4. NASOPHARYNGEAL MICROBIOTA COMMUNITY COMPOSITION AND BACTERIAL SUPER-INFECTION DURING RSV INFECTIONS
The interaction between nasopharyngeal microbiota and RSV in children can be bidirectional. As discussed above, specific nasopharyngeal microbiota compositions may modulate RSV severity but, at the same time, RSV-induced infections can stimulate proliferation of potentially pathogenic microorganisms in the upper airways and thus favor bacterial superinfections and morbidity [7,38,39]. RSV infection can promote the invasion of the respiratory epithelium by non-typeable Haemophilus influenzae andStreptococcus pneumoniae through a variety of mechanisms, including upregulation of bacterial adhesion protein expression, loss of epithelial barrier integrity and decrease of bacterial clearance mechanisms [3,8,40,41]. Using an in vitro and a lung mouse model of infection, incubation with RSV or purified G protein was shown to increase Streptococcus pneumoniae adherence to differentiated ciliated cells of the respiratory epithelium, thus enhancing the severity of the airway infection and decreasing animal survival [8,42]. Also detected in these studies were extensive changes in the pneumococcal transcriptome, with upregulation in the expression of key pneumococcal virulence genes, including those encoding the pneumococcal toxin pneumolysin [42]. On the airway epithelium end, RSV infection upregulated the cellular surface expression of other known receptors for Streptococcus pneumoniaeand non-typable Haemophilus influenzae , notably the intercellular adhesion molecule-1 (ICAM-1) and platelet-activating factor receptor [8]. In clinical practice, for infants with RSV bronchiolitis the risk of secondary bacterial infections (SBI) is present, yet the severity of the resulting bacterial infection appears to parallel that of the RSV infection (figure 3A). The low risk of SBI in mild RSV is born out in a study on 1,248 febrile infants < 60 days of age, from 88 pediatric Emergency Departments in New York. The rate of SBI was significantly lower in those with RSV infection (7.0%) than in those without RSV infection (12.5%) [43]. Additionally, as compared to the RSV-negative infants, RSV-positive infants had a lower rate of bacteremia (1.1% vs 2.3) and of urinary tract infections (UTI) (5.4% vs 10.1%). Similar results were reported in a retrospective cohort study of < 8-week-old infants, who presented with fever and a positive RSV antigen test to the Emergency Department at an urban child facility in Nashville, Tennessee [44]. Buttressing these results is a systematic review of 11 studies on “in- and out-patients”, < 60 to 90 day old, with bronchiolitis, reporting no case of bacteremia, yet the rate of UTI resulting slightly higher in the RSV-positive subgroup (5.1% vs 2.0%) [45]. These data suggest that, in nontoxic-appearing infants with a positive RSV test, full septic evaluations are not necessary, however, the finding of a low, but clinically relevant rate of urinary tract infection, probably justifies urine work up [44,45]. Conversely, the incidence of SBI and concomitant bacterial pneumonia is more frequently observed (or diagnosed) in infants with severe RSV bronchiolitis (figure 3B). Out of 127 infants (median age 1.7 months) admitted to the PICU of the Zurich University Children’s Hospital, 57 were mechanically ventilated [46]. Tracheal cultures obtained from intubated children within the first hour of intubation were positive in 25 infants, half of them for community and half for nosocomial acquired bacteria. Microorganisms associated with bacterial LRTI included Moraxella catarrhalis ,Haemophilus influenzae type b and Streptococcus pneumoniae . The incidence of sepsis was 3/127 patient. Despite only documenting 25 bacterial concomitant infection, antibiotics were administered to 73/127 infants over the first two days of hospital admission, namely, to a high proportion of those with negative bacterial cultures. The authors conclude that to avoid inappropriate antibiotic prescription, tracheal aspirates should be investigated microbiologically on a routine basis in this patient group [46]. The high frequency of bacterial coinfection in young infants with severe RSV bronchiolitis was confirmed in the study of the Royal Liverpool Children’s Hospital [47]. Out of the 165 children (median age 1.6 months) admitted to PICU with RSV bronchiolitis, 70 (42.4%) lower airway secretions tested positive for bacteria, includingHaemophilus influenzae , Moraxella catarrhalis ,Streptococcus pneumoniae and Staphylococcus aureus[47]. All those with bacterial co-infection were mechanically ventilated and required ventilatory support for longer than those with only RSV. Gender, co-morbidity, origin, prior antibiotics, time on preceding antibiotics, admission oxygen, and ventilation index were not predictive of positive bacterial cultures [47]. The risk of prolonged hospitalization and PICU admission may be even higher in preterm born infants, hospitalized with RSV disease, if concurrent bacterial infections are present. Out of a total of 464 infants, median age 2.75 months (range 0.25–96), hospitalized due RSV infection to the Graz Medical University, 42 (9.1%) were born<37 weeks of gestational age [48]. Concurrent bacterial infections were detected in 3.1% of term and 9.5% of preterm infants and the percentage of those referred to PICU was 5.2% and 23.8% in term and preterm infants respectively. In all these preterm a diagnosis of pneumonia was made, and tracheal aspirates were positive for Streptococcus pneumoniaeand Haemophilus influenzae . Mean hospital stay was longer (22.3 days) in preterm infants with bacterial co-infection than in the ones without it (10.3 days) [48]. Concomitant bacterial infection can occur also in older children hospitalized with RSV bronchopulmonary infections as shown in a study performed of 188 pediatric inpatients, aged up to 5 years, admitted to 2 hospitals in Chiba Prefecture, Japan [49]. Pathogenic bacteria were isolated from washed sputum bacterial culture in 43.6% of the children, the three most frequently isolated being Haemophilus influenzae (43.9%), Streptococcus pneumoniae (36.6%) and Moraxella catarrhalis (29.3%). Before admission, 28.7% of the patients received oral antibiotics, most often macrolides and amoxicillin. Of the Haemophilus influenzae strain isolated 38.9% were ampicillin-resistant, whilst allStreptococcus pneumoniae strains were penicillin G sensitive, minimum inhibitory concentration values were 2 µg/ml [49]. Thus, RSV bronchopulmonary infection in hospitalized children can often be associated with super-infection by antimicrobial-resistant bacterial. Prior antibiotic treatment and Gram-negative pathogen prevalence at nasopharyngeal level can be associated with more disease severity. A prospective, observational cohort study, of 136 infants with RSV bronchiolitis enrolled at 24 h of hospitalization; 104 on the ward and 32 in the PICU [50]. As compared to an age-matched group of healthy controls, RSV bronchiolitis infants had more frequent nasopharyngeal colonization withnon-typable Haemophilus influenzae or Moraxella catarrhalis (Gram-negative bacteria) andStaphylococcus aureus andStreptococcus pneumoniae (Gram-positive bacteria). Higher proportion of Gram-negative pathogens was associated with higher blood neutrophil percentages, greater number of nasal wash leukocytes, increased concentrations of plasma IL-8 and IL-6, a trend towards increased disease severity and longer need for oxygen. Thus,Staphylococcus aureus , which seems to have a protective role? on RSV infection, may be detected in the lower airways of young infants with severe RSV bronchiolitis but its presence does not seem to be associated with increase disease severity. In this study, 47% (64/136) of all RSV infants and 75% of those in PICU received antibiotics prior to sample collection. This common use of antibiotic was associated with a decreased likelihood of recovering potentially pathogenic bacteria by culture, but also with enhanced disease severity as defined by clinical parameters and radiologic findings. Infants treated with antibiotics prior to hospitalization required oxygen for a longer time and radiographic lobar consolidation was more frequently identified compared to those with no prior antibiotics. Atelectasis or lobar consolidation was associated with higher clinical disease severity scores [50]. The elevated antimicrobial-resistance in bacterial superinfection and the higher risks of enhanced disease severity highlight the need for microbiological investigation of airway secretion in this patient population. Finally, the incidence of Streptococcus pneumoniaecoinfection could be modified following the introduction of seven-valent pneumococcal conjugate vaccine (PCV7). Weinberger DM et al, evaluating hospitalization data from community hospitals in 18 U.S.A. states and comparing the 1992/1993 and 2008/2009 seasons, demonstrated a significant decline in RSV-coded hospitalizations in children aged <1 y and 1–2-year-old following PCV7 introduction in 2000 [51]. We could find no data on the effect of the effect of the subsequent introduction of PCV13.