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Coagulase-negative staphylococci septicaemia in newborns: aspects on host-bacterial interactions with special regard to neutrophil and endothelial response
Linköping University, Department of Molecular and Clinical Medicine, Pediatrics. Linköping University, Faculty of Health Sciences.
2004 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Newborn infants, especially those born preterm, are immunologically immature and prone to invasive infections. As a result of the increasing survival of very preterm (VPT < 31 weeks gestational age) newborns, nosocomial septicaemia has become a major concern the neonatal intensive care, and coagulase-negative staphylococci (CoNS) are nowadays the most frequently isolated pathogens in neonatal blood cultures. Further insight into host-bacterial interactions is required for the development of preventive strategies against CoNS septicaemia in VPT newborns.

Aim of the study: To investigate host-bacterial interactions in neonatal CoNS septicaemia with special regard to the neutrophil and endothelial response and to bacterial virulence factors.

Methods and results: Neonatal blood isolates of CoNS collected at Örebro University Hospital, Örebro, Sweden during the years 1983-1997 were characterised clinically and according to species and to phenotypic and genotypic patterns. Biochemial fingerprinting was found useful as a screening tool for selection of phenotypically related strains, but for further discrimination within a phenotypic cluster, genetic fingerprinting by pulsed field gel electrophoresis (PFGE) was required.

The isolates of S. epidermidis collected during the later part of the study period (1990-1997, n = 50) were further investigated. A hypervirulent clone of bacteria was identified, representing 7 of the 12 sepsis isolates in that cohort. These 12 isolates of S. epidermidis induced significantly higher endothelial release of neutrophil chemoattractants from human umbilical vein endothelial cell (HUVEC) cultures than did the isolates regarded as skin contaminants (n = 38). There were no differences between the sepsis and contaminant groups in the prevalence of genes for biofilm production, methicillin resistance or fibrinogen-binding protein.

The neutrophil oxidative burst occuring after stimulation by different bacterial strains was investigated by a flow cytometric method applied to a whole blood model. The oxidative activity in unstimulated neutrophils was similar in term (n = 10) and preterm (n = 10) newborns. However, the term newborns showed a significantly higher capacity to up-regulate the oxidative burst after bacterial stimulation. Significant differences in oxidative responses to different bacterial strains were observed, but these differences could not be related exclusively to species or invasive capacity.

A neutrophil granule protein, human neutrophil lipocalin (HNL), was evaluated as an early marker of neonatal septicaemia in newborns with clinical signs of infection. The serum level of HNL was significantly higher in the infected group of neonates (n = 25) than in the group with non-proven infection (n = 62). In healthy term controls the HNL level was similar at age 3 days to that at birth and close to the level reported in healthy adults.

Conclusions: The increased up-regulation of endothelial inflammatory mediators induced by sepsis isolates of S. epidermidis represents an important step in the pathogenesis of neonatal CoNS septicaemia. HNL might be useful as a marker of neutrophil activity also in VPT newborns. The laboratory assays used in the present study can be further developed for future investigations of the pathogenesis and host-bacterial interactions in neonatal CoNS septicaemia.

Place, publisher, year, edition, pages
Linköping: Linköpings universitet , 2004. , 70 p.
Series
Linköping University Medical Dissertations, ISSN 0345-0082 ; 861
National Category
Medical and Health Sciences
Identifiers
URN: urn:nbn:se:liu:diva-22100Local ID: 1194ISBN: 91-7373-836-0 (print)OAI: oai:DiVA.org:liu-22100DiVA: diva2:242413
Public defence
2004-10-15, B-husets aula, Universitetssjukhuset, Örebro, 09:00 (Swedish)
Opponent
Available from: 2009-10-07 Created: 2009-10-07 Last updated: 2012-11-01Bibliographically approved
List of papers
1. Phenotypic and genotypic characterisation of blood isolates of coagulase-negative staphylococci in the newborn
Open this publication in new window or tab >>Phenotypic and genotypic characterisation of blood isolates of coagulase-negative staphylococci in the newborn
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2002 (English)In: Acta Pathologica, Microbiologica et Immunologica Scandinavica (APMIS), ISSN 0903-4641, E-ISSN 1600-0463, Vol. 110, no 4, 332-339 p.Article in journal (Refereed) Published
Abstract [en]

Coagulase-negative staphylococci (CNS) are the leading cause of late-onset sepsis in newborns (>72 h of age). Our aim was to determine whether phenotypic and/or genotypic differences existed between blood isolates of CNS regarded as inducers of sepsis or as contaminants. Ninety-seven bloodisolates of CNS recovered from newborns at the neonatal intensive care unit, Örebro, Sweden in 1983–1997 were analysed. Twenty-nine of them (30%) were classified as sepsis isolates and 68 (70%) as contaminants. The most prevalent species was Staphylococcus epidermidis (n=59). Staphylococcus haemolyticus (n=16) was most often isolated from newborns with the lowest gestational age and birth weight. Biochemical typing using the Phene Plate system (PhP) and genotyping using pulsed-field gel electrophoresis (PFGE) showed that the S. epidermidis isolates regarded as inducers of sepsis (n=16) were more homogeneous than isolates considered contaminants (n=37). One main genotypic group, representing seven (44%) isolates, was identified among the sepsis isolates. Phenotypically the S. epidermidis sepsis isolates comprised three major clusters. In contrast, among the S. epidermidis contaminants, eight genotypic groups and two phenotypic clusters were identified. The dominating genotypic group among the sepsis isolates of S. epidermidis may represent strains with higher invasive capacity.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-85065 (URN)10.1034/j.1600-0463.2002.100408.x (DOI)
Available from: 2012-11-01 Created: 2012-11-01 Last updated: 2017-12-07
2. Human neutrophil lipocalin: normal levels and use as a marker for invasive infection in the newborn
Open this publication in new window or tab >>Human neutrophil lipocalin: normal levels and use as a marker for invasive infection in the newborn
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2004 (English)In: Acta Paediatrica, ISSN 0803-5253, E-ISSN 1651-2227, Vol. 93, no 4, 534-539 p.Article in journal (Refereed) Published
Abstract [en]

Aim: To evaluate human neutrophil lipocalin (HNL) as a marker of neonatal invasive infection and determine the normal serum levels of HNL in newborns.

Methods: HNL is released from neutrophil granulocytes and is regarded as a specific marker of neutrophil activity. In 81 newborns 28 d of age with signs of infection on a total of 87 occasions, HNL and C-reactive protein (CRP) were measured at inclusion and on the three following days. As controls, term healthy newborns were recruited at birth (cord blood, n= 45) and at ages 3-5 d (n= 46). Serum HNL was measured by a radioimmunoassay.

Results: 25/87 episodes were classified as infection and 62 as non-proven infection. HNLmax was significantly higher in the infected group (mean 587.6 μg/1) than in the non-proven infected group (mean 217.7 μg/1, p > 0.001). HNL peaked at inclusion, 1 d earlier than CRP. In the healthy controls, HNL was the same at 3-5 d of age as at birth (mean 82.4-81.7 μg/1) and similar to normal adult levels.

Conclusions: The release of HNL is not increased in healthy newborns at birth, but neonatal neutrophils rapidly release HNL upon microbial stimulation in vivo. HNL might be useful as an early marker of neonatal infection.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-85066 (URN)10.1080/08035250410024754 (DOI)
Available from: 2012-11-01 Created: 2012-11-01 Last updated: 2017-12-07
3. Defective neutrophil oxidative burst in preterm newborns on exposure to coagulase-negative staphylococci
Open this publication in new window or tab >>Defective neutrophil oxidative burst in preterm newborns on exposure to coagulase-negative staphylococci
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2004 (English)In: Pediatric Research, ISSN 0031-3998, E-ISSN 1530-0447, Vol. 55, no 6, 966-971 p.Article in journal (Refereed) Published
Abstract [en]

The neutrophil oxidative burst is a product of the regulated assembly of the multicomponent oxidase enzyme. Our aim was to compare the oxidative burst in term (n = 10) and preterm newborns <31 wk gestational age (n = 10) after stimulation with coagulase-negative staphylococci in vitro. Strains of Streptococcus epidermidis with different invasive and slime-producing properties, one strain of S. haemolyticus, and one strain of group B-streptococcus were investigated. A whole-blood flow cytometric assay using the oxidation of hydroethidine to ethidium bromide was used. The oxidative activity in unstimulated neutrophil granulocytes [polymorphonuclear leukocytes (PMNLs)] was similar in term and preterm newborns, but the preterm newborns showed a significantly lower capacity to up-regulate the oxidative burst intensity after bacterial stimulation (p = 0.004). In the term but not in the preterm group, the oxidative burst intensity after bacterial stimulation correlated with the baseline oxidative burst intensity. After bacterial stimulation, there was a trend toward a greater percentage of activated neutrophils in the term group than in the preterm group, but the difference was less pronounced than that in oxidative burst intensity. Significant differences in oxidative burst response to different bacterial strains were observed (p < 0.001), but the differences could not be correlated exclusively to invasive capacity or slime-producing properties. It is concluded that the baseline oxidative activity is similar in term and preterm PMNLs but that preterm PMNLs have a decreased capacity to increase the oxidative burst in response to bacterial stimulation.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-85067 (URN)10.1203/01.pdr.0000127018.44938.89 (DOI)
Available from: 2012-11-01 Created: 2012-11-01 Last updated: 2017-12-07
4. Increased endothelial activation in neonatal sepsis isolates of Staphylococcus epidermidis, but no differences in biofilm producing properties between sepsis and contaminant isolates
Open this publication in new window or tab >>Increased endothelial activation in neonatal sepsis isolates of Staphylococcus epidermidis, but no differences in biofilm producing properties between sepsis and contaminant isolates
(English)Manuscript (preprint) (Other academic)
Abstract [en]

Staphylococcus epidermidis is the predominating etiological agent in neonatal septicemia today, but specific specific factors associated with S. epidermidis are incompletely described. We compared neonatal blood isolates of S. epidermidis representing sepsis (n = 12) or skin contaminants (n = 38) regarding endothelial activation, and the prevalence of genes encoding for biofilm production (icaAB and D), fibrinogen-binding protein (fbe) and methicillin resistance (mecA).

Endothelial cells cultured from human umbilical veins (HUVEC) were challenged by the different isolates of S. epidermidis. Endothelial release of adhesion molecules and interleukin-8 (IL-8) was investigated by an ELISA. Endothelial cell death was determined by light microscopy. The different genes were detected by PCR, and phenotypic biofilm production was investigated by Trypan blue staining. The sepsis isolates of S. epidermidis induced significantly higher endothelial release of intracellular adhesion molecule 1 (ICAM-1, p = 0.0021), endothelial selectin (E-selectin, p = 0.002), and IL-8 (p = 0.010) compared to the contaminants. Vaseular cell adhesion molecules 1 (VCAM-1) was not released. The sepsis-isolates were more cytotoxic than the contaminants; Nine out of 12 sepsis strains induced ≥ 50% cytotoxicity to HUVEC, compared to 15/38 contaminant strains (p = 0.047). The prevalence of the ica-operon, biofilm-production, fbe-, or mecA genes did not discriminate between sepsis and contaminant isolates. It is concluded that sepsis isolates of S. epidermidis induced higher endothelial release of chemotactic inflammatory mediators compared to contaminant isolates, but that the production of biofilm might be less important in neonatal infections eaused by S. epidermidis.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-85068 (URN)
Available from: 2012-11-01 Created: 2012-11-01 Last updated: 2012-11-01

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