Ana Baće

Predominant type-2 response in infants with respiratory syncytial virus (RSV) infection demonstrated by cytokine flow cytometry

Acute RSV infection in infancy may produce some asthma‐like symptoms and may be followed by a recurrent wheeze later in childhood. It has been proposed that RSV infection stimulates type‐2 cytokine responses, resembling those found in atopy and asthma. Peripheral blood cells were obtained from RSV‐infected infants (n = 30) and healthy controls (n = 10). After in vitro restimulation of the cells, intracellular IL‐4 and interferon‐gamma (IFN‐γ) were measured by flow cytometry. The cells from RSV‐infected infants produced more IL‐4 and less IFN‐γ than those from healthy controls. IL‐4 production was more frequent in CD8 than in CD4 cells, and the bias toward IL‐4 production was greatest in infants with mild infections, whereas IFN‐γ production increased with disease severity. Our conclusions are that RSV infection is associated with IL‐4 production in peripheral T cells, and that peripheral blood in infants with severe disease may be depleted of cytokine‐producing cells.

Increased Toll-like receptor 4 expression in infants with respiratory syncytial virus bronchiolitis

The fusion protein of the respiratory syncytial virus (RSV) binds to the pattern recognition receptors, TLR4 and CD14, and initiates innate immunity response to the virus. The aim of the study was to investigate the expression of TLR4 on peripheral blood lymphocytes and monocytes in peripheral blood of infants in both acute and convalescent phase of RSV bronchiolitis (n = 26). In addition, TNF‐α expression in lipopolysaccharide‐stimulated monocytes was also assessed. The results showed TLR4 to be expressed predominantly by monocytes in both sick infants and controls. During the acute phase of infection monocytes up‐regulated TLR4 in eight infants, which returned to the levels recorded in controls 4–6 weeks from infection. There was no difference in the percentage of TNF‐α secreting monocytes. Of the clinical parameters tested, minimal oxygen saturation was found to correlate negatively with this expression in the group of infants with increased TLR4. Additional studies are under way to correlate this finding with the outcome of the immune response to RSV.

Does the viral subtype influence the biennial cycle of respiratory syncytial virus

BackgroundThe epidemic pattern of respiratory syncytial virus (RSV) is quite different in regions of Europe (biennial epidemics in alternating cycles of approximately 9 and 15 months) than in the Western Hemisphere (annual epidemics). In order to determine if these differences are accounted for by the circulation of different RSV subtypes, we studied the prevalence of RSV subtype A and B strains in Zagreb County from 1 January 2006 to 31 December 2007.ResultsRSV was identified in the nasopharyngeal secretions of 368 inpatients using direct fluorescence assays and/or by virus isolation in cell culture. The subtype of recovered strains was determined by real-time PCR. Of 368 RSV infections identified in children during this interval, subtype A virus caused 94 infections, and subtype B 270. Four patients had a dual RSV infection (subtypes A and B).The period of study was characterized by two epidemic waves of RSV infections-one, smaller, in the spring of 2006 (peaking in March), the second, larger, in December 2006/January 2007 (peaking in January). The predominant subtype in both outbreaks was RSV subtype B. Not until November 2007 did RSV subtype A predominate, while initiating a new outbreak continuing into the following calendar year.ConclusionThough only two calendar years were monitored, we believe that the biennial RSV cycle in Croatia occurs independently of the dominant viral subtype.

Detection of genetic lineages of human metapneumovirus in Croatia during the winter season 2005/2006.

Human metapneumovirus (HMPV) is an important respiratory pathogen, especially among young children. The genetic characteristics of HMPV circulating in Croatia have not been studied so far. The aim of this study was to determine the incidence of HMPV infection in hospitalized children with acute respiratory tract infection (ARTI) in the season 2005/2006 in Croatia, as well as to perform the genotypic analysis of detected HMPV strains. From December 1 to March 31 nasopharyngeal secretions (NPSs) were collected from 402 inpatients up to 5 years of age with ARTI. NPSs were tested by real‐time RT‐PCR assay targeting the nucleoprotein (N) gene of HMPV. HMPV infection was detected in 33 patients (8.2%). To perform the phylogenetic study, partial nucleotide sequences were obtained for HMPV fusion (F) gene of 30 HMPV positive samples. Phylogenetic analysis showed the circulation of two main genetic lineages (A and B), with B lineages being prevalent. It also showed the existence of two sublineages within the group B (B1 and B2) and three subclusters within lineage A (A1, A2a and A2b). Further molecular analysis revealed point mutations in HMPV strains of sublineage B1. J. Med. Virol. 80: 1282–1287, 2008.

Expression of chemokine receptor CX3CR1 in infants with respiratory syncytial virus bronchiolitis

Respiratory syncytial virus (RSV) glycoprotein G mimics fractalkine, a CX3C chemokine, which mediates chemotaxis of leukocytes expressing its receptor, CX3CR1. The aim of this study was to examine the relationship between RSV infection and expression of perforin and IFN‐γ in CX3CR1‐expressing peripheral blood CD8+ T cells. Samples were collected from infants with RSV bronchiolitis, both in the acute and convalescence phase (n = 12), and from their age‐ and sex‐matched healthy controls (n = 15). Perforin expression and IFN‐γ secretion in CX3CR1+ CD8+ T cells were assessed by four‐color flow cytometry. The NF‐κB p50 and p65 subunit levels were also determined as markers of RSV‐induced inflammation. Study results showed perforin and CX3CR1 expression to be significantly lower in the convalescent phase of infected infants than in healthy controls. There was no significant difference in IFN‐γ secretion and NF‐κB binding activity between two time‐points in RSV‐infected infants, or when compared with healthy controls. Infants with prolonged wheezing had lower acute‐phase CX3CR1 levels in peripheral blood. These data indicate existence of an event persisting after acute RSV infection that is able to modulate effector functions of cytotoxic T cells, and also link disease severity with CX3CR1 expression.