Immune response to a laminin-binding protein (Lmb) in group A streptococcal infection.

BACKGROUND: A laminin-binding protein (Lmb) similar to that of group B streptococcus is conserved in group A streptococcus (GAS) and has a role in adhesion of GAS to epithelial cells. The role of this protein is yet to be clarified in disease process and thus, it is important to know its role in binding of GAS to laminin and the immunogenic response against it in patients related with GAS infection. METHODS: A recombinant protein (rGAS-Lmb) was purified using the lmb gene from M1 GAS and tested for its role in binding of GAS with laminin. The antibody response against rGAS-Lmb in patient sera related with GAS infection was measured by ELISA. RESULTS: The rGAS-Lmb bound with laminin directly and inhibited the binding of GAS to laminin. The antibody response against rGAS-Lmb in patients with uncomplicated streptococcal infection (U. Strep) and those with rheumatic fever (RF) were significantly higher than those in the control group (P < 0.0001 and P < 0.001, respectively). No difference of anti-rGAS-Lmb antibody titer could be found between these two disease groups. CONCLUSION: The higher antibody response in patients with GAS infection implies that the protein is well expressed during the period of infection and may be related with the colonization and infection of GAS in pharyngeal mucosa.

Heart rate variability and ambulatory blood pressure monitoring in young patients with hypertrophic cardiomyopathy.

BACKGROUND: Sudden cardiac death commonly occurs in young patients with hypertrophic cardiomyopathy (HCM); however, their heart rate variability (HRV) and blood pressure (BP) response to daily life activities is not well known. METHODS AND RESULTS: HRV and ambulatory BP monitoring were performed in 20 patients (age range: 7-21 years) and 57 age-matched healthy volunteers (age range: 10-22 years). Time domain variables and spectral data were obtained at hourly intervals throughout the day. To determine the BP response to daily life activities, the ratios of the mean BP and pulse pressure in the morning, afternoon, and night to those during sleeping were calculated. The association between the BP level and HRV was also evaluated. The HCM patients showed significantly increased sympathovagal imbalance and decreased parasympathetic activity in the early morning, around noon, and in the early evening. This abnormality was independent of cardiac symptoms. Symptomatic patients showed a significantly lower systolic BP response in the morning, and a higher incidence of dissociation between sympathetic activity and BP response than asymptomatic patients. CONCLUSION: An abnormal BP response in the presence of impaired HRV appears to be predictive for cardiac events in young patients with HCM.

Effects of pure 2,4-dichlorophenoxyacetic acid on cultured rat embryos.

2,4-dichlorophenoxyacetic acid (2,4-D), a plant growth regulator, has been used worldwide as a herbicide. Previously we evaluated the prenatal developmental effects of 2,4-D by feeding it to pregnant rats and found that it is maternally toxic and embryolethal, and it induces urogenital malformations in rat fetuses. In the study presented here, we investigated the effects of pure 2,4-D on rat embryos in whole embryo culture. Rat embryos on day 9.5 of gestation were cultured for 48 h at several concentration levels with pure 2,4-D (50-500 microg/mL). 2,4-D caused a concentration-related increase in the incidence of each malformation. Significant decreases in the number of somites were observed at a concentration of 100 microg/mL or more. At the concentration of 100 microg/mL, there was normal yolk sac circulation. This result suggests that 2,4-D has a detrimental effect on somite development and directly damages developing embryos.

Typical enteroaggregative Escherichia coli is the most prevalent pathotype among E. coli strains causing diarrhea in Mongolian children.

Diarrhea remains one of the main sources of morbidity and mortality in the world, and a large proportion is caused by diarrheagenic Escherichia coli. In Mongolia, the epidemiology of diarrheagenic E. coli has not been well studied. A total of 238 E. coli strains from children with sporadic diarrhea and 278 E. coli strains from healthy children were examined by PCR for 10 virulence genes: enteropathogenic E. coli (EPEC) eae, tir, and bfpA; enterotoxigenic E. coli (ETEC) lt and st; enteroinvasive E. coli (EIEC) ipaH; enterohemorragic E. coli stx1 and stx2; and enteroaggregative E. coli (EAEC) aggR and astA. EAEC strains without AggR were identified by the HEp-2 cell adherence test. The detection of EAEC, ETEC, EPEC, and EIEC was significantly associated with diarrhea. The incidence of EAEC (15.1%), defined by either a molecular or a phenotypic assay, was higher in the diarrheal group than any other category (0 to 6.0%). The incidence of AggR-positive EAEC in the diarrheal group was significantly higher than in the control group (8.0 versus 1.4%; P = 0.0004), while that of AggR-negative EAEC was not (7.1 versus 4.3%). Nineteen AggR-positive EAEC strains harbored other EAEC virulence genes-aggA, 2 (5.5%); aafA, 4 (11.1%); agg-3a, 5 (13.8%); aap, 8 (22.2%); aatA, 11 (30.5%); capU, 9 (25.0%); pet, 6 (16.6%); and set, 3 (8.3%)-and showed 15 genotypes. EAEC may be an important pathogen of sporadic diarrhea in Mongolian children. Genetic analysis showed the heterogeneity of EAEC but illustrated the importance of the AggR regulon (denoting typical EAEC) as a marker for virulent EAEC strains.

Prevention of mildly overweight children from development of more overweight condition.

BACKGROUND: A follow-up study has reported that not only highly obese but also mildly obese children are becoming heavier during the elementary school children. Then we determined the effect of programs for the screening and treatment of overweight elementary school children whether the programs prevented mildly overweight children from development of more overweight condition. METHODS: Subjects were 40 overweight children who participated in both screening and treatment programs. As controls, only 240 children who participated in the screening program were used. The mean observation periods of the subjects and controls were 14 and 12 months, respectively. RESULTS: In both groups, older and heavier children significantly decreased their indices of overweight (percent relative body weight; %RBW), indicating that mildly obese children worsened their %RBW. However, the incidence of children who worsened their %RBW was significantly lower in the subjects (5 of 40) than in the controls (133 of 240) (P < 0.0001). Finally, the subjects significantly reduced their mean %RBW (P < 0.0001) and the controls significantly worsened it (P = 0.0093). CONCLUSION: The treatment program was found important because it prevents mildly overweight children who may worsen their overweight indices without the program from developing more overweight condition.

Who is at risk for cardiac events in young patients with long QT syndrome?

The objective of this study was to determine who is at risk for cardiac events among young patients with long QT syndrome (LQTS) with or without a past history of LQTS-related cardiac events. The subjects were young patients with LQTS who had visited one of 36 hospitals from January 1997 to August 2000 in Japan. To predict the risk factors for cardiac events, stepwise regression analyses were performed for a total of 197 cases. There were 7 of 129 cases (5%) without a past history and 32 of the 68 (47%) cases with a past history of LQTS-related cardiac events that experienced new events after diagnosis (p<0.0001). Patients with a family history showed a higher incidence of symptoms both before and after diagnosis than patients with sporadic occurrence. Analyses revealed that noncompliance with medication and a lower age at diagnosis were significant predictors for the group with a past history. A negative predictive value <4 points was 100% in the group without a past history. To prevent future cardiac events, compliance with medication must be improved in those with a past history. A total LQTS score <4 points was useful to predict the absence of cardiac events in the group without a past history.

Possible relationship between streptococcal pyrogenic exotoxin A and Kawasaki syndrome in patients older than six months of age.

BACKGROUND: We previously investigated antibody titers against four kinds of superantigens [streptococcal pyrogenic exotoxin A (SPEA), streptococcal pyrogenic exotoxin C, toxic shock syndrome toxin-1 and staphylococcal enterotoxin B] in patients with Kawasaki syndrome (KS) younger than 6 months of age and reported a relationship between toxic shock syndrome toxin-1 and KS patients. In this study we have investigated antibody titers in KS patients older than 6 months of age. METHODS: Serum of 81 patients with KS older than 6 months of age, before intravenous gamma-globulin therapy, and 88 normal age-matched children were used in this study. The IgG antibody titers against four kinds of superantigens were measured with an enzyme-linked immunosorbent assay. RESULTS: The KS patients showed significantly elevated mean SPEA titer (P = 0.006) and significantly higher incidence of high SPEA (P = 0.0024) compared with the controls. The SPEA titer in KS patients showed a significant positive correlation with the number of days from onset of illness (P = 0.0002). CONCLUSIONS: The elevated antibody titer against superantigens of KS patients older than 6 months of age was different from that of KS patients younger than 6 months of age. Our results suggest that KS patients' exposure to SPEA occurred a few weeks before the onset of KS. SPEA may be one of the possible etiologic agents of KS among patients older than 6 months of age in Kagoshima, Japan.

Effects of supplemental L-methionine on E-64 [trans-epoxysuccinyl-1-leucyl-amido (4-guanido) butane]-induced dysmorphology in rat embryos cultured in vitro.

E-64 [trans-epoxysuccinyl-1-leucyl-amido (4-guanido) butane] is teratogenic, inducing a spec-trum of malformations in vivo and producing similar effects in vitro. Numerous studies support the concept that E-64-induced malformations result from embryonic nutritionaldeficiency, without affecting the maternal nutritional status. This has provided a useful model with which to investigate the nutritional requirements of the early embryo, as well as the role of various nutrients in the etiology of congenital defects. In the current investigation, we examined effects of L-methionine on E-64-induced embryotoxicity in vitro. For these experiments, we cultured rat embryos 9.5 days postconception (p.c.) for 48 hours with E-64 and/or L-methionine. We found that the addition of L-methionine to E-64-exposed cultures reduced optic abnormality and increased embryo protein. These results suggest that embryopathy largely results from a deficiency of L-methionine although E-64 limits the supply of all amino acids to the embryo. Furthermore, although endocytosis and degradation of proteins by the visceral yolk sac (VYS) supply most amino acids to the embryo, free amino acids may be compensatory when this source is reduced. These results support those of previous investigations that suggest L-methionine is a limiting nutrient for embryonic development.

Oral antimicrobial susceptibilities of Streptococcus pneumoniae, Haemophilus influenzae, and Moraxella catarrhalis isolates from Japanese children.

The minimum inhibitory concentrations (MICs) of 11 oral antibiotics were measured for 140 Streptococcus pneumoniae, 115 Haemophilus influenzae, and 46 Moraxella catarrhalis strains isolated from Japanese children. The antibiotics selected included a range of commonly prescribed agents together with a selection of new cefems and a penem. Cefditoren was most active against the highly penicillin resistant S. pneumoniae, beta-lactamase-producing H. influenzae and beta-lactamase-negative ampicillin-resistant H. influenzae. However, amoxycillin retained good activity against the penicillin-susceptible or -intermediately resistant S. pneumoniae (88.6%) and most of ampicillin-susceptible or -intermediately resistant H. influenzae (87.9%). We thus consider that amoxycillin remains a useful initial choice for the treatment of paediatric respiratory infections in Japan.

Patients diagnosed with Kawasaki disease before the fifth day of illness have a higher risk of coronary artery aneurysm.

BACKGROUND: A fever lasting for at least 5 days is an essential characteristic of the original diagnostic criteria of Kawasaki disease (KD). However, it is not difficult for an experienced physician to confirm the diagnosis of KD before the fifth day of fever. The aim of this study is to investigate the effect of intravenous gamma globulin therapy (IVGG) in KD initiated before the fifth day of illness. METHODS: A total of 125 patients treated with IVGGwere divided into group A (IVGG was initiated before the fifth day of illness, n= 46) and group B (IVGG was initiated at the fifth day or after, n= 79). Patients' characteristics,laboratory findings, treatments and outcomes were compared between the groups. RESULTS: White blood cell count value, C-reactive protein and Harada's score showed no difference between the groups. A significantly higher average value of alanine aminotransferase(ALT) was observed in group A. Although the treatments were identical in both groups, the average duration of fever from the initial day of IVGG in group A was significantly longer than in group B. The incidence of aneurysm in group A was significantly higher than that in group B. Stepwise regression analysis using aneurysm as a dependent variable revealed that group A and ALT were significant. CONCLUSIONS: Patients diagnosed with KD before the fifth day of illness showed a poor response to IVGG. This observation might be related to high ALT values. Further examination concerning the modification of treatment in such patients is necessary.

Incidences of nasopharyngeal colonization of respiratory bacterial pathogens in Japanese children attending day-care centers.

BACKGROUND: In Japan, many younger children attending day-care centers tend to frequently experience acute respiratory infections and prolonged otitis media. OBJECTIVES: To evaluate the carriage rate of respiratory bacterial pathogens in children attending day-care centers in our district. METHODS: Nasopharyngeal cultures of 156 healthy children between the ages of 1 month and 5 years were conducted at two day-care centers in Japan, in April 1999. The carriage rates of four major pathogens (Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catarrhalis and Staphylococcus aureus) and the antibiotic susceptibilities of the isolates were examined. RESULTS: Streptococcus pneumoniae, H. influenzae, M. catarrhalis and S. aureus were detected in 94 (60.3%), 83 (53.2%), 54 (34.6%) and 28 (17.9%) children, respectively. A total of 141 (90.4%) children carried at least one pathogen among these four pathogens and 87 (55.8%) children carried more than one pathogen. Fifty-seven of the 94 (60.6%) S. pneumoniae isolates were penicillin-intermediately or highly resistant strains of S. pneumoniae (PISP/PRSP). Beta-lactamase producing H. influenzae was not detected. Twelve of the 28 (42.9%) S. aureus isolates were methicillin-resistant. The incidence of colonization by PISP/PRSP in children younger than 3 years (43/69, 62.3%) was significantly higher than that in children aged 3-5 years (14/87, 16.1%) (P < 0.0001). CONCLUSIONS: We conclude that the rates of colonization by respiratory bacterial pathogens, especially by antibiotic-resistant strains, were high in children attending day-care centers in our district, suggesting their horizontal spread among children in day-care centers. Considering that the majority of children attending day- care centers carried one or more of the bacterial pathogens, the judicious use of antimicrobials will be required to prevent the increase of antibiotic-resistant rates among the colonizing pathogens.

Diagnostic significance of increased serum hyaluronic acid in juvenile rheumatoid arthritis.

BACKGROUND: We have previously reported that serum levels of hyaluronic acid (HA) objectively reflect the severity of arthritis in juvenile rheumatoid arthritis (JRA). However, clear diagnostic standards do not exist for JRA; it is difficult to evaluate arthritis in children, particularly in small children and the diagnostic criteria for JRA requires an exclusion of several diseases. Therefore, if a specific test finding associated with JRA could be established, it would enable general pediatricians to make an objective diagnosis. METHODS: We measured the serum HA levels in children with joint symptoms as a chief complaint. The total number of subjects were 197 children; of these 89 had JRA, 39 had rheumatic diseases other than JRA, and 69 had non-rheumatic diseases (including systemic 31, polyarticular 40 and pauci-articular in 17), rheumatic diseases other than JRA in 39 subjects, and non-rheumatic diseases in 69 subjects. Sandwich enzyme-linked immunosorbent assay measured HA by using the HA binding protein. RESULTS: The serum level of HA was significantly higher in systemic and polyarticular JRA patients than in patients with pauci-articular JRA, with rheumatic diseases other than JRA, and non-rheumatic patients. With a cut-off value of 100 ng/mL, a diagnostic value of HA in all JRA patients was 48.3% sensitivity and 98.1% specificity. CONCLUSIONS: In children presenting with joint symptoms, serum HA measurement is useful for diagnosing systemic and polyarticular JRA.

Postnatal survival of rat offspring prenatally exposed to pure 2,4-dichlorophenoxyacetic acid (2,4-D).

In previous report on prenatal developmental effects of pure 2,4-dichlorophenoxyacetic acid (2,4-D) in rat, we found that this chemical was maternally toxic, embryolethal, and that it induced urogenital malformations in the fetuses. In the present report, we investigated the postnatal survival of the offspring prenatally exposed to 2,4-D during organogenesis, to determine the participation of urogenital malformations on postnatal survival. We used doses of 70 mg, 110 mg and 150 mg, which were each found to induce significant urogenital malformations, when administered in different periods of organogenesis: GD 6 to 15, GD 6 to 10, and GD 11 to 15. We found that 2,4-D has a significant influence on progeny viability by increasing the postnatal death. The kidney and urinary tract malformations induced in the fetuses might be the cause of the increased rate of postnatal death. 2,4-D did not impair the postnatal growth of the unaffected offspring.

Maternal antibody against toxic shock syndrome toxin-1 may protect infants younger than 6 months of age from developing Kawasaki syndrome.

The symptoms of Kawasaki syndrome (KS) suggest a possible relationship between KS and superantigen(s). The infrequent occurrence of KS among young infants may be due to a passive maternal antibody. We investigated the antibody titers for superantigens (toxic shock syndrome toxin [TSST]-1, staphylococcal exotoxin B, and streptococcal pyrogenic exotoxins C and A) in 15 patients with KS who were <6 months of age prior to gamma globulin therapy and in 10 mothers of patients with KS <6 months of age. Significant findings were observed for only TSST-1 among the 4 anti-superantigens. The proportion of patients with KS who had high anti-TSST-1 titers was significantly higher than that among infant control subjects (33% vs. 5%, respectively; P=.031). The mean anti-TSST-1 titer for the mothers was significantly lower than that of adult control subjects (P=.021). Among infants <6 months of age, TSST-1 may be related to KS, and a maternal antibody may protect infants from developing KS.

Characterization of Klebsiella pneumoniae and Escherichia coli strains that produce CTX-M-2-type broad spectrum beta-lactamase isolated from a child with leukemia.

An 8-year-old girl with acute leukemia had bacteremia caused by Klebsiella pneumoniae producing CTX-M-2-type broad spectrum beta-lactamase. K. pneumoniae and Escherichia coli strains producing the same enzyme and harboring identical conjugative plasmids were recovered from stoor culture. Patients with frequent episodes of neutropenia and prophylactic administration of beta-lactams are at risk of harboring colonizing strains that produce broad spectrum beta-lactamases.

Effect of age and overweight on the QT interval and the prevalence of long QT syndrome in children.

The change in QT interval with age during childhood of normal children and children with long QT syndrome (LQTS) and the effects of body mass index on the QT interval have not been studied in detail. The prevalence of LQTS in children is not well known. We measured 3 consecutive QT and RR intervals in 4,655 children. Their electrocardiograms along with their height and weight were recorded when they were in the first grade in 1994 and again when they were in the seventh grade in 2000. The QT interval was corrected by Bazett's formula. The longer corrected QT intervals in female subjects than male subjects start at elementary school age, earlier than previously reported. Overweight did not have an impact on the uncorrected or corrected QT interval. None of the 4 children diagnosed with LQTS in the seventh grade had characteristic electrocardiographic findings of LQTS in the first grade. All 4 are nonfamilial cases. The prevalence of LQTS in children was found to be 1 of 1,164. These data suggest that abnormal electrocardiographic phenotypes in children with nonfamilial LQTS may appear during the elementary school year. The longer QT intervals in female subjects than male subjects start at the same period. No correlation was found between obesity and length of the QT interval. Finally, the prevalence of LQTS in children is greater than previously suspected.

GCV resistance due to the mutation A594P in the cytomegalovirus protein UL97 is partially reconstituted by a second mutation at D605E.

A ganciclovir (GCV)-resistant human cytomegalovirus (HCMV) was isolated from an AIDS patient. Molecular analysis of the HCMV UL97 gene revealed two point mutations, A594P and D605E, respectively. In order to evaluate quantitatively the impact of the individual mutations on GCV phosphorylation, recombinant vaccinia viruses (rVVs) were generated carrying either the two mutations (rVV-594/605) or only one mutation (rVV-594 or rVV-605, respectively). In cells infected with the rVV-594/605 double mutant, the GCV phosphorylation was decreased to 50% compared with the phosphorylation in cells infected with the rVV-UL97 wild-type. In cells infected with the rVV-594, however, the GCV phosphorylation was further decreased to 30%. Interestingly, the mutation D605E led to an even better GCV phosphorylation than that measured in cells infected with the rVV-UL97 wild type. These results were confirmed by plaque reduction assays, indicating that rVV-594 was more resistant to GCV than rVV-594/605. In contrast, rVV-605 was more sensitive to GCV than the rVV-UL97 wild type. Therefore, our results demonstrated for the first time that compensatory mutations can also occur in HCMV, as already shown for human immunodeficiency virus type 1.

The Upstream Sequence of a New Growth/Differentiation Factor, Midkine (MK), Mediates Developmentally Regulated lac Z Gene Expression in Transgenic Mice.

Midkine (MK) is the product of a retinoic acid responsive gene, and is a heparin binding protein involved in the regulation of growth and differentiation. The 1.9 kb upstream region of MK gene was fused with the bacterial ß-galactosidase gene (lac Z) and injected into fertilized mouse eggs. The resulting transgenic mice were used to evaluate the in vivo transcriptional regulation through of the upstream region. Comparison of the ß-galactosidase expression and endogenous MK expression indicated that the temporal regulation of the transgene was similar to that of MK gene expression during mouse development. The transgene was neither expressed in the preimplantation period nor in 6.5-day embryos. Transgene expression was high and widely distributed on the 8.5th day, became restricted on the 10.5th and 12.5th days, and thereafter almost confined to the kidney. Thus, the 1.9 kb upstream region accounts for overall temporal regulation of MK gene expression, while there are some differences between the spatial regulation of the transgene expression and that of the endogenous MK gene expression. The transgene was expressed in a few limited regions of the brain of 17 day old embryos, and those sites consisted largely of matrix cells with columnar arrangements. These results suggests a role of MK in the brain development, and MK activity may be involved in retinoic acid induced malformations of the central nervous system.