Molecular evidence of Ureaplasma urealyticum and Ureaplasma parvum colonization in preterm infants during respiratory distress syndrome

Abstract

Background: Ureaplasma urealyticum and U. parvum have been associated with respiratory diseases in premature newborns, but their role in the pathogenesis of the respiratory distress syndrome (RDS) is unclear. The aim of this study was to detect, using molecular techniques, the role of Mycoplasma spp. and Ureaplasma spp. in respiratory secretion and blood specimens of preterm newborns with or without RDS and to evaluate the prevalence of perinatal U. urealyticum or U. parvum infection. The influence of chemotherapy on the clinical course was also evaluated.

Methods: Tracheal aspirate or nasopharingeal fluid samples from 50 preterm babies with (24) or without RDS (26) were analysed for detection of U. urealyticum and U. parvum by culture identification assay and PCR. Sequencing analysis of amplicons allowed us to verify the specificity of methods. Clarithromycin (10 mg kg-1 twice a day) was administered in ureaplasma-positive patients who presented clinical signs of RDS.

Results: 15/24 neonates with RDS (p < 0.001) and 4/26 without RDS were found PCR-positive for U. urealyticum or U. parvum. Culture identification assay was positive in 5/50 newborns, three of which with RDS. Sequencing analyses confirmed the specificity of these methods. Association of patent ductus arteriosus with ureaplasma colonization was more statistically significant (p = 0.0004) in patients with RDS than in those without RDS.

Conclusion: Colonization of the lower respiratory tract by Ureaplasma spp. and particularly by U. parvum in preterm newborns was related to RDS. The routine use of molecular methods could be useful to screen candidate babies for etiologic therapy.

Figure 1

Agarose gel electrophoresis of urease gene PCR. 20 μL of amplified products for the urease gene were electrophoresed on 0.8% agarose gel in TAE buffer (40 mM tris-acetate, 1 mM EDTA, pH 8). A) Lanes 1–14: PCR results of TA samples of newborns with RDS; lanes 15–16: U. urealyticum and U. parvum PCR positive control; lanes 17 and 24: 100 bp ladder; lanes 18–23 PCR results of TA or NF samples of newborns without RDS. B: PCR negativity for Ureaplasma spp. of respiratory secretions from the control group; lane 13: 100 bp ladder; lanes 19–20: U. urealyticum and U. parvum PCR positive control.

Figure 2

BLAST sequence analysis of Ureaplasma spp. ureasegene amplicons. BLAST sequence analysis of urease gene amplified fragments of U. urealyticum (GenBank AE002140) and U. parvum (GenBank AF085729) in two different respiratory specimens: sample SR1 (A) resulted identical to U. parvum and sample SR42 (B) resulted more identical to U. urealyticum than U. parvum. Nucleotide differences between sequences of two species are shown.

Figure 3

BLAST sequence analysis of Ureaplasma parvum 16SrRNA amplicons. BLAST sequence analysis of 16S rRNA gene amplified fragments of U. parvum (GenBank AF002112) employing U3/P6 primers, in the sample SR1 resulted identical to U. parvum. Nucleotide differences between sequences of two species are shown.

Figure 4

BLAST sequence analysis of Ureaplasma urealyticum 16S rRNA amplicons. BLAST sequence analysis of 16S rRNA gene amplified fragments of U. urealyticum (GenBank AF073455, AF073454) employing U8/P6 primers in the sample SR42 resulted identical to U. urealyticum. Nucleotide differences between sequences of two species are shown.