Pseudomonas aeruginosa diversity in distinct paediatric patient groups.

Pseudomonas aeruginosa is a pathogen that often infects patients who are either immunocompromised or have local defects in host defences. It is known that cystic fibrosis (CF) patients are sometimes infected with certain clonal isolates. It is not clear whether these clonal isolates also infect non-CF patients and whether clonality of isolates occurs in other patient groups. The aim of this study was to investigate P. aeruginosa diversity and the occurrence of clones within five distinct paediatric patient groups susceptible to P. aeruginosa infection. P. aeruginosa isolates were cultured from 157 patients (CF first infection (CF-1 group) (29); CF chronic infection (CF-chronic group) (27); urinary tract infection (34); chronic suppurative otitis media (43); and intensive-care hospitalization/immunodeficiency (24)). All 202 phenotypically different isolates were tested for antimicrobial resistance and further typed by pulsed-field gel electrophoresis. Simpson's diversity index was calculated for the five groups. CF-chronic patients carried the highest number of distinct P. aeruginosa phenotypes and genotypes per culture. Isolates from the CF-chronic group were significantly less diverse than those from the other groups. A group of clonal isolates was observed among patients from the CF-chronic and CF-1 groups. These or different clonal isolates were not encountered among the three other patient groups. No characteristic resistance pattern could be identified among isolates from the distinct patient groups and among the clonal isolates. In conclusion, isolates of the CF-chronic group were less diverse than those in the other patient groups with P. aeruginosa infection; clonal isolates were not encountered in non-CF patients. Transmission of clonal CF isolates to other patient groups was not observed.

Comparison of genotypic and phenotypic methods for species-level identification of clinical isolates of coagulase-negative staphylococci.

To compare commonly used phenotypic methods with genotypic identification methods 47 clinical isolates of coagulase-negative staphylococci (CONS), 10 CONS ATCC strains, and a Staphylococcus aureus clinical isolate were identified using the API Staph ID test, BD Phoenix Automated Microbiology System, and 16S rRNA gene and tuf gene sequencing. When necessary part of the sodA gene was sequenced for definitive identification. The results show that tuf gene sequencing is the best method for identification of CONS, but the API Staph ID test is a reasonably reliable phenotypic alternative. The performance of the BD Phoenix Automated Microbiology System for identification of CONS is poor. The present study also showed that although genotypic methods are clearly superior to phenotypic identifications, a drawback of sequence-based genotypic methods may be a lack of quality of deposited sequences in data banks. In particular, 16S rRNA gene sequencing suffers from the lack of high quality among sequences deposited in GenBank. Furthermore, genotypic identification based on 16S rRNA sequences has limited discriminating power for closely related Staphylococcus species. We propose partial sequencing of the tuf gene as a reliable and reproducible method for identification of CONS species.

Congenital staphylococcal scalded skin syndrome in a premature infant.

UNLABELLED: A case of congenital staphylococcal scalded skin syndrome (SSSS) with fatal outcome in a premature infant is reported. An intrauterine infection with Staphylococcus aureus was probably the cause for the fulminant course of the disease. Despite adequate antibiotic treatment, the infant died within 24 h after birth because of respiratory failure. CONCLUSION: Although rare, infection may occur in the perinatal period and SSSS may present within the first 24 h of life. In this situation, early administering of appropriate antibiotics is essential.

Microbiological factors associated with neonatal necrotizing enterocolitis: protective effect of early antibiotic treatment.

AIM: The incidence of necrotizing enterocolitis (NEC) strongly increased in an neonatal intensive care unit (NICU) in 1997 and 1998 compared with previous years, which coincided with increased incidence of nosocomial sepsis. Specific risk factors related to this NICU and a possible relationship between NEC and nosocomial sepsis were studied retrospectively, including all patients with NEC since 1990 and matched controls. METHODS: Clinical and bacteriological data from the period before the development of NEC and a similar period for the controls were collected retrospectively and corrected for birthweight and gestational age. Statistical analysis was performed by a stepwise regression model. RESULTS: Data of 104 neonates with NEC and matched controls were analysed. The median day of onset of NEC was 12 d (range 1-63 d). Significant risk factors for NEC were: insertion of a peripheral artery catheter [odds ratio (OR) 2.3, 95% confidence interval (95% CI) 1.3-3.9] and a central venous catheter (OR 5.6, 95% CI 3.1-10.1), colonization with Klebsiella sp. (OR 3.4, 95% CI 1.5-7.5) and Escherichia coli (OR 2.1, 95% CI 1.0-4.5), and the occurrence of sepsis, in particular due to coagulase-negative staphylococci (OR 2.6, 95% CI 1.4-5.1). The risk for NEC was decreased after the early use (< 48 h after birth) of amoxicillin-clavulanate and gentamicin (OR 0.3, 95% CI 0.2-0.6). CONCLUSION: Insertion of central venous and peripheral arterial catheters is positively associated with NEC, as is colonization with the Gram-negative bacilli Klebsiella and E. coli and the occurrence of sepsis, particularly due to coagulase-negative staphylococci. Early treatment with amoxicillin-clavulanate and gentamicin is negatively associated with NEC and may be protective against NEC.

Diagnosis of enterovirus infection in the first 2 months of life by real-time polymerase chain reaction.

During summer and fall, enterovirus infections are responsible for a considerable proportion of hospitalizations of young infants. We prospectively studied the incidence of enterovirus infections via real-time polymerase chain reaction (PCR) in blood, feces, and cerebrospinal fluid samples from infants

[Invasive infection with Moraxella catarrhalis in two children with lymphatic leukemia and granulocytopenia]. / Invasieve infectie met Moraxella catarrhalis bij twee kinderen met lymfatische leukemie en granulocytopenie.

In two young children with leukaemia, a girl and a boy aged 5 and 4 years, respectively, an invasive infection due to Moraxella catarrhalis was diagnosed at the time of granulocytopenia. They were treated with antibiotics. The first child developed pneumonia and recovered, the other developed severe septic shock and died. M. catarrhalis is a Gram-negative diplococcus, frequently colonising the upper respiratory tract in young children. In childhood this pathogen mainly causes infections such as otitis media and sinusitis, while in adults it primarily causes laryngitis, bronchitis and pneumonia. Immunocompromised patients or patients with chronic cardiopulmonary disease have an increased risk of severe infections.

[Fatal pneumonia in an adolescent due to community-acquired methicillin-resistant Staphylococcus aureus positive for Panton-Valentine-leukocidin]. / Fatale pneumonie bij een adolescent door thuis opgelopen meticillineresistente Staphylococcus aureus positief voor Panton-Valentine-leukocidine.

A 15-year-old girl developed a severe Staphylococcus aureus pneumonia following an influenza virus infection. The patient was admitted to a paediatric intensive-care facility because of respiratory and circulatory failure. Despite aggressive therapy, she died on the third day following admission to the intensive care unit due to secondary hypoxic-ischaemic encephalopathy. Blood and respiratory aspirate cultures showed community-acquired methicillin-resistant S. aureus (CA-MRSA) with a normal antibiotic sensitivity except for betalactam antibiotics. PCR-based methods demonstrated that the isolate possessed the Panton-Valentine-leukocidin (PVL) gene, encoding an S. aureus exotoxin that is associated with fulminant necrotising pneumonia. This case shows that clinicians in the Netherlands should also be aware of the possibility of CA-MRSA in patients without risk factors for MRSA carriage. Especially in children and adolescents with an influenza virus infection, pneumonia due to PVL-positive S. aureus strains may be life-threatening.

Pneumococcal immune adherence to human erythrocytes.

BACKGROUND: Human red blood cells bind various C3b-coated microorganisms via their C3b/CR1 receptor, a phenomenon referred to as immune adherence. The aim of the present study was to measure pneumococcal adherence to human red blood cells by flow cytometry and to study kinetic aspects of this binding. MATERIAL AND METHODS: We quantified pneumococcal adherence to human erythrocytes by FACS analysis and tested the involvement of antibodies and complement activation in this process. RESULTS: Pneumococci are able to bind to human red blood cells in the presence of human serum. Coating with C3b/C4b appeared obligatory for pneumococcal adherence to red blood cells. The ligand on erythrocytes was confirmed to be complement receptor 1. Kinetic studies showed that innate (mannose-binding lectin) and specific immune factors (IgG antibodies) contributed to the binding of C3b-coated pneumococci to human erythrocytes. After initial binding, serum-derived factor I was found to induce bacterial detachment from the erythrocyte. CONCLUSIONS: Pneumococci are able to adhere to red blood cells. Both the classical and lectin complement pathways are important for optimal C3b-coating of pneumococci for immune adherence. Bound pneumococci are detached from red blood cells by factor I. These findings are in line with the hypothesis of immune adherence in which human erythrocytes are able to bind pneumococci and target the bacteria to the reticulo-endothelial system in the spleen.

Osseointegration of hydroxyapatite-coated and noncoated Ti6Al4V implants in the presence of local infection: a comparative histomorphometrical study in rabbits.

A study was designed to investigate the osseointegration of titanium implants, either noncoated or coated with hydroxyapatite (HA), into rabbit tibiae in the presence of local infection compared with osseointegration in the absence of local infection. HA-coated or noncoated Ti cylinders were implanted into both tibiae of 32 rabbits (New Zealand Whites). Before implantation the left tibia was contaminated with different quantities of Staphylococcus aureus (10(2)-10(5) CFU). Four weeks after surgery the tibiae were explanted and prepared for microbiological and histomorphometrical examination. Histomorphometrical data, as a representation of implant fixation, were obtained by measuring the percentage of bone around the implants (within a radius of 1 mm from the outer diameter of the implants) and the percentage of the circumference of the implant that was in direct contact with bone. Histomorphometry revealed, in particular for the HA implants, a relationship between the inoculum concentration and/or the presence or absence of infection with the bone contact at the distal implant side. This confirms a relationship between peri-implant infection and bone contact or remodeling. HA-coated implants developed, in the presence of bacteria, more easily a more severe infection than noncoated Ti implants, and we show in the present study that local infection will influence histomorphometrical parameters (bone-implant contact) that determine implant fixation. Precautions to prevent contamination (asepsis) and/or infection (perioperative antibiotics) are even more important for the highly biocompatible HA-coated implant.

Epidemiological survey of neonatal non-polio enterovirus infection in the Netherlands.

The epidemiological, virological, and clinical data of 119 infants less than 30 days of age with enteroviral infection collected from January 1993 to November 1995 by the diagnostic virology laboratories were analyzed retrospectively. Ninety-eight isolates (83%) were obtained in the period of May 1 to December 1 with a peak in the summer months. Sixty-five percent (n = 78) of neonates became ill within the first 2 weeks of life. Echoviruses and Coxsackie virus type B were isolated most frequently, in 77 (65%) and 29 (24%) infants, respectively. Diagnosis was made by viral isolation from stool, nasopharyngeal swab, cerebrospinal fluid, and blood. One hundred four (87%) infants developed fever and 25 (21%) infants had diarrhea. A clinical diagnosis of sepsis was made in 42 (35%) infants and meningitis was diagnosed in 28 (24%) cases. The great majority of sepsis cases (36/86%) occurred in infants less than 15 days of age. In conclusion, non-polio enteroviruses (especially echoviruses) are a common and underreported cause of neonatal infection in the Netherlands in the summer months and are associated with a clinical diagnosis of sepsis or meningitis cases in the first 2 weeks of life in a high proportion of cases.

Tobramycin-containing bone cement and systemic cefazolin in a one-stage revision. Treatment of infection in a rabbit model.

The efficacy of tobramycin-containing bone cement with that of systemic cefazolin for treatment of infection in a one-stage revision model is compared. In addition, the value of detecting bacterial DNA after antibiotic treatment was investigated. An implant was inserted into the right tibia of rabbits after inoculation with Staphylococcus aureus. At 28 days, the implant was removed. Subsequently, either plain bone cement with or without systemic administration of cefazolin, or tobramycin-containing bone cement was injected into the medullary canal. The tibiae were cultured 14 days after revision (Day 42), and showed a significant decrease in bacterial counts for both antibiotic groups compared with the control group (p

Carriage of gram-negative bacilli in young Brazilian children with community-acquired pneumonia.

BACKGROUND: Gram-negative bacilli are not infrequently encountered as etiologic organisms of pneumonia in children in warm-climate countries. OBJECTIVES: To investigate the nasopharyngeal carriage rate and antimicrobial susceptibility patterns of gram-negative bacilli colonizing children with community-acquired pneumonia in Fortaleza, Brazil. METHODS: A single nasopharyngeal specimen was collected from children 2 months to 5 years of age presenting at one of the three children's hospitals in Fortaleza and fulfilling the World Health Organization criteria for pneumonia. Randomly recruited healthy children from public daycare centers and immunization clinics served as controls. RESULTS: The study included 912 children, 482 (53%) with pneumonia and 430 (47%) controls. Aerobic gram-negative bacilli were seen in 79 (16%) of the 482 children with pneumonia and 51 (12%) of the 430 healthy controls. Nonfermentative gram-negative bacilli were seen in 85 (18%) of children with pneumonia and 54 (13%) of healthy controls. Neither gender, nutritional status, season, previous hospital admission nor antibiotic use was associated with carriage with gram-negative bacilli. However, pneumonia was associated with increased carriage, whereas concomitant colonization with Streptococcus pneumoniae or Haemophilus influenzae was associated with decreased carriage with gram-negative bacilli. Only 36% of all Escherichia species and 76% of all Klebsiella isolates were susceptible to cotrimoxazole; 90% of all Acinetobacter species were susceptible to gentamicin. CONCLUSION: Nasopharyngeal carriage with gram-negative bacilli, in particular with Acinetobacter species, is common and associated with a clinical diagnosis of community-acquired pneumonia in children in Fortaleza, Brazil.

Once-daily versus multiple-daily gentamicin in infants and children.

In this prospective randomized trial, the efficacy and safety of once-daily administration of gentamicin were compared with multiple-daily administration in infants and children. In addition, pharmacokinetic variables were calculated. Gentamicin therapy was started at a dose of 5 mg/kg per day under individual dose or dosage interval adjustments to achieve target levels. Fifty-two infants and children aged 1 month (postterm) to 16 years were enrolled. The duration of fever from the start of therapy, the percentage decline of C-reactive protein (CRP) on day 3 of treatment, and the clinical outcome were used as efficacy parameters. Nephrotoxicity was evaluated using creatinine serum levels. Basic characteristics in both groups were comparable. A good clinical response was observed in both groups. Fever may have resolved faster with multiple-daily administration, but this was not statistically significant. The percentage of decline of CRP was also comparable in both groups. Nephrotoxicity occurred in six patients, three per group. Many patients were too ill or too young to perform hearing tests, but no clinical signs of ototoxicity were observed. Mean doses of 6.8 mg/kg per day (multiple-daily administration) and 7.3 mg/kg per day (once-daily administration) were necessary to meet the target gentamicin levels. Triple-daily doses had to be reduced to a twice-daily regimen in 17 of 26 children. Dose and dosage interval adaptations can be performed by Bayesian forecasting using a one-compartment model with one set of K(e) and V(d) parameters. The authors consider both regimens equally effective, with a comparable incidence of nephrotoxicity. A starting dose of 6.5 mg/kg once daily is advised.

Prevalence of molecular types and mecA gene carriage of coagulase-negative Staphylococci in a neonatal intensive care unit: relation to nosocomial septicemia.

Molecular typing of isolates revealed that neonatal coagulase-negative staphylococcal (CONS) septicemia is most frequently caused by predominant, antibiotic-resistant CONS types, which are widely distributed among both neonates and staff of the neonatal unit, suggesting cross-contamination. Therefore, infection control measures may be valuable in the prevention of this common nosocomial septicemia.

In-vivo transfer of mecA DNA to Staphylococcus aureus [corrected].

Staphylococcus aureus is thought to have acquired mecA DNA by horizontal transfer. DNA fingerprints made by restriction nucleases that cut certain sequences of DNA are used to compare complete genomes or particular genes between bacteria. We isolated an epidemic mecA(-) meticillin-susceptible S aureus genotype and, subsequently, a rare isogeneic mecA(+) meticillin-resistant S aureus (MRSA) genotype from a neonate who had never been in contact with MRSA. This MRSA contained mecA DNA that was identical to that in a coagulase-negative staphylococcal strain isolated from this patient, but different from other MRSA genotypes. We believe that this MRSA was formed in vivo by horizontal transfer of the mecA DNA between two staphylococcal species.

Neonatal sepsis by Campylobacter jejuni: genetically proven transmission from a household puppy.

We report a case of neonatal Campylobacter jejuni sepsis in a 3-week-old infant who acquired the infection through transmission from a recently acquired household puppy. Genotyping of Campylobacter strains obtained from puppy and child resulted in highly homogeneous findings. This represents the first genetically proven C. jejuni dog-human transmission.

Oral antibiotics as a novel therapy for arthritis: evidence for a beneficial effect of intestinal Escherichia coli.

OBJECTIVE: The intestinal flora is thought to play an important role in regulation of immune responses. We investigated the effects of changing the intestinal flora on the course of adjuvant-induced arthritis (AIA) and on experimental autoimmune encephalomyelitis (EAE) by the use of oral antibiotics. METHODS: Oral treatment with either vancomycin or vancomycin, tobramycin, and colistin was started after AIA and EAE induction. Clinical symptoms of AIA and EAE were monitored, and microbial analysis of ileal samples was performed. RESULTS: Oral vancomycin treatment after disease induction significantly decreased clinical symptoms of AIA. Simultaneously, increased concentrations of Escherichia coli were detected in the distal ileum of vancomycin-treated rats. Ileal concentrations of E coli were inversely related to disease scores in rats with AIA. Coadministration of colistin/tobramycin to prevent the increase in E coli abrogated the beneficial effect of vancomycin on AIA. Vancomycin treatment also reduced the clinical symptoms of EAE. CONCLUSION: We propose oral vancomycin as a novel therapeutic strategy in autoimmune diseases.

Molecular epidemiology of penicillin-resistant Streptococcus pneumoniae colonizing children with community-acquired pneumonia and children attending day-care centres in Fortaleza, Brazil.

To study clonal diversity of penicillin-resistant Streptococcus pneumoniae, 161 randomly selected isolates with reduced susceptibility to penicillin, collected from the nasopharynx of children under 5 years of age with community-acquired pneumonia and healthy controls from public day-care and immunization centres in Fortaleza, Brazil, were characterized by microbiological and serological techniques and automated ribotyping. Also included were 44 randomly selected penicillin-susceptible strains and three international reference strains. With automated ribotyping 75 ribopatterns were observed: 50 ribogroups were unique and 25 ribogroups were represented by two or more isolates. Genetic diversity was extensive but some degree of genetic homogeneity was found in strains from children with pneumonia, strains from children in day-care centres, isolates with reduced susceptibility to penicillin and isolates expressing 'paediatric' serogroups. Fourteen (56%) clusters contained both isolates with reduced penicillin susceptibility and penicillin-susceptible isolates, suggesting emergence of penicillin resistance. In general, there was a good correlation between ribogroups and serogroups, but 12 (48%) clusters contained isolates with alternative serogroups. Isolates with such alternative serogroups were more often encountered in penicillin-susceptible strains (41%) than in strains with reduced susceptibility to penicillin (7%). Thirty-eight (19%) isolates (including seven penicillin-susceptible strains) showed ribotypes indistinguishable from those of two international epidemic clones of S. pneumoniae: ribogroup 54-S-1 (15 isolates) with a ribopattern characteristic of the 23F multiresistant 'Spanish/USA' clone and ribogroup 74-S-3 (23 isolates) with a pattern similar to that of the 6B multiresistant 'Spanish' clone.

Prevention of infection with tobramycin-containing bone cement or systemic cefazolin in an animal model.

We investigated in an animal model the efficacy of tobramycin-containing bone cement and systemic cefazolin for infection prophylaxis. In 18 female rabbits, the femoral cavity was inoculated with Staphylococcus aureus before injection of bone cement. The first group of six rabbits received tobramycin-containing Simplex-P bone cement. Two other groups of six rabbits received plain Simplex-P bone cement. Preoperatively, in one of the two latter groups cefazolin was administered intravenously. The other group served as untreated controls. The rabbits were monitored for clinical signs of infection. At 7 days' follow-up, the femora were harvested and cultures from the bone adjacent to the cement plug were quantified. Cultures from the rabbits which received antibiotic prophylaxis (either cefazolin systemically or tobramycin-containing bone cement) were all negative. In contrast, all rabbits in the untreated control group had positive cultures. These rabbits also had other signs of infection such as an elevated erythrocyte sedimentation rate and loss of body weight. Culture results were confirmed by the absence of bacterial DNA in the polymerase chain reaction hybridization assay. In conclusion, we found that both tobramycin-containing bone cement and systemic cefazolin are effective in preventing implant bed infection in rabbits up to 7 days after contamination with S. aureus.