoyan; Chen, Jun-Jiang; Wang, Yange; Ouyang, Shi; Ji, Tianxing; Xia, Yong; Guo, Xu‐Guang

doi:10.1186/s12866-020-01857-w

Cited by 19 publications

(7 citation statements)

References 24 publications

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“…Then, PCR was performed on a Bio-Rad T100™ PCR Thermal Cycler using the following conditions: predenaturation for 1 cycle at 95°C for 3 min; denaturation for 40 cycles at 95°C for 30 s; and annealing and extension for 40 cycles at 59°C for 1 min (with a ramp rate of 2.5°C/s). Finally, the uorescence signal in each plate was analyzed by a QX200™ Droplet Reader and QuantaSoft™ Version 1.7.4, and each reaction used a negative control (12). The threshold can be manually set according to the results of the negative control.…”

Section: Droplet Digital Pcr (Ddpcr) Assay For Gbsmentioning

confidence: 99%

To evaluate the performance of simultaneous amplification and testing assay for group B Streptococcus detection: comparison with real-time PCR and ddPCR assays

Lu,

Chen,

Wang

et al. 2024

Preprint

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Background To evaluate the performance of simultaneous amplification and testing (SAT) assay for the detection of group B Streptococcus (GBS) in maternal vaginal and perianal swabs compared with real-time polymerase chain reaction (RT‒PCR). Methods We obtained vaginal and perianal swabs from 1474 pregnant women at the Obstetrics and Gynecology Hospital of Fudan University (Shanghai, China) between April 2023 and June 2023. Vaginal and perianal swabs were collected at 35–37 weeks of gestation. Swabs were tested for GBS simultaneously by using the SAT assay and RT‒PCR, and a comparative analysis (kappa coefficient) was performed. Furthermore, we conducted additional droplet digital PCR (ddPCR) tests to confirm the results when there were controversial results between SAT and RT‒PCR. In addition, we compared the limit of detection, technical specificity, repeatability and reproducibility of SAT-GBS with those of routine RT‒PCR assays. Results In our study, the rate of clinical GBS colonization according to the SAT assay was 11.5% (169/1471). The SAT assay showed a sensitivity of 91.8%, a specificity of 99.9%, a diagnostic accuracy of 98.9%, a positive predictive value (PPV) of 99.4% and a negative predictive value (NPV) of 98.8%. The kappa value between RT‒PCR and SAT was 0.917. Conclusions This SAT assay for the detection of group B Streptococcus is not only easy to perform but can also detect GBS sensitively and specifically and may be used in the regular molecular diagnosis of GBS in cases of newborn sepsis and meningitis.

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Section: Droplet Digital Pcr (Ddpcr) Assay For Gbsmentioning

confidence: 99%

To evaluate the performance of simultaneous amplification and testing assay for group B Streptococcus detection: comparison with real-time PCR and ddPCR assays

Lu,

Chen,

Wang

et al. 2024

Preprint

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“…Colonization of S. agalactiae exhibits significant differences between different populations, which may be related to race, ethnicity, and genetic composition (19). It is worth noting that the prevalence is high in Italy (21.9%, sampling from ear, throat and rectum) and Gambia (24.8%, sampling from nasopharynx and rectum), but low in Pakistan (3.1%, sampling from abdominal skin and ears) and Greece (2.4%, sampling from the ear canal, throat and umbilical cord) (20). These findings indicate the importance of sampling points in determining the colonization of neonatal S. agalactiae.…”

Section: S Agalactiae Genotype and Erythromycin Resistance In Neonata...mentioning

confidence: 99%

Experimental Study on Streptococcus agalactiae Genotype and Erythromycin Resistance in Neonatal Sepsis

Shen

Huang

Xie

2022

Cell Mol Biol (Noisy-le-grand)

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This study aimed to evaluate Streptococcus agalactiae genotype and erythromycin resistance in neonatal sepsis. After obtaining the mothers’ informed consent, trained nurses sampled 430 neonatal specimens of sepsis from the ear canal, oral cavity and umbilical cord immediately after childbirth and implemented a cross-sectional study. By Gram staining, morphology, hemolysis mode, catalase and CAMP tests, the isolate was identified as S. agalactiae. All 455 isolates were tested for antimicrobial susceptibility by the disc diffusion method. Multilocus sequence typing was used to serotype S. agalactiae involving sequencing of 7 housekeeping genes. The erythromycin resistance genes-erm (B), erm (A) and mef (A) were detected by PCR. Results showed that there were 286 cases (66.51%) of neonates delivered naturally, and 144 cases (33.49%) of neonates delivered by cesarean section. A total of 455 strains were tested, including 253 strains (55.60%) of Gram-positive bacteria with 100 strains (21.98%) of S. agalactiae and 52 strains (11.43%) of Staphylococcus epidermidis, 178 strains (39.12%) of Gram-negative bacteria with 45 strains of Klebsiella pneumoniae (9.89%), 36 strains of Escherichia coli (7.91%), 36 strains of Pseudomonas aeruginosa (7.91%), and 323 strains of Citrobacter freundii (7.03%). S. agalactiae had the highest resistance of 87 (87.00%) to erythromycin, followed by resistance to azithromycin 83 (83.00%) and clindamycin 78 (78.00%). In children with neonatal sepsis, S. agalactiae serotypes were mainly Ia, Ib, and III, accounting for 29.00%, 35.00%, and 19.00% respectively. The main genotypes were ST651, ST103 and ST176, which account for 19.00%, 17.00% and 15.00% respectively. The ST19 type 13.00%, ST27 type 8.00%, ST17 Type 11.00%, ST10 type 12.00%, ST485 type 5.00%. The ST103 and ST485 isolates were classified as serotype Ia, the ST10 and ST176 isolates were classified as serotype Ib, and ST17 and ST19 isolates were classified as serotype III. Among the strains of S. agalactiae, 40.23% (35/87) carry erm (A) gene, 35.63% (31/87) carry erm (B) gene, and 24.14% (21/87) carry mef (A) gene. erm (A) gene was the most common gene in ST19 strain (7/11, 63.64%), and erm (B) gene was the most common gene in ST176 and ST651 strains (6/12, 50.00%; 8/18, 44.44%), while mef (A) gene was the most common gene in ST17 strain (5/11, 45.45%). In general, S. agalactiae genotypes in neonatal sepsis were mainly ST651, ST103 and ST176, and the main serotypes are Ia, Ib, and III. There was good consistency between ST and serotype, and a significant difference was shown in erythromycin resistance and ST distribution, which highlights the value of new epidemiological trend detection by monitoring multiple characteristics and provides inspiration for the development of multivalent S. agalactiae vaccines.

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“…Another drawback is the need for qualified personnel and specialist knowledge of the techniques used. High requirements limit the use of these methods in many laboratories [ 20 , 21 ]. For all these reasons, it is important to create and develop fast, less costly tests for the specific detection of GBS, which can help in the early diagnosis of GBS infection.…”

Section: Introductionmentioning

confidence: 99%

Ultra-Fast Impedimetric Immunoassay for Detection of Streptococcus agalactiae Using Carbon Electrode with Nanodiamonds Film

Bigus,

Lewandowska,

Bięga

et al. 2023

Micromachines

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This publication presents the results of work on the development of a quick and cheap electrochemical immunosensor for the diagnosis of infections with the pathogen Streptococcus agalactiae. The research was carried out on the basis of the modification of the well-known glassy carbon (GC) electrodes. The surface of the GC (glassy carbon) electrode was covered with a film made of nanodiamonds, which increased the number of sites for the attachment of anti-Streptococcus agalactiae antibodies. The GC surface was activated with EDC/NHS (1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide/N-Hydroxysuccinimide). Determination of electrode characteristics after each modification step, performed using cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS).

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Development of a droplet digital PCR method for detection of Streptococcus agalactiae

Cited by 19 publications

References 24 publications

To evaluate the performance of simultaneous amplification and testing assay for group B Streptococcus detection: comparison with real-time PCR and ddPCR assays

To evaluate the performance of simultaneous amplification and testing assay for group B Streptococcus detection: comparison with real-time PCR and ddPCR assays

Experimental Study on Streptococcus agalactiae Genotype and Erythromycin Resistance in Neonatal Sepsis

Ultra-Fast Impedimetric Immunoassay for Detection of Streptococcus agalactiae Using Carbon Electrode with Nanodiamonds Film

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