Mycoplasma infection is the most prevalent contamination in cell culture. Analysis of cell culture in laboratories from different countries shows that mycoplasma contamination ranges from 15% to 80% and, in some cases, even reaches 100% (Chernov et al., 2014). Whilst mycoplasma infection is not visible to the naked eye in cell culture, the consequences of mycoplasma contamination have been shown to induce a number of cellular changes, for example, increased resistance to chemotherapeutic drugs. Therefore, any results obtained from tissue culture studies, in the presence of mycoplasma contamination, potentially render the data invalid (Kim et al., 2015; Gedye et al., 2016). As such, mycoplasmas are not harmless bystanders and cannot be ignored in in vitro studies.
Arginine/pharmacology* ; HEK293 Cells ; Humans ; Mycoplasma/isolation & purification* ; Plasmids ; Transfection

OBJECTIVEMycoplasma pneumoniae (Mp) infection is one of major causes of community-acquired pneumonia. Isolation and culture of Mp are very difficult, fluorescent quantitative PCR is a new technique to detect Mp. The aim of this study was to explore the diagnostic value of fluorescent quantitation PCR for Mp infection.

METHODMp-DNA from the deep respiratory tract secretion of children suffering from pneumonia was tested by a fluorescent quantitative PCR. Totally 256 cases who were positive for Mp DNA were enrolled into this study, 164 (64.1%) were male, 92 (35.9%) were female; the age ranged from 9 days to 16 years. All the patients also had results of Mp-IgM test. These patients were divided into 2 groups according to the result of Mp-IgM detection, namely, Mp-IgM positive and negative groups. Area under the roc curve (Az) was used as the index to evaluate the diagnostic value of fluorescent quantitation PCR for Mp detection. The number of Mp-DNA copies, age and course of disease of the 2 groups were also compared.

RESULTS(1) Diagnostic accuracy of fluorescent quantitative PCR for detecting Mp infection was that Az = 0.641. (2) The number of copies of the cases in Mp-IgM positive group was 5.42 +/- 1.26 [log(Mp-DNA copy/ml)], while that of Mp-IgM-negative group was 4.87 +/- 1.29 [log(Mp-DNA copy/ml), t = 3.43, P < 0.05]. (3) The age of Mp-IgM positive group was dramatically younger than Mp-IgM negative group (P < 0.001).

CONCLUSIONThe diagnostic accuracy of fluorescent quantitative PCR for mycoplasma pneumoniae (Mp) infection is low; however for children whose immunologic systems are not fully developed, this technique has some diagnostic value, and higher number of Mp-DNA copies may support diagnosis of Mp infection.

Adolescent ; Child ; Child, Preschool ; Female ; Humans ; Immunoglobulin M ; blood ; Infant ; Infant, Newborn ; Male ; Mycoplasma pneumoniae ; genetics ; immunology ; isolation & purification ; Pneumonia, Mycoplasma ; diagnosis ; Polymerase Chain Reaction ; methods

OBJECTIVEThe P1 protein of Mycoplasma pneumoniae (MP) plays an important role in the pathogenesis of MP pneumonia. It mediates the attachment of the pathogen to host cells and elicits a strong humoral immune response during infection. In early studies, only two types of MP P1 genes were assumed to exist. Later, eight subtypes of MP P1 genes and some variations of P1 gene were reported. However, there are no related reports in China until now. This study aimed to understand epidemiology of MP subtype in Zhejiang province, China, as well as the relationship between MP subtype and clinical severity of MP pneumonia.

METHODClinical samples were collected by nasopharyngeal aspiration from children with MP pneumonia hospitalized in the Children's Hospital of Zhejiang University School of Medicine from February to December in 2009. P1 gene fragment was amplified by using PCR method (with primers of ADH1/ADH2 and ADH3/ADH4, respectively). Then ADH1/ADH2-generated fragments were digested with HaeIII, HpaII, Sau3A, and the ADH3/ADH4-generated fragments digested with HaeIII, Sau3A, HhaI, RsaI. The MP P1 subtypes were determined based on resulting fragments. Part of samples were selected for sequencing. The clinical data of different MP subtype pneumonia were compared.

RESULTA total of 300 hospitalized children with MP pneumonia were enrolled in this study. All the samples produced specific bands for MP P1 gene after PCR with primers of ADH1/ADH2 and ADH3/ADH4 respectively. By restrictive fragment length polymorphism analysis, 297 clinical specimens showed the characteristic band patterns for P1 type 1 identical to Mp129, and only 3 clinical specimens showed the characteristic band pattern for P1 type 2 identical to MP-FH. All P1 type 1 and P1 type 2 showed the same subtype bands respectively, as subtype 1b and 2a. After sequencing, one synonymous point mutation in P1 type 1 was identified relative to the MP129 P1 sequence at nucleotide position (nt) 208(G→A). Three cases with P1 type 2 MP pneumonia were found to have liver damage, and longer hospital stay and fever duration than P1 type 1, but no statistically significant difference was found.

CONCLUSIONClinical samples can be used directly for genotyping of MP. The dominating type of MP in Zhejiang Province was P1 type 1 subtype 1b. But whether there was any relationship between MP subtype and clinical severity remains to be clarified.

Adhesins, Bacterial ; genetics ; Child ; China ; DNA, Bacterial ; genetics ; Genotype ; Humans ; Mycoplasma pneumoniae ; genetics ; isolation & purification ; Nasopharynx ; microbiology ; Pneumonia, Mycoplasma ; microbiology ; Polymorphism, Restriction Fragment Length

Anti-Bacterial Agents ; pharmacology ; therapeutic use ; Child ; Drug Resistance, Bacterial ; Humans ; Mycoplasma pneumoniae ; drug effects ; isolation & purification ; Pneumonia, Mycoplasma ; drug therapy ; microbiology

OBJECTIVETo summarize the epidemiology and clinical characteristics of Mycoplasma pneumoniae (MP) infection in children with acute respiratory tract infection (ARI) in Guangzhou.

METHODSMP was detected using an indirect immunofluorescent method in 2084 children with ARI. The relations between MP infection rate and the gender, age, season, site of infection and wheezing diseases were analyzed.

RESULTSA total of 433 children (20.8%) were positive for MP, including 222 boys (19.8%) and 211 girls (21.9%) without significant difference in the infection rate between the genders (P>0.05). In 0- to 3-year-old group, 106 children were positive for MP (15.0%), while in 3- to 5-year-old group and 5- to 14-year-old group, 163 (25.2%) and 164 (22.5%) were positive, respectively, showing a significant difference in the infection rate between the 3 groups (P<0.05). The MP infection rate was 18.0% in January to March, 25.1% in April to June, 17.7% in July to September, and 20.5% in October to December, showing significant differences between the periods (P<0.05). No significant difference was found in the infection rate between children with acute upper respiratory tract infection (URI) and those with lower respiratory tract infection (LRI) (P>0.05). Among the children with LRI, those having wheezing disease had significantly higher MP positivity rate than those without wheezing.

CONCLUSIONMP is a common causative agent for ARI in children. MP infection is not related to gender and infection site, but to age and season. Children over 3 years old are vulnerable to MP infection. MP infection can be associated with wheezing in LRI.

Adolescent ; Age Factors ; Child ; Child, Preschool ; China ; epidemiology ; Female ; Humans ; Male ; Mycoplasma pneumoniae ; isolation & purification ; Pneumonia, Mycoplasma ; epidemiology ; microbiology ; Prevalence ; Respiratory Tract Infections ; epidemiology ; microbiology ; Retrospective Studies ; Seasons

Objective To explore the clinical value of one-step visualization loop-mediated isothermal amplification(LAMP)in the detection of Mycoplasma pneumoniae(Mp). Methods One-step visualized LAMP,polymerase chain reaction(PCR),and enzyme-linked immunosorbent assay(ELISA)were used to simultaneously detect 108 clinical Mp specimens in children,which included 73 cases of Mp infection diagnosed by PCR and 35 cases of other chronic/acute respiratory tract infections.On the first day of admission,one-step visualization LAMP,PCR(fluorimetric method),and ELISA were used to test the throat swab and serum sample obtained from the same patient,and the Kappa value was calculated.The consistence between LAMP and PCR and that between LAMP and ELISA were compared.On the fifth day of admission,40 patients were resampled and the findings of these three tests on the first day and on the fifth day were compared. Results One-step visualization LAMP had a sensitivity of 100% and a specificity of 94.3%,whereas ELISA had a sensitivity of 65.8% and a specificity of 82.9%.The ratio of Kappa camparing one-step visualization LAMP and PCR was 0.956 and the ratio of Kappa camparing one-step visualization LAMP and ELISA was 0.38.The number of positive specimens detected by LAMP was higher than that by ELISA on the first day. Conclusions One-step visualization LAMP has excellent sensitivity and specificity in detecting early acute Mp infection.It has high consistency with PCR and can be applied to detect Mp.
Child ; Enzyme-Linked Immunosorbent Assay ; Humans ; Mycoplasma pneumoniae ; isolation & purification ; Nucleic Acid Amplification Techniques ; Pneumonia, Mycoplasma ; diagnosis ; Polymerase Chain Reaction ; Sensitivity and Specificity

OBJECTIVETo investigate the incidence and clinical features of mixed infections in children with Mycoplasma pneumoniae (MP) pneumonia.

METHODA total of 201 cases diagnosed as MP pneumonia were investigated for mixed infections by sputum bacterial culture, respiratory virus antigen detection and serum Chlamydia pneumoniae antibody test. For those with the indications for bronchoscopy, we also did bronchoalveolar lavage and lavage bacterial culture.

RESULTA high incidence (103/201, 51.2%) of mixed infections in children with MP pneumonia was revealed. The most frequent co-infected pathogen was Chlamydia pneumoniae (52, 25.9%), followed by viruses (29, 14.4%), and bacteria (22, 10.9%). Among viruses, respiratory syncytial virus was the most common (17, 8.5%), followed by adenovirus (6, 3.0%), parainfluenza virus type III (4, 2.0%) and influenza virus type B (2, 1.0%). Sputum bacterial culture was positive in 14/201 (7.0%) cases, Streptococcus pneumonia being most common (6, 3.0%). BALF culture yielded positive results in 11.6% (8/69), Streptococcus pneumonia was also common (5, 7.3%). Among 29 cases with MP and virus coinfection, 26 were younger than 3 years (89.7%), while for MP and Chlamydia pneumoniae coinfection, most of them were older than 3 years (40/52, 76.9%). Compared with non-mixed infections, those with mixed infections had longer fever duration (24.5% and 40.8% longer than 10 d), more frequently developed pleural effusion (11.2%, 23.3%) and large area of shadow in chest imaging (35.7%, 51.5%). White blood cell [(14.28 ± 4.99) × 10(9)/L], C-reactive protein (CRP) [69(32.5 - 99.5) mg/L] and neutrophil ratio in BALF [0.86 (0.63 - 0.91)] were much higher in children with mixed bacterial infections than that in non-mixed infections [(9.06 ± 3.47) × 10(9)/L, 3 (0 - 31.0) mg/L, 0.44 (0.03 - 0.88)]. But no significant difference was found in peripheral blood neutrophil proportion between mixed bacterial infections (0.38 ± 0.25) and non-mixed infections (0.51 ± 0.19).

CONCLUSIONMore than half of cases with MP pneumonia had mixed infections, most commonly caused by Chlamydia pneumonia followed by viruses. The incidence of mixed infections with bacteria was low. Mixed infections with virus were more common in young children, while mixed infection with Chlamydia pneumoniae was more common in older ones. Bacterial infections should be paid more attention, especially those caused by Streptococcus pneumoniae, for those with high peripheral white blood cell counts, high CRP levels and high proportion of neutrophils in BALF.

Adolescent ; Child ; Child, Preschool ; Chlamydophila pneumoniae ; isolation & purification ; Coinfection ; Female ; Humans ; Infant ; Inpatients ; Male ; Mycoplasma pneumoniae ; isolation & purification ; Pneumonia, Mycoplasma ; diagnosis ; microbiology ; virology ; Pneumonia, Viral ; diagnosis ; Respiratory Syncytial Viruses ; isolation & purification

Anti-Bacterial Agents ; therapeutic use ; Anti-Inflammatory Agents ; therapeutic use ; Asthma ; drug therapy ; etiology ; Child ; Child, Preschool ; Humans ; Macrolides ; therapeutic use ; Mycoplasma Infections ; complications ; drug therapy ; Mycoplasma pneumoniae ; isolation & purification ; Pneumonia, Mycoplasma ; complications ; drug therapy ; Respiratory Sounds ; drug effects ; etiology

OBJECTIVETo explore the relationship between the colonization of group B streptococci (GBS), mycoplasma,and Chlamydia trachomatis (CT) infections and spontaneous abortion due to early embryonic death.

METHODSTotally 74 patients (study group) who experienced the missed abortion during their first or second trimester and 62 women (control group) who underwent induced termination of normal pregnancy during the first or second trimester were enrolled in this study. The vaginal secretions, intrauterine aspirates, and amniotic fluids were collected for GBS culture. Cervical mycoplasma (UU+MH) and CT were detected at the same time.

RESULTSPositive results of GBS culture of vaginal secretions were detected in 9 patients (12.16%) in the study group, but in only 6 patients (9.68%) in control group (P=0.662). The intrauterine aspirate samples (as well as the amniotic fluid samples) of all cases were negative in GBS culture. The positive rates of UU and MH were 32.43% (24/74) and 16.22% (12/74) in the study group, but were only 10.35% (12/62) (P=0.0103) and 6.45 (4/62) (P=0.042) in the control group. The positive rate of CT was 8.11% (6/74) in the study group and 8.06 % (5/62) in the control group (P=0.905). The rate of concurrent infection of GBS and mycoplasma was 4.05% (3/74) in the study group and 6.45% (4/62) in the control group (P=0.743). The rate of concurrent infection of GBS and CT was 0 in the study group and 1.61% (1/62) in the control group (P=0.475). The rate of concurrent mycoplasma and CT infection was 2.70% (2/74) in the study group and 0 in the control group (P=0.325). Furthermore, no one was positive for poly infection of all these three pathogens.

CONCLUSIONSGBS may be positive in the genital tract of some pregnant women but is not related with early abortion. The mycoplasma infection may be one of the reasons leading to arrested intrauterine pregnancy.

Abortion, Spontaneous ; microbiology ; Adult ; Cervix Uteri ; microbiology ; Chlamydia trachomatis ; isolation & purification ; Female ; Humans ; Mycoplasma ; isolation & purification ; Pregnancy ; Streptococcus agalactiae ; isolation & purification ; Young Adult

OBJECTIVETo investigate the distribution of pathogens of children with community acquired pneumonia (CAP) from the Chongqing area.

METHODSNasopharyngeal specimens and blood specimens of 1 613 children with CAP were collected between January 2014 and December 2014 for bacterial culture and detection of 7 respiratory viruses and antibodies against Mycoplasma pneumoniae (MP).

RESULTSThe overall positive rate of bacteria was 50.22% (810 cases). Hemophilus parainfluenzae (40.8%), Streptococcus pneumonia (29.7%) and Moraxelle catarrhalis (7.3%) were the predominant ones. Among the viruses, the top detected virus was respiratory syncytial virus (RSV, 58.3%), followed by parainfluenza virus type3 (17.4%) and adenovirus (14.3%). A total of 481 cases (29.82%) were MP-positive. The co-infection rate was 32.18% (519 cases), and the mixed infections of bacteria and viruses were common (47.4%).

CONCLUSIONSRSV and Hemophilus parainfluenzae are the major pathogens of CAP in children from the Chongqing area. MP is also an important pathogen. The co-infection of bacteria and viruses is prevalent.

Adolescent ; Child ; Child, Preschool ; Community-Acquired Infections ; etiology ; Female ; Haemophilus parainfluenzae ; isolation & purification ; Hospitalization ; Humans ; Infant ; Male ; Mycoplasma pneumoniae ; isolation & purification ; Pneumonia ; etiology ; Respiratory Syncytial Viruses ; isolation & purification