Identification of Risk Factors for Infection in an Outbreak of Mycoplasma pneumoniae Respiratory Tract Disease

Klement, Eyal; Talkington, Deborah F.; Wasserzug, Oshri; Kayouf, Raid; Davidovitch, Nadav; Dumke, Roger; Bar‐Zeev, Yael; Ron, M.; Boxman, Jonathan; Thacker, W L; Wolf, Dana; Lazarovich, Tsilia; Shemer‐Avni, Yonat; Glikman, Daniel; Jacobs, Enno; Grotto, Itamar; Block, Colin; Nir-Paz, Ran

doi:10.1086/508458

Cited by 55 publications

(40 citation statements)

References 31 publications

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“…Further community outbreaks of acute respiratory tract infections due to M. pneumoniae have been described worldwide (9,22), and it would be very interesting to evaluate our MLVA scheme in such epidemic situations. The level of discrimination of this typing method should be confirmed by comparing outbreak-related strains to a set of control strains that were isolated from a similar time period and geographical area but that are not epidemiologically related.…”

Section: Resultsmentioning

confidence: 99%

Development of Multiple-Locus Variable-Number Tandem-Repeat Analysis for Molecular Typing of Mycoplasma pneumoniae

et al. 2009

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In this study we report on the development of a multilocus variable-number tandem-repeat (VNTR) analysis (MLVA) method for the molecular typing of Mycoplasma pneumoniae. The genomic content of M. pneumoniae M129 was analyzed for VNTRs, and 5 of the 17 VNTRs identified were selected for use in an MLVA assay. The method was based on a GeneScan analysis of VNTR loci labeled with fluorescent dyes by multiplex PCR and capillary electrophoresis. This approach was applied to a collection of 265 isolates from various European countries, Japan, and Tunisia; and 26 distinct VNTR types were found. The VNTR assay was compared to the P1 adhesin PCRrestriction fragment length polymorphism (RFLP) typing method and showed a far better resolution than the P1 PCR-RFLP method. The discriminatory power of MLVA (Hunter-Gaston diversity index [HGDI], 0.915) for the 265 isolates was significantly higher than that of the P1 PCR-RFLP method (HGDI, 0.511). However, there was a correlation between the typing results obtained by MLVA and the P1 gene PCR-RFLP method. The potential value of MLVA of M. pneumoniae as an epidemiological tool is discussed, and the use of the VNTR markers in further investigations of the potential use of MLVA in outbreaks of M. pneumoniae infections is proposed.Mycoplasma pneumoniae represents one of the most common etiological agents of community-acquired respiratory tract infections, with the clinical courses ranging from mild forms of pharyngitis and tracheobronchitis to severe cases of interstitial pneumonia. M. pneumoniae infections are responsible for 20% or more of all cases of community-acquired pneumonia (22) and occur endemically, with epidemic peaks occurring every 4 to 7 years. School-age children and young adults are the populations that are the most affected.Strain subtyping by molecular methods is a powerful tool for surveillance and outbreak investigation. However, molecular typing is hampered by the fact that M. pneumoniae is a genetically homogeneous species and isolates are poorly differentiated by PCR-restriction fragment length polymorphism (RFLP) analysis of the P1 gene, the most common genotyping method. PCR-RFLP analysis of the gene encoding the P1 protein, a major adhesin that induces a strong humoral immune response during infection, enables the separation of isolates into two types, types 1 and 2 (1, 2, 17). More recent studies used repetitive regions, RepMp2/3 and RepMp4, which can be found in the P1 gene, for molecular typing. Besides the subtype classification, those methods allow the identification of variants of subtypes 1 and 2 (3,5,8).In addition to these molecular typing methods that target only one gene, other methods based on the study of the whole genome have been adapted to M. pneumoniae, such as pulsedfield electrophoresis (1) and multilocus sequence typing (MLST) (4). Pulsed-field electrophoresis is able to differentiate strains into two groups (groups 1 and 2), like P1 gene PCR-RFLP analysis can, and can subdivide group 2 into two subgroups, subgroups 2a and 2b (1). Due t...

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Section: Resultsmentioning

confidence: 99%

Development of Multiple-Locus Variable-Number Tandem-Repeat Analysis for Molecular Typing of Mycoplasma pneumoniae

et al. 2009

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“…It is also a significant cause of community-wide outbreaks which are reported to occur in 3-to 7-year intervals with various incidence rates (10,12,28). Upper and lower respiratory tract symptoms are often mild and self-limiting; however, occasional extrapulmonary complications may develop, sometimes resulting in death (5,7,23).…”

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confidence: 99%

Multilocus Variable-Number Tandem-Repeat Analysis of Mycoplasma pneumoniae Clinical Isolates from 1962 to the Present: a Retrospective Study

et al. 2012

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In this study, we evaluated a recently developed multilocus variable-number tandem-repeat (VNTR) analysis (MLVA) method for the molecular typing of Mycoplasma pneumoniae. The method is based on GeneScan analysis of five VNTR loci throughout the genome which define a specific genotype based on the number of tandem repeats within each locus. A retrospective analysis of 154 M. pneumoniae clinical isolates collected over the last 50 years and a limited (n ‫؍‬ 4) number of M. pneumoniae-positive primary specimens acquired by the CDC was performed using MLVA. Eighteen distinct VNTR types were identified, including two previously unidentified VNTR types. Isolates from several M. pneumoniae community outbreaks within the United States were also analyzed to examine clonality of a specific MLVA type. Observed in vitro variability of the Mpn1 VNTR locus prompted further analysis, which showed multiple insertions or deletions of tandem repeats within this locus for a number of specimens and isolates. To our knowledge, this is the first report showing variation within the Mpn1 locus, thus affecting precise and reliable classification using the current MLVA typing system. The superior discriminatory capability of MLVA provides a powerful tool for greater resolution of M. pneumoniae strains and could be useful during outbreaks and epidemiological investigations. Mycoplasma pneumoniae is one of the most common bacterial etiologies of community-acquired pneumonia (CAP), representing 15 to 20% of cases in some studies (28). It is also a significant cause of community-wide outbreaks which are reported to occur in 3-to 7-year intervals with various incidence rates (10,12,28). Upper and lower respiratory tract symptoms are often mild and self-limiting; however, occasional extrapulmonary complications may develop, sometimes resulting in death (5,7,23).Strain subtyping is an important epidemiological tool for surveillance and outbreak investigations. Until recently, molecular typing of M. pneumoniae had been hindered by its minimal and highly uniform genome. Molecular typing was often restricted to a single gene encoding the P1 protein. Methods such as PCR-restriction fragment length polymorphism (RFLP) analysis of the P1 gene (4, 18) and real-time PCR followed by high-resolution melt analysis (HRM) targeting the region of M. pneumoniae repetitive elements 2 and 3 (RepMp2/3) of the P1 gene (19,20) were used to genotype M. pneumoniae into two subtypes (PCR-RFLP) and variants of subtypes 1 and 2 (PCR-HRM). Both sequencing (9) and pyrosequencing (6, 21, 22) techniques have also been developed to differentiate subtypes of M. pneumoniae. Recently, Degrange et al. (6) developed a multiple-locus variable-number tandem-repeat (VNTR) analysis (MLVA) method for M. pneumoniae based on whole-genome analysis that was able to differentiate 26 distinct VNTR types. This procedure takes advantage of the variation in the copy numbers of tandemly repeated sequences within five different loci within the genome and has proven to be highly discriminatory (...

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“…This scheme provides a more discriminatory method than both the MLVA and P1 typing methods currently in use, allow-ing further and more detailed analysis of observed epidemic peaks of M. pneumoniae infection. Community outbreaks of pneumonia caused by M. pneumoniae have been described worldwide (36)(37)(38), and it would be interesting to evaluate this MLST scheme in such epidemic situations. The level of discrimination of this typing method and its usefulness in epidemic analysis should be confirmed by comparing outbreak-related strains to a set of control strains that were isolated from a similar time period and geographical area but that are not epidemiologically related.…”

Section: Discussionmentioning

confidence: 99%

Development of a Multilocus Sequence Typing Scheme for Molecular Typing of Mycoplasma pneumoniae

et al. 2015

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c Mycoplasma pneumoniae is a major human respiratory pathogen causing both upper and lower respiratory disease in humans of all ages, and it can also result in other serious extrapulmonary sequelae. A multilocus sequence typing (MLST) scheme for M. pneumoniae was developed based on the sequences of eight housekeeping genes (ppa, pgm, gyrB, gmk, glyA, atpA, arcC, and adk) and applied to 55 M. pneumoniae clinical isolates and the two type strains M129 and FH. A total of 12 sequence types (STs) resulted for 57 M. pneumoniae isolates tested, with a discriminatory index of 0.21 STs per isolate. The MLST loci used in this scheme were shown to be stable in 10 strains following 10 sequential subculture passages. Phylogenetic analysis of concatenated sequences of the eight loci indicated two distinct genetic clusters that were directly linked to multilocus variable-number tandem repeat analysis (MLVA) type. Genetic MLST clustering was confirmed by genomic sequence analysis, indicating that the MLST scheme developed in this study is representative of the genome. Furthermore, this MLST scheme was shown to be more discriminatory than both MLVA and P1 typing for the M. pneumoniae isolates examined, providing a method for further and more detailed analysis of observed epidemic peaks of M. pneumoniae infection. This scheme is supported by a public Web-based database (http://pubmlst.org/mpneumoniae). Mycoplasma pneumoniae is a common cause of communityacquired pneumonia (CAP) transmitted by aerosol or close contact (1). M. pneumoniae may cause other serious extrapulmonary sequelae, such as encephalitis (2). The pathogen is found in all age groups, with a higher prevalence in children age 5 to 14 years (3, 4). Admissions to a United Kingdom hospital in patients with CAP that were attributed to M. pneumoniae were estimated at 18% in 1982 and 4% in 1999 (5). Major increases and decreases in M. pneumoniae infection have occurred periodically in the United Kingdom; historically, epidemics have occurred at approximately 4-year intervals and have lasted 12 to 15 months, concurrent with sporadic infection at a lower level and seasonal peaks from December to February (4, 6). However, globally, peaks of infection have been observed in either summer or autumn, with no obvious explanation for this seasonal variation (7-10).Typing of clinical isolates by molecular methods is of importance for the understanding of the epidemiology of M. pneumoniae infection and for an analysis of endemic outbreaks. It is generally considered that molecular typing of M. pneumoniae is hampered by the fact that the pathogen is a genetically homologous species (11). Initial molecular typing targeted the gene encoding the major surface protein (P1) of M. pneumoniae. PCRrestriction fragment length polymorphism (PCR-RFLP) analysis of the P1 gene, which encodes a major adhesion, is the most common genotyping method. This enables the separation of isolates into two types, 1 and 2 (11-13). More recent studies utilize the repetitive regions, RepMp2/3 and RepMp4, which ...

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Identification of Risk Factors for Infection in an Outbreak of Mycoplasma pneumoniae Respiratory Tract Disease

Cited by 55 publications

References 31 publications

Development of Multiple-Locus Variable-Number Tandem-Repeat Analysis for Molecular Typing of Mycoplasma pneumoniae

Development of Multiple-Locus Variable-Number Tandem-Repeat Analysis for Molecular Typing of Mycoplasma pneumoniae

Multilocus Variable-Number Tandem-Repeat Analysis of Mycoplasma pneumoniae Clinical Isolates from 1962 to the Present: a Retrospective Study

Development of a Multilocus Sequence Typing Scheme for Molecular Typing of Mycoplasma pneumoniae

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