Thiopurine S-methyltransferase genotype and the use of thiopurines in paediatric inflammatory bowel disease Greek patients.

BACKGROUND AND OBJECTIVE: Azathioprine (AZA) and 6-mercaptopurine (6MP) are used in the treatment of paediatric inflammatory bowel disease (IBD). Genetic variations in thiopurine S-methyltranfarase (TPMT) gene have been correlated with enzyme activity and with the occurrence of adverse events to AZA and 6MP. The aim of the present study was to investigate the frequency of the functional TPMT polymorphisms and their association with the occurrence of adverse events during azathioprine therapy in a paediatric IBD cohort. METHODS: Ninety-seven thiopurine-treated paediatric IBD patients (41.24% boys and 58.76% girls) with a mean age 11.25 years (range 3-16), were assessed for TPMT polymorphisms and adverse events. RESULTS: Of the 97 patients enrolled in the study, 18 (18.56%) were heterozygous mutated; two (2.06%) were homozygous for a mutated TPMT gene. Ten patients (10.31%) developed adverse effects, and four of them (40%) had one of the variant alleles. CONCLUSIONS: In this small cohort of subjects, no association was found between TPMT polymorphisms and the occurrence of thiopurines-related adverse events.

Dietary compliance and life style of children with coeliac disease.

BACKGROUND: Coeliac disease (CD) is common and requires a permanent strict gluten-free diet (GFD). However, data concerning how the situation is experienced by children are limited. The present study aimed to investigate the compliance with a GFD and the impact of CD and GFD on the lifestyle of patients and their families, together with proposed recommendations for improvement of quality of life. METHODS: Children with biopsy confirmed CD were recruited consecutively from the outpatient gastroenterology clinic. Participants were evaluated by a special questionnaire for compliance with the GFD, patients' knowledge about CD, and the well-being and lifestyle of children and their families. Comparisons between discrete variables were performed by a chi-square test. RESULTS: Seventy-three children of median age 9.4 (interquartile range = 5-14.5) years were evaluated. Compliance to diet was reported by 58%. Reasons for noncompliance were: poor palatability (32%), dining outside home (17%), poor availability of products (11%), and asymptomatic disease diagnosed by screening (11%). The acceptance of the GFD was reported as good in 65%, whereas avoidance of travelling and restaurants was stated by 17% and 46% of families, respectively. Most families experienced difficulties detecting gluten from the food label. Proposed factors for improvement of quality of life were: better labelling of gluten-containing ingredients (76%) and more gluten-free (GF) foods in supermarkets (58%) and restaurants (42%). CONCLUSIONS: Children with CD have low compliance with the GFD. Better education about the disease, the availability of GF products, and appropriate food labelling could improve compliance and quality of life.

Delayed exposure to infections and childhood lymphomas: a case-control study.

OBJECTIVES: Delayed exposure to common infections during childhood, have been implied to cause strong immunological response to a single infectious agent that eventually triggers leukemogenesis. The aim of the present study was to investigate whether decreased exposure to infections, as reflected in a more seronegative spectrum to several common infectious agents, is associated with increased risk for the development of childhood lymphomas. METHODS: All 125 children (up to 14 years old), with Hodgkin (HL, n = 52) and non-Hodgkin lymphomas (NHL, n = 73) diagnosed through the national network of childhood Hematology-Oncology units during an 8-year period were enrolled in the study along with 125 age- and gender-matched controls. Past exposure to nine common infections [respiratory syncytial virus (RSV), influenza A and B, parainfluenza type 1, adenovirus, Epstein-Barr virus (EBV), cytomegalovirus (CMV), human herpes virus 6 (HHV6), Bartonella henselae] was assessed using serological markers. RESULTS: After controlling for possible confounding factors, the overall seronegativity status upon diagnosis was statistically significantly associated with NHL [odds ratio; 95% CI: 1.45 (1.10-1.93), p = 0.01] and less so with HL risk [odds ratio; 95% CI: 1.30 (0.83-2.05), p = 0.25]. A statistically significant association of seronegativity with the development of NHL was evident for RSV [odds ratio; 95% CI: 7.27 (1.59-33.28), p = 0.01], EBV [odds ratio; 95% CI: 6.73 (1.45-31.20), p = 0.01] and suggestive association for influenza B [odds ratio; 95% CI: 2.60 (0.90-7.55), p = 0.08] and influenza A [odds ratio; 95% CI: 2.35 (0.81-6.80), p = 0.11]. In contrast, there was no evidence for association of HL with negative serology for any of the infectious agents tested. CONCLUSIONS: The risk of lymphomas, especially NHL, might be higher when, due to lower exposure to several infectious agents, the relatively unmodulated immune system of a child is challenged by environmental stimuli that can trigger development of lymphomas. The results, however, need further confirmation, through more pertinent methodological designs.

Long-term follow-up of children with vesicoureteral reflux with and without antibiotic prophylaxis.

The aim of the present study was to obtain data on the outcome of children with persistent vesicoureteral reflux (VUR) after cessation of antibiotic prophylaxis. Children with VUR who had been on antibiotic prophylaxis for at least 2 y and were free of urinary tract infections (UTI), had normal voiding patterns, and no hydronephrosis or new kidney scarring, had antibiotic prophylaxis discontinued, were followed up prospectively with urine cultures, voiding cystourethrography, and technecium-99m dimercaptosuccinate renal scintigraphy. The findings were compared with those of the same patients while they were on antibiotic prophylaxis. In 54 children (39 girls and 15 boys), antibiotic prophylaxis was discontinued. The mean follow-up time on and off antibiotic prophylaxis was 4.4+/-2.1 and 4.4+/-2.2 y, respectively. Nine UTI episodes occurred during the on- and 8 during the off-prophylaxis period. In 80 of 96 and in 68 of 74 ureters the reflux resolved or downgraded during the on- and off-prophylaxis periods, respectively. No new scar lesions were detected in any of the children. In conclusion, in children with persistent VUR and certain characteristics, antibiotic prophylaxis can be safely discontinued.

Brain abscesses complicating Staphylococcus aureus sepsis in a premature infant.

Brain abscess is a rare complication of staphylococcal bacteremia in infants. Here we present a case of a premature infant who developed multiple brain abscesses 12 weeks following an episode of inadequately treated Staphylococcus aureus sepsis. The abscess developed in the absence of trauma, prior surgery, cyanotic heart disease, or immune defect. The initial staphylococcal isolate exhibited identical pulsed-field gel electrophoresis pattern with that of the isolate cultured from abscess aspirate. The infant was successfully treated by surgical drainage and administration of antibiotics for 12 weeks, initially teicoplanin and meropenem followed by trimethoprim/sulfamethoxazole, without neurological or developmental sequelae. Staphylococcal bacteremia in neonates should be vigorously treated to prevent life-threatening complications.

Carriage of Neisseria meningitidis by Greek children: risk factors and strain characteristics.

Oropharyngeal swabs were cultured from 554 children aged 2-19 years attending nurseries, primary schools and secondary schools in the central Athens area. A questionnaire was completed to identify risk factors for carriage. Susceptibility to antimicrobial agents was determined by Etest. The genetic relatedness of the strains was examined by pulsed-field gel electrophoresis (PFGE), and isolate serogrouping was performed by slide agglutination. Twenty-two (4%) children were carriers of Neisseria meningitidis; seven isolates belonged to serogroup C, and five to serogroup B. One isolate was resistant to co-trimoxazole, and five showed intermediate resistance to penicillin. DNA analysis of 16 isolates revealed six distinct PFGE patterns. Clusters with indistinguishable PFGE patterns were noted in the same school. More than one serogroup was included in the same clonal group. On multivariate logistic regression analysis, only age > 12 years remained independently associated with the carrier state (odds ratio, 7.96; 95% CI, 2.24-28.33; p < 0.001). Overall, the N. meningitidis carriage rate among Greek schoolchildren increased with age, and the predominant serogroups in the Athens region were groups C and B. These findings may have important implications for future immunisation strategies with conjugate vaccines.

Drug-resistant tuberculous mastoiditis in 2 children.

Tuberculosis of the middle ear and mastoid is uncommon nowadays. Two cases of drug-resistant tuberculous mastoiditis in immunocompetent Greek native children are reported and the diagnostic and therapeutic difficulties of this rare condition are discussed.

Gentamicin penetration into cerebrospinal fluid in experimental Haemophilus influenzae meningitis.

We studied the effect of meningitis and the method of parenteral gentamicin administration (intramuscular injection, a 30-min intravenous infusion, or intravenous bolus administration) on achievable concentrations of drug in cerebrospinal fluid (CSF). In normal animals, only intravenous bolus administration of 2 to 8 mg/kg produced a gentamicin concentration of greater than 0.1 microgram/ml in CSF in some animals. All CSF samples contained less than the limit of detection (0.1 microgram/ml) after the intramuscular administration of 6 mg/kg. In animals with meningitis, gentamicin penetration into cisternal CSF was increased significantly after a bolus administration of 6 mg/kg (mean, 0.197 +/- 0.063 microgram/ml in normal animals versus 1.68 +/- 0.38 micrograms/ml in animals with meningitis; P less than 0.01). In meningitic animals that received 6 mg/kg as an intravenous bolus, lumbar CSF had the highest maximum concentration (4.25 +/- 1.08 micrograms/ml), in comparison with ventricular CSF (3.10 +/- 0.66 micrograms/ml). The gentamicin concentration in cisternal CSF decreased more slowly than it did in serum (elimination half-life, 238.70 +/- 64.56 min in cisternal CSF versus 82.73 +/- 2.91 min in serum), yielding a relative increase in the percentage of penetration. We conclude that maximum penetration by gentamicin into CSF occurs after intravenous bolus administration and that the maximum concentration occurs in lumbar CSF.

Molecular epidemiology of plasmid-mediated ampicillin resistance in Haemophilus influenzae type b isolates from Alaska.

Native Alaskans have a high incidence of disease caused by invasive Haemophilus influenzae type b and represent an isolated population for epidemiological study. We used plasmid DNA analysis and subtyping of outer membrane proteins as markers to characterize 29 ampicillin-resistant, invasive strains and seven ampicillin-resistant, noninvasive strains of this organism from distinct geographic regions. All 36 strains produced beta-lactamase; 34 strains transferred resistance by conjugation. Seven of the 36 strains harbored detectable plasmid DNA: four had a molecular mass of 40 MDa, and three had a molecular mass of 3 MDa. Furthermore, 20 transconjugants had a similar large plasmid, and four had a similar small plasmid. Ten of 12 transconjugants with either the large, small, or undetectable plasmid DNA were able to retransfer resistance. Transformation of resistance was successful with two large plasmids. DNA-DNA hybridization studies revealed that 33 of 36 strains had DNA homology. Restriction endonuclease digestion of 10 large plasmids revealed five patterns; identity was evident within a geographic region, and similarity existed between regions. Seven restricted plasmids demonstrated an identical pattern with a small beta-lactamase probe. Ampicillin resistance in these isolates from Alaska is primarily due to a common, 40-MDa conjugative plasmid that mediates beta-lactamase production, a finding which differs from that for ampicillin-resistant plasmids isolated elsewhere in the United States. Despite variable outer membrane protein profiles of the distinct strains of H. influenzae type b, the plasmids shared significant DNA homology. It appears that a common genetic element was responsible for the dissemination of this phenotype for resistance in Alaska.

Enterobacter sepsis in infants and children due to contaminated intravenous fluids.

Sixty-three cases of nosocomial sepsis occurring from April through October 1981, in a 500-bed pediatric hospital, were traced to bacterial contamination of intravenous fluid produced by a single manufacturer. Two species of uncommon blood stream pathogens, Enterobacter cloacae and Enterobacter agglomerans contaminated the fluid. Infections with these organisms might have contributed to the death of four patients; two who were immunosuppressed, one who was asplenic and one premature infant. Epidemiologic and laboratory investigations identified the site of contamination to be within the screw-caps of the bottles containing the intravenous fluid. Contamination occurred during insertion of the intravenous fluid administration set into the bottle. The "epidemic" terminated when the hospital discontinued the use of infusion fluids from that manufacturer. We conclude that intravenous fluids should be examined during outbreaks of nosocomial bacteremia due to unusual pathogens.

Loss of plasmid DNA coding for beta-lactamase during experimental infection with Haemophilus influenzae type b.

Apparent R factor-negative segregation was documented during infection of infant monkeys with two of three strains of ampicillin-resistant Haemophilus influenzae type b. In vitro the bacterial population of one strain (A-Sm) uniformly produced beta-lactamase. All bacteria isolated from blood or cerebrospinal fluid from both animals inoculated with strain A-Sm produced the enzyme. In contrast, 98% and 96% of bacteria from two other strains produced beta-lactamase in vitro. After intranasal inoculation of infant Macacca mulatta with these two strains, bacteria isolated from blood and cerebrospinal fluid uniformly did not produce beta-lactamase. Loss of the beta-lactamase-producing phenotype was associated with loss of plasmid DNA. Strains containing a mixed population of bacteria may undergo spontaneous loss of plasmid DNA during experimental infection. It is suggested that in these strains the bacteria carrying plasmids are less virulent.

In vitro antibacterial activity of fluorinated analogs of chloramphenicol and thiamphenicol.

We evaluated the in vitro antimicrobial activity of Sch 24893, Sch 25298, and Sch 25393, three novel analogs of chloramphenicol and thiamphenicol. All of the analogs had minimal inhibitory concentrations of less than or equal to 10 micrograms/ml for 18 chloramphenicol-thiamphenicol-resistant strains of Shigella dysenteriae and 21 strains of resistant Salmonella typhi. The analogs were also more active than were chloramphenicol and thiamphenicol against chloramphenicol-resistant enteric bacteria, including six strains of Escherichia coli, seven strains of Klebsiella pneumoniae, and two strains of Enterobacter cloacae. Fifty-three strains of ampicillin-resistant Haemophilus influenzae were uniformly susceptible to chloramphenicol, thiamphenicol, and the three analogs. Sch 25298 was the most active compound tested (minimal inhibitory concentration, 0.5 microgram/ml for all strains). Four of seven chloramphenicol-thiamphenicol-resistant Haemophilus strains were susceptible to the fluorinated analogs. Of the three Haemophilus strains which were resistant to chloramphenicol, thiamphenicol, and the analogs, two contained less than 10% of the chloramphenicol acetyltransferase activity of the strains which were resistant to only chloramphenicol and thiamphenicol. We conclude that fluorinated analogs of chloramphenicol and thiamphenicol have considerable in vitro activity against a broad spectrum of chloramphenicol-thiamphenicol-resistant, gram-negative bacteria.

Comparison of two antigen detection techniques in a primate model of Haemophilus influenzae type b infection.

Rapid diagnosis of Haemophilus influenzae type b meningitis is possible using immunological tests for capsular antigen (polyribophosphate, PRP), such as countercurrent immunoelectrophoresis (CIE) and latex particle agglutination (LPA). We compared two tests in monkeys with evolving, serially quantitated H. influenzae type b bacteremia (n = 23) and meningitis (n = 21). In vitro, the LPA test was sensitive to 0.5 ng of PRP/ml of saline, and the CIE test was sensitive to 1.0 ng/ml; in serum, however, CIE detected 5.0 ng of PRP/ml, whereas the sensitivity of LPA was unchanged. LPA detected PRP earlier in the course of bacteremia (mean, 12 h after onset; range, 4 to 36 h) than did CIE (mean, 45 h; range, 4 to 168 h) (P less than 0.01). A positive LPA test required greater than or equal to 100 bacteria per ml of blood, whereas CIE required greater than or equal to 1,000/ml. PRP accumulated with continuing blood stream infection, aiding detection of low-grade bacteremia. LPA detected antigen in cerebrospinal fluid (CSF) earlier in the course of meningitis and at a lower bacteria density than did CIE. Both methods detected antigen reliably with greater than or equal to 1,000 bacteria per ml of CSF. A close correlation existed between CSF concentrations of capsular antigen and bacteria (r = 0.90, P less than 0.001). We conclude that the LPA method permits earlier diagnosis of H. influenzae type b infection in part because of its greater sensitivity.

Effect of inoculum size on the susceptibility of Haemophilus influenzae b to beta-lactam antibiotics.

The current prevalence of ampicillin-resistant Haemophilus influenzae b meningitis requires accurate knowledge of susceptibility to alternative antibiotics. One variable affecting susceptibility is inoculum size. We studied the susceptibility of 200 clinical isolates of H. influenzae b to ampicillin, carbenicillin, and cefamandole at inocula of 10(5) and 10(7) CFU by two techniques. Fifty ampicillin-susceptible and fifty ampicillin-resistant strains were tested for susceptibility to ampicillin by broth dilution while 100 of each were tested by agar dilution. An inoculum effect was found, being greatest with the ampicillin-resistant strains. The range of minimal inhibitory concentrations for the resistant strains was 25 to 800 microgram of ampicillin per ml at an inoculum of 10(5) and 2,000 to less than 6,000 microgram of ampicillin at 10(7); 1.0 to 150 microgram of carbenicillin per ml at 10(5) and 6.2 to 2,000 microgram of carbenicillin per ml at 10(7); 0.4 to 2.0 microgram of cefamandole at 10(5) and 1.0 to 125 microgram/ml at 10(7). Because of this inoculum effect, we would not recommend the use of carbenicillin or cefamandole for therapy of ampicillin-resistant H. influenzae meningitis.

Ventricular involvement in experimental Hemophilus influenzae meningitis.

We determined the frequency of ventricular involvement during Hemophilus influenzae type b meningitis in ten bacteremic infant rhesus monkeys. Meningitis was defined as cerebrospinal fluid obtained from the lumbar subarachnoid space or cisterna magna containing bacteria and ten or more leukocytes per mm3. HIB were cultured from 22 of 22 ventricular CSF samples. In 17 of 18 comparisons of bacterial density in ventricular and cisternal CSF, the values were within one log10: similarly, 5 of 8 quantitative ventricular-lumbar comparisons were within one log10. This concordance was present at bacterial densities of 2 x 10(1) to 1 x 10(9) CFU/ml. When discordance was present, the ventricular CSF contained more bacteria. In 15 of 20 comparisons of leukocyte density in ventricular and cisternal CSF, the ventricular pleocytosis was lower (mean ventricular/cisternal ratio 0.08). We conclude that infection of the lateral cerebral ventricle is a uniform feature of HIB meningitis in infant monkeys, but the cellular inflammatory component is less.

Evaluation of a rapid beta-lactamase test for detecting ampicillin-resistant strains of Hemophilus influenzae type b.

Chloramphenicol is presently the drug of choice in the initial treatment of serious infections due to Hemophilus influenzae type b. Rapid detection of ampicillin resistance in clinical isolates would facilitate early discontinuation of chloramphenicol therapy in patients infected with ampicillin-sensitive bacteria. A total of 160 strains of H. influenzae type b were tested with a one-hour acidimetric microassay for beta-lactamase activity. All ampicillin-resistant strains rapidly hydrolysed the beta-lactam ring of penicillin. When isolates were encoded and tested without knowledge of their MICs, the 40 ampicillin-resistant strains (MIC greater than or equal to 2 mug/ml) were readily distinguished from 120 sensitive strains. Rapid beta-lactamase assay is therefore a reliable detector of ampicillin resistance in H. influenzae type b.